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Medina-Chávez NO, Rodriguez-Cruz UE, Souza V, De la Torre-Zavala S, Travisano M. Salty secrets of Halobacterium salinarum AD88: a new archaeal ecotype isolated from Cuatro Cienegas Basin. BMC Genomics 2025; 26:399. [PMID: 40275130 PMCID: PMC12023398 DOI: 10.1186/s12864-025-11550-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2024] [Accepted: 03/28/2025] [Indexed: 04/26/2025] Open
Abstract
The Cuatro Cienegas Basin (CCB) in Mexico, represents a unique ecological habitat, characterized by extreme and fluctuating conditions, providing a window into ancient evolutionary processes. This basin, characterized by hypersalinity and phosphorus scarcity, harbors diverse microbial communities that exhibit remarkable adaptations to oligotrophic conditions. Among these, Halobacterium salinarum, a halophilic archaeon known for its polyploid genome and metabolic versatility, has been extensively studied as a model for extremophile survival. However, only a limited number of H. salinarum strains have been successfully cultured and characterized to date. Here, we report the isolation and genomic analysis of a novel Halobacterium salinarum strain, AD88, from microbial mats at the Archaean Domes site in the CCB. This strain displays unique genomic features, including smaller plasmid sizes and distinctive metabolic pathways for phosphorus and sulfur utilization. Comparative analyses with other Halobacterium strains revealed genetic innovations, such as genes involved in sulfolipid biosynthesis, enabling membrane stability in phosphorus-depleted environments, and adaptations for horizontal gene transfer, which facilitate genomic flexibility in response to environmental pressures. This study reveals that H. salinarum AD88 is the first recorded diploid strain of Halobacterium, a feature previously undocumented in this genus. Phylogenomic reconstruction positioned AD88 tightly within the Halobacterium clade, reflecting its evolutionary history within the genus. Pangenome analysis further highlighted the open nature of the Halobacterium genus, with AD88 contributing novel accessory genes linked to ecological specialization. These findings emphasize the evolutionary significance of the CCB as a natural laboratory for studying microbial adaptation and expand our understanding of archaeal genomic diversity and functional innovation under extreme conditions.
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Affiliation(s)
- Nahui Olin Medina-Chávez
- Ecology, Evolution and Behavior, University of Minnesota, St. Paul, MN, USA.
- BioTechnology Institute, University of Minnesota, St. Paul, MN, USA.
| | - Ulises E Rodriguez-Cruz
- Departamento de Ecología Evolutiva, Instituto de Ecología, Universidad Nacional Autónoma de México, Ciudad de México, México
- Doctorado en Ciencias Biomédicas, Universidad Nacional Autónoma de México, Ciudad de México, México
| | - Valeria Souza
- Departamento de Ecología Evolutiva, Instituto de Ecología, Universidad Nacional Autónoma de México, Ciudad de México, México
- Centro de Estudios del Cuaternario de Fuego-Patagonia y Antártica (CEQUA), Punta Arenas, Chile
| | - Susana De la Torre-Zavala
- Instituto de Biotecnología, Facultad de Ciencias Biológicas, Universidad Autónoma de Nuevo León, San Nicolas de los Garza, Nuevo León, México
| | - Michael Travisano
- Ecology, Evolution and Behavior, University of Minnesota, St. Paul, MN, USA
- BioTechnology Institute, University of Minnesota, St. Paul, MN, USA
- Minnesota Center for the Philosophy of Science, University of Minnesota, Minneapolis, MN, USA
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2
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Liu W, Wang Z, Huang Y, Liu Y, Li R, Wang M, Zhang H, Meng C, Xiao X. Acetylshikonin reduces the spread of antibiotic resistance via plasmid conjugation. Int J Antimicrob Agents 2024; 64:107370. [PMID: 39481662 DOI: 10.1016/j.ijantimicag.2024.107370] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2024] [Revised: 09/27/2024] [Accepted: 10/22/2024] [Indexed: 11/02/2024]
Abstract
The plasmid-mediated conjugative transfer of antibiotic resistance genes (ARGs) stands out as the primary driver behind the dissemination of antimicrobial resistance (AMR). Developing effective inhibitors that target conjugative transfer represents an potential strategy for addressing the issue of AMR. Here, we studied the effect of acetylshikonin (ASK), a botanical derivative, on plasmid conjugation. The conjugative transfer of RP4-7 plasmid inter and intra species was notably reduced by ASK. The conjugation process of IncI2 and IncX4 plasmids harbouring the mobile colistin resistance gene (mcr-1), IncX4 and IncX3 plasmids containing the carbapenem resistance gene (blaNDM-5), and IncFI and IncFII plasmids possessing the tetracycline resistance gene [tet(X4)] were also reduced by ASK. Importantly, the conjugative transfer frequency of mcr-1 positive IncI2 plasmid in mouse peritoneal conjugation model and gut conjugation model was reduced by ASK. The mechanism investigation showed that ASK disrupted the functionality of the bacterial cell membrane. Furthermore, the proton motive force (PMF) was dissipated. In addition, ASK blocked the electron transmission in bacteria's electron transport chain (ETC) through disturbing the quinone interaction, resulting in an insufficient energy supply for conjugation. Collectively, ASK is a potential conjugative transfer inhibitor, providing novel strategies to prevent the spread of AMR.
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Affiliation(s)
- Wei Liu
- College of Veterinary Medicine, Yangzhou University, Yangzhou, China
| | - Zhiqiang Wang
- College of Veterinary Medicine, Yangzhou University, Yangzhou, China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, China; Jiangsu Key Laboratory of Zoonosis, Yangzhou University, Yangzhou, China; Institute of Comparative Medicine, Yangzhou University, Yangzhou, China
| | - Yanhu Huang
- College of Veterinary Medicine, Yangzhou University, Yangzhou, China
| | - Yuan Liu
- College of Veterinary Medicine, Yangzhou University, Yangzhou, China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, China; Institute of Comparative Medicine, Yangzhou University, Yangzhou, China
| | - Ruichao Li
- College of Veterinary Medicine, Yangzhou University, Yangzhou, China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, China; Institute of Comparative Medicine, Yangzhou University, Yangzhou, China
| | - Mianzhi Wang
- College of Veterinary Medicine, Yangzhou University, Yangzhou, China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, China
| | - Haijie Zhang
- College of Veterinary Medicine, Yangzhou University, Yangzhou, China
| | - Chuang Meng
- Jiangsu Key Laboratory of Zoonosis, Yangzhou University, Yangzhou, China
| | - Xia Xiao
- College of Veterinary Medicine, Yangzhou University, Yangzhou, China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, China; Jiangsu Key Laboratory of Zoonosis, Yangzhou University, Yangzhou, China.
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Burckhardt RM, Escalante-Semerena JC. Sirtuin-dependent reversible lysine acetylation of the o-succinylbenzoyl-coenzyme A synthetase of Bacillus subtilis. Microbiol Spectr 2024; 12:e0201124. [PMID: 39422507 PMCID: PMC11619455 DOI: 10.1128/spectrum.02011-24] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2024] [Accepted: 09/09/2024] [Indexed: 10/19/2024] Open
Abstract
Reversible lysine acylation (RLA) is a conserved posttranslational modification that cells of all domains of life use to regulate the biological function of proteins, some of which have enzymatic activity. Many AMP-forming organic acid:CoA ligases are regulated via acylation in prokaryotes and eukaryotes. Here, we report the acetylation of the o-succinylbenzoyl-CoA synthetase (EC 6.2.1.26) of Bacillus subtilis (BsMenE) by the GCN5-related acetyltransferase (GNAT) AcuA enzyme of this bacterium. BsMenE is part of the metabolic pathway that assembles menaquinone (MK), an essential component of the electron transport chain in B. subtilis. We demonstrate that the active-site lysine 471 (K471) of BsMenE is acetylated specifically by BsAcuA, and that acetylated BsMenE (BsMenEAc) is deacetylated by the NAD+-dependent sirtuin (BsSrtN) of this bacterium. The in vivo analyses performed in this study were done in an Escherichia coli ΔmenE strain because the enzymatic activity of MenE is essential in B. subtilis, but not in E. coli. The use of a heterologous system allowed us to assess the effect of acetylation on BsMenE function under MK-dependent growth conditions. Based on our in vivo data, we suggest that regulation of BsMenE by RLA reduces MK production, negatively affecting the growth rate and yield of the culture.IMPORTANCEReversible lysine acylation (RLA) is a posttranslational modification used by all cells to rapidly control the biological function of proteins. Herein, we identify an acetyltransferase and deacetylase in the soil bacterium Bacillus subtilis that can modify/demodify an enzyme required for the synthesis of menaquinone (MK), an essential electron carrier involved in respiration in cells of all domains of life. Based on our data, we suggest that under some as-yet-undefined physiological conditions, B. subtilis modulates MK biosynthesis, which changes the flux of electrons through the electron transport chain of this bacterium. To our knowledge, this is the first example of control of respiration by RLA.
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Bargabos R, Iinishi A, Hawkins B, Privalsky T, Pitt N, Son S, Corsetti R, Gates MF, Miller RD, Lewis K. Small molecule produced by Photorhabdus interferes with ubiquinone biosynthesis in Gram-negative bacteria. mBio 2024; 15:e0116724. [PMID: 39254306 PMCID: PMC11481567 DOI: 10.1128/mbio.01167-24] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2024] [Accepted: 06/26/2024] [Indexed: 09/11/2024] Open
Abstract
We report the identification of 3,6-dihydroxy-1,2-benzisoxazole (DHB) in a screen of Photorhabdus and Xenorhabdus, whose symbiotic relationship with eukaryotic nematodes favors secondary metabolites that meet several requirements matching those for clinically useful antibiotics. DHB is produced by Photorhabdus laumondii and is selective against the Gram-negative species Escherichia coli, Enterobacter cloacae, Serratia marcescens, Klebsiella pneumoniae, Proteus mirabilis, and Acinetobacter baumannii. It is inactive against anaerobic gut bacteria and nontoxic to human cells. Mutants resistant to DHB map to the ubiquinone biosynthesis pathway. DHB binds to 4-hydroxybenzoate octaprenyltransferase (UbiA) and prevents the formation of 4-hydroxy-3-octaprenylbenzoate. Remarkably, DHB itself is prenylated, forming an unusable chimeric product that likely contributes to the toxic effect of this antimicrobial. DHB appears to be both a competitive enzyme inhibitor and a prodrug; this dual mode of action is unusual for an antimicrobial compound. IMPORTANCE The spread of resistant pathogens has led to the antimicrobial resistance crisis, and the need for new compounds acting against Gram-negative pathogens is especially acute. From a screen of Photorhabdus symbionts of nematodes, we identified 3,6-dihydroxy-1,2-benzisoxazole (DHB) that acts against a range of Gram-negative bacteria, including Escherichia coli, Enterobacter cloacae, Klebsiella pneumoniae, and Acinetobacter baumannii. DHB had previously been isolated from other bacterial species, but its mechanism of action remained unknown. We show that DHB is unique among antimicrobials, with dual action as an inhibitor of an important enzyme, UbiA, in the biosynthesis pathway of ubiquinone and as a prodrug. DHB is a mimic of the natural substrate, and UbiA modifies it into a toxic product, contributing to the antimicrobial action of this unusual antibiotic. We also uncover the mechanism of DHB selectivity, which depends on a particular fold of the UbiA enzyme.
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Affiliation(s)
- Rachel Bargabos
- Antimicrobial Discovery Center, Northeastern University, Boston, Massachusetts, USA
| | - Akira Iinishi
- Antimicrobial Discovery Center, Northeastern University, Boston, Massachusetts, USA
| | - Bryson Hawkins
- Antimicrobial Discovery Center, Northeastern University, Boston, Massachusetts, USA
| | - Thomas Privalsky
- Antimicrobial Discovery Center, Northeastern University, Boston, Massachusetts, USA
| | - Norman Pitt
- Antimicrobial Discovery Center, Northeastern University, Boston, Massachusetts, USA
| | - Sangkeun Son
- Antimicrobial Discovery Center, Northeastern University, Boston, Massachusetts, USA
| | - Rachel Corsetti
- Antimicrobial Discovery Center, Northeastern University, Boston, Massachusetts, USA
| | - Michael F. Gates
- Antimicrobial Discovery Center, Northeastern University, Boston, Massachusetts, USA
| | - Ryan D. Miller
- Antimicrobial Discovery Center, Northeastern University, Boston, Massachusetts, USA
| | - Kim Lewis
- Antimicrobial Discovery Center, Northeastern University, Boston, Massachusetts, USA
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Wang Y, Lilienfeldt N, Hekimi S. Understanding coenzyme Q. Physiol Rev 2024; 104:1533-1610. [PMID: 38722242 PMCID: PMC11495197 DOI: 10.1152/physrev.00040.2023] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2023] [Revised: 04/08/2024] [Accepted: 05/01/2024] [Indexed: 08/11/2024] Open
Abstract
Coenzyme Q (CoQ), also known as ubiquinone, comprises a benzoquinone head group and a long isoprenoid side chain. It is thus extremely hydrophobic and resides in membranes. It is best known for its complex function as an electron transporter in the mitochondrial electron transport chain (ETC) but is also required for several other crucial cellular processes. In fact, CoQ appears to be central to the entire redox balance of the cell. Remarkably, its structure and therefore its properties have not changed from bacteria to vertebrates. In metazoans, it is synthesized in all cells and is found in most, and maybe all, biological membranes. CoQ is also known as a nutritional supplement, mostly because of its involvement with antioxidant defenses. However, whether there is any health benefit from oral consumption of CoQ is not well established. Here we review the function of CoQ as a redox-active molecule in the ETC and other enzymatic systems, its role as a prooxidant in reactive oxygen species generation, and its separate involvement in antioxidant mechanisms. We also review CoQ biosynthesis, which is particularly complex because of its extreme hydrophobicity, as well as the biological consequences of primary and secondary CoQ deficiency, including in human patients. Primary CoQ deficiency is a rare inborn condition due to mutation in CoQ biosynthetic genes. Secondary CoQ deficiency is much more common, as it accompanies a variety of pathological conditions, including mitochondrial disorders as well as aging. In this context, we discuss the importance, but also the great difficulty, of alleviating CoQ deficiency by CoQ supplementation.
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Affiliation(s)
- Ying Wang
- Department of Biology, McGill University, Montreal, Quebec, Canada
| | - Noah Lilienfeldt
- Department of Biology, McGill University, Montreal, Quebec, Canada
| | - Siegfried Hekimi
- Department of Biology, McGill University, Montreal, Quebec, Canada
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6
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Mondal A, Roy P, Carrannanto J, Datar PM, DiRocco DJ, Hunter K, Marsh ENG. Surveying the scope of aromatic decarboxylations catalyzed by prenylated-flavin dependent enzymes. Faraday Discuss 2024; 252:208-222. [PMID: 38837123 DOI: 10.1039/d4fd00006d] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/06/2024]
Abstract
The prenylated-flavin mononucleotide-dependent decarboxylases (also known as UbiD-like enzymes) are the most recently discovered family of decarboxylases. The modified flavin facilitates the decarboxylation of unsaturated carboxylic acids through a novel mechanism involving 1,3-dipolar cyclo-addition chemistry. UbiD-like enzymes have attracted considerable interest for biocatalysis applications due to their ability to catalyse (de)carboxylation reactions on a broad range of aromatic substrates at otherwise unreactive carbon centres. There are now ∼35 000 protein sequences annotated as hypothetical UbiD-like enzymes. Sequence similarity network analyses of the UbiD protein family suggests that there are likely dozens of distinct decarboxylase enzymes represented within this family. Furthermore, many of the enzymes so far characterized can decarboxylate a broad range of substrates. Here we describe a strategy to identify potential substrates of UbiD-like enzymes based on detecting enzyme-catalysed solvent deuterium exchange into potential substrates. Using ferulic acid decarboxylase (FDC) as a model system, we tested a diverse range of aromatic and heterocyclic molecules for their ability to undergo enzyme-catalysed H/D exchange in deuterated buffer. We found that FDC catalyses H/D exchange, albeit at generally very low levels, into a wide range of small, aromatic molecules that have little resemblance to its physiological substrate. In contrast, the sub-set of aromatic carboxylic acids that are substrates for FDC-catalysed decarboxylation is much smaller. We discuss the implications of these findings for screening uncharacterized UbiD-like enzymes for novel (de)carboxylase activity.
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Affiliation(s)
- Anushree Mondal
- Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA.
| | - Pronay Roy
- Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA.
| | - Jaclyn Carrannanto
- Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA.
| | - Prathamesh M Datar
- Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA.
| | - Daniel J DiRocco
- Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA.
| | - Katherine Hunter
- Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA.
| | - E Neil G Marsh
- Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA.
- Department of Biological Chemistry, University of Michigan, Ann Arbor, Michigan, USA
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7
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Thitirungreangchai T, Roytrakul S, Aunpad R. Deciphering the Intracellular Action of the Antimicrobial Peptide A11 via an In-Depth Analysis of Its Effect on the Global Proteome of Acinetobacter baumannii. ACS Infect Dis 2024; 10:2795-2813. [PMID: 39075773 PMCID: PMC11320580 DOI: 10.1021/acsinfecdis.4c00160] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2024] [Revised: 07/17/2024] [Accepted: 07/17/2024] [Indexed: 07/31/2024]
Abstract
The potential antimicrobial activity and low propensity to induce the development of bacterial resistance have rendered antimicrobial peptides (AMPs) as novel and ideal candidate therapeutic agents for the treatment of infections caused by drug-resistant pathogenic bacteria. The targeting of bacterial membranes by AMPs has been typically considered their sole mode of action; however, increasing evidence supports the existence of multiple and complementary functions of AMPs that result in bacterial death. An in-depth characterization of their mechanism of action could facilitate further research and development of AMPs with higher potency. The current study employs biophysics and proteomics approaches to unveil the mechanisms underlying the antibacterial activity of A11, a potential candidate AMP, against Acinetobacter baumannii, a leading cause of hospital-acquired infections (HAIs) and consequently, a serious global threat. A11 peptide was found to induce membrane depolarization to a high extent, as revealed by flow cytometry and electron microscopy analyses. The prompt intracellular penetration of A11 peptide, observed using confocal microscopy, was found to occur concomitantly with a very low degree of membrane lysis, suggesting that its mode of action predominantly involves a nonlytic killing mechanism. Quantitative proteomics analysis employed for obtaining insights into the mechanisms underlying the antimicrobial activity of A11 peptide revealed that it disrupted energy metabolism, interfered with protein homeostasis, and inhibited fatty acid synthesis that is essential for cell membrane integrity; all these impacted the cellular functions of A. baumannii. A11 treatment also impacted signal transduction associated with the regulation of biofilm formation, hindered the stress response, and influenced DNA repair processes; these are all crucial survival mechanisms of A. baumannii. Additionally, robust antibacterial activity was exhibited by A11 peptide against multidrug-resistant (MDR) and extensively drug-resistant (XDR) clinical isolates of A. baumannii; moreover, A11 peptide exhibited synergy with levofloxacin and minocycline as well as low propensity for inducing resistance. Taken together, the findings emphasize the therapeutic potential of A11 peptide as an antibacterial agent against drug-resistant A. baumannii and underscore the need for further investigation.
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Affiliation(s)
- Thanit Thitirungreangchai
- Graduate
Program in Biomedical Sciences, Faculty of Allied Health Sciences, Thammasat University, Khlong Luang, Pathum Thani 12120, Thailand
| | - Sittiruk Roytrakul
- Functional
Proteomics Technology Laboratory, National Center for Genetic Engineering
and Biotechnology, National Science and
Technology Development Agency, Khlong Luang, Pathum Thani 12120, Thailand
| | - Ratchaneewan Aunpad
- Graduate
Program in Biomedical Sciences, Faculty of Allied Health Sciences, Thammasat University, Khlong Luang, Pathum Thani 12120, Thailand
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8
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Miranda S, Koop M, Angeli A, Lagrèze J, Malnoy M, Martens S. Assessment and Partial Characterization of Candidate Genes in Dihydrochalcone and Arbutin Biosynthesis in an Apple-Pear Hybrid by De Novo Transcriptome Assembly. JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY 2024; 72:11804-11819. [PMID: 38717061 DOI: 10.1021/acs.jafc.4c01006] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/23/2024]
Abstract
Apples (Malus × domestica Borkh.) and pears (Pyrus communis L.) are valuable crops closely related within the Rosaceae family with reported nutraceutical properties derived from secondary metabolites including phloridzin and arbutin, which are distinctive phenolic metabolites characterizing apples and pears, respectively. Here, we generated a de novo transcriptome assembly of an intergeneric hybrid between apple and pear, accumulating intermediate levels of phloridzin and arbutin. Combining RNA-seq, in silico functional annotation prediction, targeted gene expression analysis, and expression-metabolite correlations, we identified candidate genes for functional characterization, resulting in the identification of active arbutin synthases in the hybrid and parental genotypes. Despite exhibiting an active arbutin synthase in vitro, the natural lack of arbutin in apples is reasoned by the absence of the substrate and broad substrate specificity. Altogether, our study serves as the basis for future assessment of potential physiological roles of identified genes by genome editing of hybrids and pears.
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Affiliation(s)
- Simón Miranda
- Research and Innovation Centre, Edmund Mach Foundation, San Michele all'Adige 38098, Italy
| | - Marion Koop
- Research and Innovation Centre, Edmund Mach Foundation, San Michele all'Adige 38098, Italy
| | - Andrea Angeli
- Research and Innovation Centre, Edmund Mach Foundation, San Michele all'Adige 38098, Italy
| | - Jorge Lagrèze
- Research and Innovation Centre, Edmund Mach Foundation, San Michele all'Adige 38098, Italy
| | - Mickael Malnoy
- Research and Innovation Centre, Edmund Mach Foundation, San Michele all'Adige 38098, Italy
| | - Stefan Martens
- Research and Innovation Centre, Edmund Mach Foundation, San Michele all'Adige 38098, Italy
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9
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Tang J, Matsuda Y. Discovery of fungal onoceroid triterpenoids through domainless enzyme-targeted global genome mining. Nat Commun 2024; 15:4312. [PMID: 38773118 PMCID: PMC11109268 DOI: 10.1038/s41467-024-48771-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2023] [Accepted: 05/09/2024] [Indexed: 05/23/2024] Open
Abstract
Genomics-guided methodologies have revolutionized the discovery of natural products. However, a major challenge in the field of genome mining is determining how to selectively extract biosynthetic gene clusters (BGCs) for untapped natural products from numerous available genome sequences. In this study, we developed a fungal genome mining tool that extracts BGCs encoding enzymes that lack a detectable protein domain (i.e., domainless enzymes) and are not recognized as biosynthetic proteins by existing bioinformatic tools. We searched for BGCs encoding a homologue of Pyr4-family terpene cyclases, which are representative examples of apparently domainless enzymes, in approximately 2000 fungal genomes and discovered several BGCs with unique features. The subsequent characterization of selected BGCs led to the discovery of fungal onoceroid triterpenoids and unprecedented onoceroid synthases. Furthermore, in addition to the onoceroids, a previously unreported sesquiterpene hydroquinone, of which the biosynthesis involves a Pyr4-family terpene cyclase, was obtained. Our genome mining tool has broad applicability in fungal genome mining and can serve as a beneficial platform for accessing diverse, unexploited natural products.
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Affiliation(s)
- Jia Tang
- Department of Chemistry, City University of Hong Kong, Tat Chee Avenue, Kowloon, Hong Kong SAR, China
| | - Yudai Matsuda
- Department of Chemistry, City University of Hong Kong, Tat Chee Avenue, Kowloon, Hong Kong SAR, China.
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10
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Karlo J, Gupta A, Singh SP. In situ monitoring of the shikimate pathway: a combinatorial approach of Raman reverse stable isotope probing and hyperspectral imaging. Analyst 2024; 149:2833-2841. [PMID: 38587502 DOI: 10.1039/d4an00203b] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 04/09/2024]
Abstract
Sensing and visualization of metabolites and metabolic pathways in situ are significant requirements for tracking their spatiotemporal dynamics in a non-destructive manner. The shikimate pathway is an important cellular mechanism that leads to the de novo synthesis of many compounds containing aromatic rings of high importance such as phenylalanine, tyrosine, and tryptophan. In this work, we present a cost-effective and extraction-free method based on the principles of stable isotope-coupled Raman spectroscopy and hyperspectral Raman imaging to monitor and visualize the activity of the shikimate pathway. We also demonstrated the applicability of this approach for nascent aromatic amino acid localization and tracking turnover dynamics in both prokaryotic and eukaryotic model systems. This method can emerge as a promising tool for both qualitative and semi-quantitative in situ metabolomics, contributing to a better understanding of aromatic ring-containing metabolite dynamics across various organisms.
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Affiliation(s)
- Jiro Karlo
- Department of Biosciences and Bioengineering, Indian Institute of Technology Dharwad, Dharwad, Karnataka, 580011, India.
| | - Aryan Gupta
- Department of Biosciences and Bioengineering, Indian Institute of Technology Dharwad, Dharwad, Karnataka, 580011, India.
| | - Surya Pratap Singh
- Department of Biosciences and Bioengineering, Indian Institute of Technology Dharwad, Dharwad, Karnataka, 580011, India.
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11
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Gheorghe CE, Leigh SJ, Tofani GSS, Bastiaanssen TFS, Lyte JM, Gardellin E, Govindan A, Strain C, Martinez-Herrero S, Goodson MS, Kelley-Loughnane N, Cryan JF, Clarke G. The microbiota drives diurnal rhythms in tryptophan metabolism in the stressed gut. Cell Rep 2024; 43:114079. [PMID: 38613781 DOI: 10.1016/j.celrep.2024.114079] [Citation(s) in RCA: 10] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2023] [Revised: 02/09/2024] [Accepted: 03/22/2024] [Indexed: 04/15/2024] Open
Abstract
Chronic stress disrupts microbiota-gut-brain axis function and is associated with altered tryptophan metabolism, impaired gut barrier function, and disrupted diurnal rhythms. However, little is known about the effects of acute stress on the gut and how it is influenced by diurnal physiology. Here, we used germ-free and antibiotic-depleted mice to understand how microbiota-dependent oscillations in tryptophan metabolism would alter gut barrier function at baseline and in response to an acute stressor. Cecal metabolomics identified tryptophan metabolism as most responsive to a 15-min acute stressor, while shotgun metagenomics revealed that most bacterial species exhibiting rhythmicity metabolize tryptophan. Our findings highlight that the gastrointestinal response to acute stress is dependent on the time of day and the microbiome, with a signature of stress-induced functional alterations in the ileum and altered tryptophan metabolism in the colon.
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Affiliation(s)
- Cassandra E Gheorghe
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Department of Psychiatry and Neurobehavioural Science, University College Cork, T12 CY82 Cork, Ireland; Department of Anatomy and Neuroscience, University College Cork, T12 CY82 Cork, Ireland
| | - Sarah-Jane Leigh
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Department of Psychiatry and Neurobehavioural Science, University College Cork, T12 CY82 Cork, Ireland; Department of Anatomy and Neuroscience, University College Cork, T12 CY82 Cork, Ireland
| | - Gabriel S S Tofani
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Department of Anatomy and Neuroscience, University College Cork, T12 CY82 Cork, Ireland
| | - Thomaz F S Bastiaanssen
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Department of Anatomy and Neuroscience, University College Cork, T12 CY82 Cork, Ireland
| | - Joshua M Lyte
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Department of Psychiatry and Neurobehavioural Science, University College Cork, T12 CY82 Cork, Ireland; Department of Anatomy and Neuroscience, University College Cork, T12 CY82 Cork, Ireland
| | - Elisa Gardellin
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Department of Psychiatry and Neurobehavioural Science, University College Cork, T12 CY82 Cork, Ireland
| | - Ashokkumar Govindan
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Teagasc Moorepark Food Research Centre, Fermoy Co, P61 C996 Cork, Ireland
| | - Conall Strain
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Teagasc Moorepark Food Research Centre, Fermoy Co, P61 C996 Cork, Ireland
| | - Sonia Martinez-Herrero
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Department of Psychiatry and Neurobehavioural Science, University College Cork, T12 CY82 Cork, Ireland; Department of Anatomy and Neuroscience, University College Cork, T12 CY82 Cork, Ireland
| | - Michael S Goodson
- 711th Human Performance Wing, Air Force Research Laboratory, Wright-Patterson Air Force Base, Dayton, OH 45324, USA
| | - Nancy Kelley-Loughnane
- 711th Human Performance Wing, Air Force Research Laboratory, Wright-Patterson Air Force Base, Dayton, OH 45324, USA
| | - John F Cryan
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Department of Anatomy and Neuroscience, University College Cork, T12 CY82 Cork, Ireland
| | - Gerard Clarke
- APC Microbiome Ireland, University College Cork, T12 CY82 Cork, Ireland; Department of Psychiatry and Neurobehavioural Science, University College Cork, T12 CY82 Cork, Ireland.
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Kuper TJ, Islam MM, Peirce-Cottler SM, Papin JA, Ford RM. Spatial transcriptome-guided multi-scale framework connects P. aeruginosa metabolic states to oxidative stress biofilm microenvironment. PLoS Comput Biol 2024; 20:e1012031. [PMID: 38669236 PMCID: PMC11051585 DOI: 10.1371/journal.pcbi.1012031] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/12/2024] [Accepted: 03/29/2024] [Indexed: 04/28/2024] Open
Abstract
With the generation of spatially resolved transcriptomics of microbial biofilms, computational tools can be used to integrate this data to elucidate the multi-scale mechanisms controlling heterogeneous biofilm metabolism. This work presents a Multi-scale model of Metabolism In Cellular Systems (MiMICS) which is a computational framework that couples a genome-scale metabolic network reconstruction (GENRE) with Hybrid Automata Library (HAL), an existing agent-based model and reaction-diffusion model platform. A key feature of MiMICS is the ability to incorporate multiple -omics-guided metabolic models, which can represent unique metabolic states that yield different metabolic parameter values passed to the extracellular models. We used MiMICS to simulate Pseudomonas aeruginosa regulation of denitrification and oxidative stress metabolism in hypoxic and nitric oxide (NO) biofilm microenvironments. Integration of P. aeruginosa PA14 biofilm spatial transcriptomic data into a P. aeruginosa PA14 GENRE generated four PA14 metabolic model states that were input into MiMICS. Characteristic of aerobic, denitrification, and oxidative stress metabolism, the four metabolic model states predicted different oxygen, nitrate, and NO exchange fluxes that were passed as inputs to update the agent's local metabolite concentrations in the extracellular reaction-diffusion model. Individual bacterial agents chose a PA14 metabolic model state based on a combination of stochastic rules, and agents sensing local oxygen and NO. Transcriptome-guided MiMICS predictions suggested microscale denitrification and oxidative stress metabolic heterogeneity emerged due to local variability in the NO biofilm microenvironment. MiMICS accurately predicted the biofilm's spatial relationships between denitrification, oxidative stress, and central carbon metabolism. As simulated cells responded to extracellular NO, MiMICS revealed dynamics of cell populations heterogeneously upregulating reactions in the denitrification pathway, which may function to maintain NO levels within non-toxic ranges. We demonstrated that MiMICS is a valuable computational tool to incorporate multiple -omics-guided metabolic models to mechanistically map heterogeneous microbial metabolic states to the biofilm microenvironment.
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Affiliation(s)
- Tracy J. Kuper
- Department of Chemical Engineering, University of Virginia, Charlottesville, Virginia, United States of America
| | - Mohammad Mazharul Islam
- Department of Biomedical Engineering, University of Virginia, Charlottesville, Virginia, United States of America
| | - Shayn M. Peirce-Cottler
- Department of Biomedical Engineering, University of Virginia, Charlottesville, Virginia, United States of America
| | - Jason A. Papin
- Department of Biomedical Engineering, University of Virginia, Charlottesville, Virginia, United States of America
| | - Roseanne M Ford
- Department of Chemical Engineering, University of Virginia, Charlottesville, Virginia, United States of America
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Datar PM, Joshi SY, Deshmukh SA, Marsh ENG. Probing the role of protein conformational changes in the mechanism of prenylated-FMN-dependent phenazine-1-carboxylic acid decarboxylase. J Biol Chem 2024; 300:105621. [PMID: 38176649 PMCID: PMC10850782 DOI: 10.1016/j.jbc.2023.105621] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/21/2023] [Revised: 11/30/2023] [Accepted: 12/27/2023] [Indexed: 01/06/2024] Open
Abstract
Phenazine-1-carboxylic acid decarboxylase (PhdA) is a prenylated-FMN-dependent (prFMN) enzyme belonging to the UbiD family of decarboxylases. Many UbiD-like enzymes catalyze (de)carboxylation reactions on aromatic rings and conjugated double bonds and are potentially valuable industrial catalysts. We have investigated the mechanism of PhdA using a slow turnover substrate, 2,3-dimethylquinoxaline-5-carboxylic acid (DQCA). Detailed analysis of the pH dependence and solvent deuterium isotope effects associated with the reaction uncovered unusual kinetic behavior. At low substrate concentrations, a substantial inverse solvent isotope effect (SIE) is observed on Vmax/KM of ∼ 0.5 when reaction rates of DQCA in H2O and D2O are compared. Under the same conditions, a normal SIE of 4.15 is measured by internal competition for proton transfer to the product. These apparently contradictory results indicate that the SIE values report on different steps in the mechanism. A proton inventory analysis of the reaction under Vmax/KM and Vmax conditions points to a "medium effect" as the source of the inverse SIE. Molecular dynamics simulations of the effect of D2O on PhdA structure support that D2O reduces the conformational lability of the enzyme and results in a more compact structure, akin to the active, "closed" conformer observed in crystal structures of some UbiD-like enzymes. Consistent with the simulations, PhdA was found to be more stable in D2O and to bind DQCA more tightly, leading to the observed rate enhancement under Vmax/KM conditions.
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Affiliation(s)
- Prathamesh M Datar
- Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA
| | - Soumil Y Joshi
- Department of Chemical Engineering, Virginia Polytechnic Institute and State University, Blacksburg, Virginia, USA
| | - Sanket A Deshmukh
- Department of Chemical Engineering, Virginia Polytechnic Institute and State University, Blacksburg, Virginia, USA
| | - E Neil G Marsh
- Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA; Department of Biological Chemistry, University of Michigan, Ann Arbor, Michigan, USA.
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Lyu Y, Xu J, Verdoodt F, Vanhaecke L, Hemeryck LY, Hesta M. Faecal metabolome responses to an altered dietary protein:carbohydrate ratio in adult dogs. Vet Q 2023; 43:1-10. [PMID: 37869782 PMCID: PMC10614716 DOI: 10.1080/01652176.2023.2273891] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2023] [Accepted: 10/17/2023] [Indexed: 10/24/2023] Open
Abstract
High-protein diets may aid weight loss and weight maintenance programs in both humans and dogs, although the effect of dietary protein levels on gut metabolism and functionality has not been studied in depth. The current study aimed to investigate the effect of an altered dietary protein:carbohydrate ratio on gut function in adult dogs by means of faecal metabolomic fingerprinting. More specifically, functional metabolic differences in dogs fed a high-protein/low-carbohydrate (HPLC) vs. low-protein/high-carbohydrate (LPHC) diet were studied by equally allocating twelve clinically healthy (6 lean and 6 obese) Beagles into two groups in a cross-over design, with each group receiving two isocaloric diets for four weeks. The faecal metabolome revealed that different protein:carbohydrate ratio can influence host and/or gut microbiome metabolism and function, while no effect was observed on the body condition. Targeted analysis demonstrated that the HPLC diet significantly increased the concentration of indole, spermidine, and pipecolinic acid and decreased the concentration of azelaic acid, D-fructose, mannose, and galactose (p < 0.05). Multivariate modelling (OPLS-DA) of the untargeted faecal metabolome revealed distinctly different metabolomic profiles following the HPLC vs. LPHC diet, with 18 altered pathways. The HPLC diet influenced amino acid and lipid metabolism, potentially promoting weight loss and immune function, whereas the LPHC diet affected carbohydrate fermentation and may promote anti-oxidative function.
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Affiliation(s)
- Yang Lyu
- ECAN Equine and Companion Animal Nutrition, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
| | - Jia Xu
- ECAN Equine and Companion Animal Nutrition, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
| | - Fien Verdoodt
- ECAN Equine and Companion Animal Nutrition, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
| | - Lynn Vanhaecke
- Laboratory of Integrative Metabolomics, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
| | - Lieselot Y. Hemeryck
- Laboratory of Integrative Metabolomics, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
| | - Myriam Hesta
- ECAN Equine and Companion Animal Nutrition, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
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Back D, O’Donnell TJ, Axt KK, Gurr JR, Vanegas JM, Williams PG, Philmus B. Identification, Heterologous Expression, and Characterization of the Tolypodiol Biosynthetic Gene Cluster through an Integrated Approach. ACS Chem Biol 2023; 18:1797-1807. [PMID: 37487226 PMCID: PMC10529828 DOI: 10.1021/acschembio.3c00225] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 07/26/2023]
Abstract
Cyanobacteria are tremendous producers of biologically active natural products, including the potent anti-inflammatory compound tolypodiol. However, linking biosynthetic gene clusters with compound production in cyanobacteria has lagged behind that in other bacterial genera. Tolypodiol is a meroterpenoid originally isolated from the cyanobacterium HT-58-2. Here we describe the identification of the tolypodiol biosynthetic gene cluster through heterologous expression in Anabaena and in vitro protein assays of a methyltransferase found in the tolypodiol biosynthetic gene cluster. We have also identified similar biosynthetic gene clusters in cyanobacterial and actinobacterial genomes, suggesting that meroterpenoids with structural similarity to the tolypodiols may be synthesized by other microbes. We also report the identification of two new analogs of tolypodiol that we have identified in both the original and heterologous producer. This work further illustrates the usefulness of Anabaena as a heterologous expression host for cyanobacterial compounds and how integrated approaches can help to link natural product compounds with their producing biosynthetic gene clusters.
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Affiliation(s)
- Daniel Back
- Department of Pharmaceutical Sciences, Oregon State University, Corvallis, OR 97331, U.S.A
| | - Timothy J. O’Donnell
- Department of Chemistry, University of Hawai’i at Mānoa, Honolulu, HI 96822, U.S.A
| | - Kyle K. Axt
- Department of Pharmaceutical Sciences, Oregon State University, Corvallis, OR 97331, U.S.A
| | - Joshua R. Gurr
- Department of Chemistry, University of Hawai’i at Mānoa, Honolulu, HI 96822, U.S.A
| | - Juan M. Vanegas
- Department of Biochemistry and Biophysics, Oregon State University, Corvallis, OR 97331, U.S.A
| | - Philip G. Williams
- Department of Chemistry, University of Hawai’i at Mānoa, Honolulu, HI 96822, U.S.A
| | - Benjamin Philmus
- Department of Pharmaceutical Sciences, Oregon State University, Corvallis, OR 97331, U.S.A
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Liebl M, Huber L, Elsaman H, Merschak P, Wagener J, Gsaller F, Müller C. Quantifying Isoprenoids in the Ergosterol Biosynthesis by Gas Chromatography-Mass Spectrometry. J Fungi (Basel) 2023; 9:768. [PMID: 37504756 PMCID: PMC10381423 DOI: 10.3390/jof9070768] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/30/2023] [Revised: 07/14/2023] [Accepted: 07/19/2023] [Indexed: 07/29/2023] Open
Abstract
The ergosterol pathway is a promising target for the development of new antifungals since its enzymes are essential for fungal cell growth. Appropriate screening assays are therefore needed that allow the identification of potential inhibitors. We developed a whole-cell screening method, which can be used to identify compounds interacting with the enzymes of isoprenoid biosynthesis, an important part of the ergosterol biosynthesis pathway. The method was validated according to the EMEA guideline on bioanalytical method validation. Aspergillus fumigatus hyphae and Saccharomyces cerevisiae cells were lysed mechanically in an aqueous buffer optimized for the enzymatic deconjugation of isoprenoid pyrophosphates. The residual alcohols were extracted, silylated and analyzed by GC-MS. The obtained isoprenoid pattern provides an indication of the inhibited enzyme, due to the accumulation of specific substrates. By analyzing terbinafine-treated A. fumigatus and mutant strains containing tunable gene copies of erg9 or erg1, respectively, the method was verified. Downregulation of erg9 resulted in a high accumulation of intracellular farnesol as well as elevated levels of geranylgeraniol and isoprenol. The decreased expression of erg1 as well as terbinafine treatment led to an increased squalene content. Additional analysis of growth medium revealed high farnesyl pyrophosphate levels extruded during erg9 downregulation.
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Affiliation(s)
- Maximilian Liebl
- Department of Pharmacy, Center for Drug Research, Ludwig-Maximilians University, 81377 Munich, Germany; (M.L.); (L.H.)
| | - Ludwig Huber
- Department of Pharmacy, Center for Drug Research, Ludwig-Maximilians University, 81377 Munich, Germany; (M.L.); (L.H.)
| | - Hesham Elsaman
- Institute for Hygiene and Microbiology, Julius-Maximilians-University Wuerzburg, 97080 Wuerzburg, Germany; (H.E.); (J.W.)
| | - Petra Merschak
- Institute of Molecular Biology, Biocenter, Medical University of Innsbruck, 6020 Innsbruck, Austria; (P.M.); (F.G.)
| | - Johannes Wagener
- Institute for Hygiene and Microbiology, Julius-Maximilians-University Wuerzburg, 97080 Wuerzburg, Germany; (H.E.); (J.W.)
- Department of Clinical Microbiology, School of Medicine, Trinity College Dublin, The University of Dublin, D08 RX0X Dublin, Ireland
| | - Fabio Gsaller
- Institute of Molecular Biology, Biocenter, Medical University of Innsbruck, 6020 Innsbruck, Austria; (P.M.); (F.G.)
| | - Christoph Müller
- Department of Pharmacy, Center for Drug Research, Ludwig-Maximilians University, 81377 Munich, Germany; (M.L.); (L.H.)
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17
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Golparian D, Jacobsson S, Holley CL, Shafer WM, Unemo M. High-level in vitro resistance to gentamicin acquired in a stepwise manner in Neisseria gonorrhoeae. J Antimicrob Chemother 2023; 78:1769-1778. [PMID: 37253051 PMCID: PMC10517096 DOI: 10.1093/jac/dkad168] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2023] [Accepted: 05/10/2023] [Indexed: 06/01/2023] Open
Abstract
OBJECTIVES Gentamicin is used in several alternative treatments for gonorrhoea. Verified clinical Neisseria gonorrhoeae isolates with gentamicin resistance are mainly lacking and understanding the mechanisms for gonococcal gentamicin resistance is imperative. We selected gentamicin resistance in gonococci in vitro, identified the novel gentamicin-resistance mutations, and examined the biofitness of a high-level gentamicin-resistant mutant. METHODS Low- and high-level gentamicin resistance was selected in WHO X (gentamicin MIC = 4 mg/L) on gentamicin-gradient agar plates. Selected mutants were whole-genome sequenced. Potential gentamicin-resistance fusA mutations were transformed into WT strains to verify their impact on gentamicin MICs. The biofitness of high-level gentamicin-resistant mutants was examined using a competitive assay in a hollow-fibre infection model. RESULTS WHO X mutants with gentamicin MICs of up to 128 mg/L were selected. Primarily selected fusA mutations were further investigated, and fusAR635L and fusAM520I + R635L were particularly interesting. Different mutations in fusA and ubiM were found in low-level gentamicin-resistant mutants, while fusAM520I was associated with high-level gentamicin resistance. Protein structure predictions showed that fusAM520I is located in domain IV of the elongation factor-G (EF-G). The high-level gentamicin-resistant WHO X mutant was outcompeted by the gentamicin-susceptible WHO X parental strain, suggesting lower biofitness. CONCLUSIONS We describe the first high-level gentamicin-resistant gonococcal isolate (MIC = 128 mg/L), which was selected in vitro through experimental evolution. The most substantial increases of the gentamicin MICs were caused by mutations in fusA (G1560A and G1904T encoding EF-G M520I and R635L, respectively) and ubiM (D186N). The high-level gentamicin-resistant N. gonorrhoeae mutant showed impaired biofitness.
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Affiliation(s)
- Daniel Golparian
- Department of Laboratory Medicine, Microbiology, Faculty of Medicine and Health, WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, Örebro University, Örebro, Sweden
| | - Susanne Jacobsson
- Department of Laboratory Medicine, Microbiology, Faculty of Medicine and Health, WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, Örebro University, Örebro, Sweden
| | - Concerta L Holley
- Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA, USA
| | - William M Shafer
- Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA, USA
- The Emory Antibiotic Resistance Center, Emory University School of Medicine, Atlanta, GA, USA
- Laboratories of Bacterial Pathogenesis, Veterans Affairs Medical Center, Decatur, GA, USA
| | - Magnus Unemo
- Department of Laboratory Medicine, Microbiology, Faculty of Medicine and Health, WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, Örebro University, Örebro, Sweden
- Institute for Global Health, University College London (UCL), London, UK
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Li YX, Lin W, Han YH, Wang YQ, Wang T, Zhang H, Zhang Y, Wang SS. Biodegradation of p-hydroxybenzoic acid in Herbaspirillum aquaticum KLS-1 isolated from tailing soil: Characterization and molecular mechanism. JOURNAL OF HAZARDOUS MATERIALS 2023; 456:131669. [PMID: 37236108 DOI: 10.1016/j.jhazmat.2023.131669] [Citation(s) in RCA: 14] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/02/2023] [Revised: 05/09/2023] [Accepted: 05/19/2023] [Indexed: 05/28/2023]
Abstract
The wide distribution of p-hydroxybenzoic acid (PHBA) in the environments has attracted great concerns due to its potential risks to organisms. Bioremediation is considered a green way to remove PHBA from environment. Here, a new PHBA-degrading bacterium Herbaspirillum aquaticum KLS-1was isolated and its PHBA degradation mechanisms were fully evaluated. Results showed that strain KLS-1 could utilize PHBA as the sole carbon source and completely degrade 500 mg/L PHBA within 18 h. The optimal conditions for bacterial growth and PHBA degradation were pH values of 6.0-8.0, temperatures of 30 °C-35 °C, shaking speed of 180 rpm, Mg2+ concentration of 2.0 mM and Fe2+ concentration of 1.0 mM. Draft genome sequencing and functional gene annotations identified three operons (i.e., pobRA, pcaRHGBD and pcaRIJ) and several free genes possibly participating in PHBA degradation. The key genes pobA, ubiA, fadA, ligK and ubiG involved in the regulation of protocatechuate and ubiquinone (UQ) metabolisms were successfully amplified in strain KLS-1 at mRNA level. Our data suggested that PHBA could be degraded by strain KLS-1 via the protocatechuate ortho-/meta-cleavage pathway and UQ biosynthesis pathway. This study has provided a new PHBA-degrading bacterium for potential bioremediation of PHBA pollution.
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Affiliation(s)
- Yi-Xi Li
- College of Environmental and Resource Science, Fujian Normal University, Fuzhou 350117, Fujian, China; Fujian Key Laboratory of Pollution Control and Resource Reuse, Fuzhou 350117, Fujian, China
| | - Wei Lin
- College of Life Science, Fujian Normal University, Fuzhou 350117, Fujian, China
| | - Yong-He Han
- College of Environmental and Resource Science, Fujian Normal University, Fuzhou 350117, Fujian, China; Fujian Key Laboratory of Pollution Control and Resource Reuse, Fuzhou 350117, Fujian, China.
| | - Yao-Qiang Wang
- College of Environmental and Resource Science, Fujian Normal University, Fuzhou 350117, Fujian, China; Fujian Key Laboratory of Pollution Control and Resource Reuse, Fuzhou 350117, Fujian, China
| | - Tao Wang
- College of Environmental and Resource Science, Fujian Normal University, Fuzhou 350117, Fujian, China; Fujian Key Laboratory of Pollution Control and Resource Reuse, Fuzhou 350117, Fujian, China
| | - Hong Zhang
- College of Environmental and Resource Science, Fujian Normal University, Fuzhou 350117, Fujian, China; Fujian Key Laboratory of Pollution Control and Resource Reuse, Fuzhou 350117, Fujian, China
| | - Yong Zhang
- College of Environmental and Resource Science, Fujian Normal University, Fuzhou 350117, Fujian, China; Fujian Key Laboratory of Pollution Control and Resource Reuse, Fuzhou 350117, Fujian, China
| | - Shan-Shan Wang
- College of Integrative Medicine, Fujian University of Traditional Chinese Medicine, Fuzhou 350122, China.
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Pujari V, Rozman K, Dhiman RK, Aldrich CC, Crick DC. Mycobacterial MenG: Partial Purification, Characterization, and Inhibition. ACS Infect Dis 2022; 8:2430-2440. [PMID: 36417754 DOI: 10.1021/acsinfecdis.2c00190] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022]
Abstract
Menaquinone (MK) is an essential component of the electron transport chain (ETC) in the gram-variable Mycobacterium tuberculosis and many Gram-positive pathogens. Three genes in the M. tuberculosis genome were annotated as methyltransferases involved in lipoquinone synthesis in mycobacteria. Heterologous expression of Rv0558 complemented an ubiE (the quinone C-methyltransferase involved in ubiquinone and menaquinone synthesis) deletion in Escherichia coli, and expression in a wild-type E. coli strain increased quinone C-methyltransferase specific activity by threefold. Rv0558 encodes a canonical C-methyltransferase or, more specifically, a S-adenosylmethionine/demethylmenaquinol methyltransferase. Partially purified recombinant protein catalyzed the formation of MK from demethylmenaquinone (DMK), although the activity of the recombinant protein was low and appeared to require a cofactor or intact membrane structure for activity. Membrane preparations from irradiated M. tuberculosis also showed poor activity; however, membrane preparations from wild-type Mycobacterium smegmatis showed robust, substrate-dependent activity. The apparent Km values for demethylmenaquinone and SAM were 14 ± 5.0 and 17 ± 7.0 μM, respectively. Interestingly, addition of dithiothreitol, dithionite, NADH, or other substrates of primary dehydrogenases to reaction mixtures containing membrane preparations stimulated the activity. Thus, these observations strongly suggest that demethylmenaquinol is the actual substrate of MenG. Ro 48-8071, previously reported to inhibit mycobacterial MK synthesis and growth, inhibited Rv0558 activity with an IC50 value of 5.1 ± 0.5 μM, and DG70 (GSK1733953A), first described as a respiration inhibitor in M. tuberculosis, inhibits MenG activity with an IC50 value of 2.6 ± 0.6 μM.
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Affiliation(s)
- Venugopal Pujari
- Mycobacteria Research Laboratories, Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, Colorado 80523, United States
| | - Kaja Rozman
- Department of Medicinal Chemistry, University of Minnesota, 308 Harvard Street Southeast, Minneapolis, Minnesota 55455, United States
| | - Rakesh K Dhiman
- Mycobacteria Research Laboratories, Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, Colorado 80523, United States
| | - Courtney C Aldrich
- Department of Medicinal Chemistry, University of Minnesota, 308 Harvard Street Southeast, Minneapolis, Minnesota 55455, United States
| | - Dean C Crick
- Mycobacteria Research Laboratories, Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, Colorado 80523, United States
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Zhang L, Li YL, Hu JH, Liu ZY. Overexpression of enzymes in glycolysis and energy metabolic pathways to enhance coenzyme Q10 production in Rhodobacter sphaeroides VK-2-3. Front Microbiol 2022; 13:931470. [PMID: 36033867 PMCID: PMC9412181 DOI: 10.3389/fmicb.2022.931470] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2022] [Accepted: 07/26/2022] [Indexed: 12/02/2022] Open
Abstract
We subjected the components of the glycolysis and energy metabolism pathways of Rhodobacter sphaeroides (R. sphaeroides) to metabolic engineering to improve the titer and yield of coenzyme Q10 (CoQ10). Phosphofructokinase (PFK), cyclic adenylate-dependent protein kinase (PKAC), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and adenosine triphosphate hydrolase (KdpC) were overexpressed in R. sphaeroides VK-2-3 (VK-2-3). The strains were labeled R. sphaeroides PFK (RS.PFK), RS.PKAC, RS.PFK–PKAC, RS.KdpC, RS.GAPDH, and RS.KdpC–GAPDH. Results showed that the CoQ10 titers of RS.PFK, RS.PKAC, and RS.PFK–PKAC were 300.96 ± 0.87, 405.94 ± 4.77, and 379.94 ± 0.42 mg/l, respectively. The CoQ10 titers of RS.PFK and VK-2-3 were not significantly different; however, those for RS.PKAC and RS.PFK–PKAC were 13 and 6% higher than that of VK-2-3, respectively. Further, the titers of RS.KdpC, RS.GAPDH, and RS.KdpC–GAPDH were 360.17 ± 0.39, 409.79 ± 0.76, and 359.87 ± 1.14 mg/l, respectively. The titers of RS.KdpC and RS.KdpC–GAPDH were not significantly different from that for VK-2-3, whereas that for RS.GAPDH was 14% higher than that of VK-2-3. Finally, when the cultures of RS.GAPDH and VK-2-3 were scaled up in 5-L fermenters, the CoQ10 titers and RS.GAPDH yields increased by 44.3 and 37.8%, respectively, compared with VK-2-3.To the best of our knowledge, the glycolysis pathway of R. sphaeroides was studied for the first time in this study. We genetically modified the components of the energy metabolism pathway to obtain the strain with high yield of CoQ10 mutant RS.GAPDH. The findings of this study can serve as a basis for future studies involving metabolic engineering of CoQ10-producing strains.
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Affiliation(s)
- Long Zhang
- Inner Mongolia Energy Conservation and Emission Reduction Engineering Technology Research Center for Fermentation Industry, Hohhot, Inner Mongolia, China
- Engineering Research Center of Inner Mongolia for Green Manufacturing in Bio-fermentation Industry, Hohhot, Inner Mongolia, China
- College of Chemical Engineering, Inner Mongolia University of Technology, Hohhot, Inner Mongolia, China
| | - Yong-li Li
- Inner Mongolia Energy Conservation and Emission Reduction Engineering Technology Research Center for Fermentation Industry, Hohhot, Inner Mongolia, China
- Engineering Research Center of Inner Mongolia for Green Manufacturing in Bio-fermentation Industry, Hohhot, Inner Mongolia, China
- College of Chemical Engineering, Inner Mongolia University of Technology, Hohhot, Inner Mongolia, China
| | - Jian-hua Hu
- Inner Mongolia Energy Conservation and Emission Reduction Engineering Technology Research Center for Fermentation Industry, Hohhot, Inner Mongolia, China
- Engineering Research Center of Inner Mongolia for Green Manufacturing in Bio-fermentation Industry, Hohhot, Inner Mongolia, China
- College of Chemical Engineering, Inner Mongolia University of Technology, Hohhot, Inner Mongolia, China
| | - Zhan-ying Liu
- Inner Mongolia Energy Conservation and Emission Reduction Engineering Technology Research Center for Fermentation Industry, Hohhot, Inner Mongolia, China
- Engineering Research Center of Inner Mongolia for Green Manufacturing in Bio-fermentation Industry, Hohhot, Inner Mongolia, China
- College of Chemical Engineering, Inner Mongolia University of Technology, Hohhot, Inner Mongolia, China
- *Correspondence: Zhan-ying Liu,
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Prince RC, Dutton PL, Gunner MR. The aprotic electrochemistry of quinones. BIOCHIMICA ET BIOPHYSICA ACTA. BIOENERGETICS 2022; 1863:148558. [PMID: 35413248 DOI: 10.1016/j.bbabio.2022.148558] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/19/2021] [Revised: 03/26/2022] [Accepted: 04/05/2022] [Indexed: 05/09/2023]
Abstract
Quinones play important roles in biological electron transfer reactions in almost all organisms, with specific roles in many physiological processes and chemotherapy. Quinones participate in two-electron, two-proton reactions in aqueous solution at equilibrium near neutral pH, but protons often lag behind the electron transfers. The relevant reactions in proteins are often sequential one electron redox processes without involving protons. Here we report the aprotic electrochemistry of the two half-couples, Q/Q.- and Q.-/Q=, of 11 parent quinones and 118 substituted 1,4-benzoquinones, 91 1,4-naphthoquinones, and 107 9,10-anthraquinones. The measured redox potentials are fit quite well with the Hammett para sigma (σpara) parameter. Occasional exceptions can involve important groups, such as methoxy substituents in ubiquinone and hydroxy substituents in therapeutics. These can generally be explained by reasonable conjectures involving steric clashes and internal hydrogen bonds. We also provide data for 25 other quinones, 2 double quinones and 15 non-quinones, all measured under similar conditions.
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Affiliation(s)
| | - P Leslie Dutton
- The Johnson Research Foundation, Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, PA 10104, USA
| | - M R Gunner
- Physics Department City College of New York in the City University of New York, NY 10031, USA.
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22
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Roberts GW, Leys D. Structural insights into UbiD reversible decarboxylation. Curr Opin Struct Biol 2022; 75:102432. [PMID: 35843126 DOI: 10.1016/j.sbi.2022.102432] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/26/2022] [Revised: 06/14/2022] [Accepted: 06/17/2022] [Indexed: 11/03/2022]
Abstract
The ubiquitous UbiX-UbiD system is associated with a wide range of microbial (de)carboxylation reactions. Recent X-ray crystallographic studies have contributed to elucidating the enigmatic mechanism underpinning the conversion of α,β-unsaturated acids by this system. The UbiD component utilises a unique cofactor, prenylated flavin (prFMN), generated by the bespoke action of the associated UbiX flavin prenyltransferase. Structure determination of a range of UbiX/UbiD representatives has revealed a generic mode of action for both the flavin-to-prFMN metamorphosis and the (de)carboxylation. In contrast to the conserved UbiX, the UbiD superfamily is associated with a versatile substrate range. The latter is reflected in the considerable variety of UbiD quaternary structure, dynamic behaviour and active site architecture. Directed evolution of UbiD enzymes has taken advantage of this apparent malleability to generate new variants supporting in vivo hydrocarbon production. Other applications include coupling UbiD to carboxylic acid reductase to convert alkenes into α,β-unsaturated aldehydes via enzymatic CO2 fixation.
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Affiliation(s)
- George W Roberts
- Manchester Institute of Biotechnology, Department of Chemistry, University of Manchester, 131 Princess Street, Manchester, M1 7DN, UK
| | - David Leys
- Manchester Institute of Biotechnology, Department of Chemistry, University of Manchester, 131 Princess Street, Manchester, M1 7DN, UK.
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23
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Feng S, Kong L, Gee S, Im W. Molecular Condensate in a Membrane: A Tugging Game between Hydrophobicity and Polarity with Its Biological Significance. LANGMUIR : THE ACS JOURNAL OF SURFACES AND COLLOIDS 2022; 38:5955-5962. [PMID: 35503859 DOI: 10.1021/acs.langmuir.2c00876] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/14/2023]
Abstract
Lipid self-organization and lipid-water interfaces have been an increasingly important topic positioned at the crossroads of physical chemistry and biology. Some neutral lipids can partition into the biomembrane and play an important biological role. In this study, we have used all-atom molecular dynamics simulations to dissect the partition, aggregation, flip-flop, and modulation of neutral lipids including (i) menaquinone/menaquinol, (ii) ubiquinone/ubiquinol, and (iii) triacylglycerol. The partitioning of these molecules is driven by the balancing force between headgroup hydrophilicity and acyl chain hydrophobicity as well as the lipid shapes. We then discuss the emerging questions in this area, share our own perspectives, and mention the development of the CHARMM-GUI membrane modeling platform, which enables further computational investigations into those questions.
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24
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Yi B, Dalpke AH. Revisiting the intrageneric structure of the genus Pseudomonas with complete whole genome sequence information: Insights into diversity and pathogen-related genetic determinants. INFECTION, GENETICS AND EVOLUTION : JOURNAL OF MOLECULAR EPIDEMIOLOGY AND EVOLUTIONARY GENETICS IN INFECTIOUS DISEASES 2022; 97:105183. [PMID: 34920102 DOI: 10.1016/j.meegid.2021.105183] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/06/2021] [Revised: 08/09/2021] [Accepted: 12/08/2021] [Indexed: 06/14/2023]
Abstract
Pseudomonas spp. exhibit considerable differences in host specificity and virulence. Most Pseudomonas species were isolated exclusively from environmental sources, ranging from soil to plants, but some Pseudomonas species have been detected from versatile sources, including both human host and environmental sources. Understanding genome variations that generate the tremendous diversity in Pseudomonas biology is important in controlling the incidence of infections. With a data set of 704 Pseudomonas complete whole genome sequences representing 186 species, Pseudomonas intrageneric structure was investigated by hierarchical clustering based on average nucleotide identity, and by phylogeny analysis based on concatenated core-gene alignment. Further comparative functional analyses indicated that Pseudomonas species only living in natural habitats lack multiple functions that are important in the regulation of bacterial pathogenesis, indicating the possession of these functions might be characteristic of Pseudomonas human pathogens. Moreover, we have performed pan-genome based homogeneity analyses, and detected genes with conserved structures but diversified functions across the Pseudomonas genomes, suggesting these genes play a role in driving diversity. In summary, this study provided insights into the dynamics of genome diversity and pathogen-related genetic determinants in Pseudomonas, which might help the development of more targeted antibiotics for the treatment of Pseudomonas infections.
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Affiliation(s)
- Buqing Yi
- Institute of Medical Microbiology and Virology, Medical Faculty, Technische Universität Dresden, Dresden, Germany.
| | - Alexander H Dalpke
- Institute of Medical Microbiology and Virology, Medical Faculty, Technische Universität Dresden, Dresden, Germany.
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25
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The Protective Effect of Ubiquinone against the Amyloid Peptide in Endothelial Cells Is Isoprenoid Chain Length-Dependent. Antioxidants (Basel) 2021; 10:antiox10111806. [PMID: 34829677 PMCID: PMC8615161 DOI: 10.3390/antiox10111806] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2021] [Revised: 11/09/2021] [Accepted: 11/10/2021] [Indexed: 12/23/2022] Open
Abstract
Vascular brain pathology constitutes a common feature in neurodegenerative diseases that could underlie their development. Indeed, vascular dysfunction acts synergistically with neurodegenerative changes to exacerbate the cognitive impairment found in Alzheimer’s disease. Different injuries such as hypertension, high glucose, atherosclerosis associated with oxidized low-density lipoprotein or inflammation induce NADPH oxidase activation, overproduction of reactive oxygen species, and apoptosis in endothelial cells. Since it has been shown that pretreatment of cultured endothelial cells with the lipophilic antioxidant coenzyme Q10 (CoQ10) displays a protective effect against the deleterious injuries caused by different agents, this study explores the cytoprotective role of different CoQs homologues against Aβ25–35-induced damage and demonstrates that only pretreatment with CoQ10 protects endothelial brain cells from Aβ25–35-induced damage. Herein, we show that CoQ10 constitutes the most effective ubiquinone in preventing NADPH oxidase activity and reducing both reactive oxygen species generation and the increase in free cytosolic Ca2+ induced by Aβ25–35, ultimately preventing apoptosis and necrosis. The specific cytoprotective effect of CoQ with a side chain of 10 isoprenoid units could be explained by the fact that CoQ10 is the only ubiquinone that significantly reduces the entry of Aβ25–35 into the mitochondria.
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26
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Datar PM, Marsh ENG. Decarboxylation of Aromatic Carboxylic Acids by the Prenylated-FMN-dependent Enzyme Phenazine-1-carboxylic Acid Decarboxylase. ACS Catal 2021. [DOI: 10.1021/acscatal.1c03040] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023]
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27
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Mudge MC, Nunn BL, Firth E, Ewert M, Hales K, Fondrie WE, Noble WS, Toner J, Light B, Junge KA. Subzero, saline incubations of Colwellia psychrerythraea reveal strategies and biomarkers for sustained life in extreme icy environments. Environ Microbiol 2021; 23:3840-3866. [PMID: 33760340 PMCID: PMC8475265 DOI: 10.1111/1462-2920.15485] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2020] [Accepted: 03/22/2021] [Indexed: 11/26/2022]
Abstract
Colwellia psychrerythraea is a marine psychrophilic bacterium known for its remarkable ability to maintain activity during long-term exposure to extreme subzero temperatures and correspondingly high salinities in sea ice. These microorganisms must have adaptations to both high salinity and low temperature to survive, be metabolically active, or grow in the ice. Here, we report on an experimental design that allowed us to monitor culturability, cell abundance, activity and proteomic signatures of C. psychrerythraea strain 34H (Cp34H) in subzero brines and supercooled sea water through long-term incubations under eight conditions with varying subzero temperatures, salinities and nutrient additions. Shotgun proteomics found novel metabolic strategies used to maintain culturability in response to each independent experimental variable, particularly in pathways regulating carbon, nitrogen and fatty acid metabolism. Statistical analysis of abundances of proteins uniquely identified in isolated conditions provide metabolism-specific protein biosignatures indicative of growth or survival in either increased salinity, decreased temperature, or nutrient limitation. Additionally, to aid in the search for extant life on other icy worlds, analysis of detected short peptides in -10°C incubations after 4 months identified over 500 potential biosignatures that could indicate the presence of terrestrial-like cold-active or halophilic metabolisms on other icy worlds.
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Affiliation(s)
- Miranda C. Mudge
- Department of Genome Sciences, University of Washington, Seattle, WA
- Department of Molecular and Cellular Biology, University of Washington, Seattle, WA
| | - Brook L. Nunn
- Department of Genome Sciences, University of Washington, Seattle, WA
- Astrobiology Program, University of Washington, Seattle, WA
| | - Erin Firth
- Applied Physics Lab, Polar Science Center, University of Washington, Seattle, WA
| | - Marcela Ewert
- Applied Physics Lab, Polar Science Center, University of Washington, Seattle, WA
| | - Kianna Hales
- Department of Genome Sciences, University of Washington, Seattle, WA
| | | | - William S. Noble
- Department of Genome Sciences, University of Washington, Seattle, WA
- Paul G. Allen School of Computer Science and Engineering, University of Washington, Seattle, WA
| | - Jonathan Toner
- Department of Earth and Space Sciences, University of Washington, Seattle, WA
| | - Bonnie Light
- Applied Physics Lab, Polar Science Center, University of Washington, Seattle, WA
| | - Karen A. Junge
- Applied Physics Lab, Polar Science Center, University of Washington, Seattle, WA
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28
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Deering RW, Whalen KE, Alvarez I, Daffinee K, Beganovic M, LaPlante KL, Kishore S, Zhao S, Cezairliyan B, Yu S, Rosario M, Mincer TJ, Rowley DC. Identification of a bacteria-produced benzisoxazole with antibiotic activity against multi-drug resistant Acinetobacter baumannii. J Antibiot (Tokyo) 2021; 74:370-380. [PMID: 33580212 PMCID: PMC7879144 DOI: 10.1038/s41429-021-00412-7] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/29/2020] [Revised: 12/21/2020] [Accepted: 01/04/2021] [Indexed: 01/05/2023]
Abstract
The emergence of multi-drug resistant pathogenic bacteria represents a serious and growing threat to national healthcare systems. Most pressing is an immediate need for the development of novel antibacterial agents to treat Gram-negative multi-drug resistant infections, including the opportunistic, hospital-derived pathogen, Acinetobacter baumannii. Herein we report a naturally occurring 1,2-benzisoxazole with minimum inhibitory concentrations as low as 6.25 μg ml-1 against clinical strains of multi-drug resistant A. baumannii and investigate its possible mechanisms of action. This molecule represents a new chemotype for antibacterial agents against A. baumannii and is easily accessed in two steps via de novo synthesis. In vitro testing of structural analogs suggest that the natural compound may already be optimized for activity against this pathogen. Our results demonstrate that supplementation of 4-hydroxybenzoate in minimal media was able to reverse 1,2-benzisoxazole's antibacterial effects in A. baumannii. A search of metabolic pathways involving 4-hydroxybenzoate coupled with molecular modeling studies implicates two enzymes, chorismate pyruvate-lyase and 4-hydroxybenzoate octaprenyltransferase, as promising leads for the target of 3,6-dihydroxy-1,2-benzisoxazole.
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Affiliation(s)
- Robert W Deering
- Department of Biomedical and Pharmaceutical Sciences, College of Pharmacy, University of Rhode Island, Kingston, RI, USA
| | | | - Ivan Alvarez
- Department of Biomedical and Pharmaceutical Sciences, College of Pharmacy, University of Rhode Island, Kingston, RI, USA
| | - Kathryn Daffinee
- Department of Pharmacy Practice, College of Pharmacy, University of Rhode Island, Kingston, RI, USA
- Infectious Diseases Research Program, Providence Veterans Affairs Medical Center, Providence, RI, USA
| | - Maya Beganovic
- Department of Pharmacy Practice, College of Pharmacy, University of Rhode Island, Kingston, RI, USA
- Infectious Diseases Research Program, Providence Veterans Affairs Medical Center, Providence, RI, USA
| | - Kerry L LaPlante
- Department of Pharmacy Practice, College of Pharmacy, University of Rhode Island, Kingston, RI, USA
- Infectious Diseases Research Program, Providence Veterans Affairs Medical Center, Providence, RI, USA
| | - Shreya Kishore
- Department of Biology, Haverford College, Haverford, PA, USA
| | - Sijing Zhao
- Department of Biology, Haverford College, Haverford, PA, USA
| | | | - Shen Yu
- Octagon Therapeutics, Inc., Cambridge, MA, USA
| | - Margaret Rosario
- Department of Biomedical and Pharmaceutical Sciences, College of Pharmacy, University of Rhode Island, Kingston, RI, USA
| | - Tracy J Mincer
- Wilkes Honors College and Harbor Branch Oceanographic Institute, Florida Atlantic University, Boca Raton, FL, USA.
| | - David C Rowley
- Department of Biomedical and Pharmaceutical Sciences, College of Pharmacy, University of Rhode Island, Kingston, RI, USA.
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29
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The In Vitro Production of prFMN for Reconstitution of UbiD Enzymes. Methods Mol Biol 2021; 2280:219-227. [PMID: 33751438 DOI: 10.1007/978-1-0716-1286-6_14] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023]
Abstract
Prenylated flavin (prFMN) is a modified FMN cofactor, the isoalloxazine is extended by an additional six membered nonaromatic ring. The modification confers azomethine ylide characteristics on the oxidised prFMN, allowing it to support the reversible nonoxidative decarboxylation of unsaturated acids by the UbiD family of decarboxylases. In absence of a chemical synthesis route for prFMN, enzymatic production by the flavin prenyltransferase, UbiX, is required for in vitro reconstitution of prFMN-dependent enzymes. Here we provide an overview of the methods for producing prFMN in vitro using the flavin prenyltransferase UbiX, and the subsequent reconstitution and activation of UbiD enzymes.
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30
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Bell RT, Wolf YI, Koonin EV. Modified base-binding EVE and DCD domains: striking diversity of genomic contexts in prokaryotes and predicted involvement in a variety of cellular processes. BMC Biol 2020; 18:159. [PMID: 33148243 PMCID: PMC7641849 DOI: 10.1186/s12915-020-00885-2] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/14/2020] [Accepted: 10/01/2020] [Indexed: 12/11/2022] Open
Abstract
BACKGROUND DNA and RNA of all cellular life forms and many viruses contain an expansive repertoire of modified bases. The modified bases play diverse biological roles that include both regulation of transcription and translation, and protection against restriction endonucleases and antibiotics. Modified bases are often recognized by dedicated protein domains. However, the elaborate networks of interactions and processes mediated by modified bases are far from being completely understood. RESULTS We present a comprehensive census and classification of EVE domains that belong to the PUA/ASCH domain superfamily and bind various modified bases in DNA and RNA. We employ the "guilt by association" approach to make functional inferences from comparative analysis of bacterial and archaeal genomes, based on the distribution and associations of EVE domains in (predicted) operons and functional networks of genes. Prokaryotes encode two classes of EVE domain proteins, slow-evolving and fast-evolving ones. Slow-evolving EVE domains in α-proteobacteria are embedded in conserved operons, potentially involved in coupling between translation and respiration, cytochrome c biogenesis in particular, via binding 5-methylcytosine in tRNAs. In β- and γ-proteobacteria, the conserved associations implicate the EVE domains in the coordination of cell division, biofilm formation, and global transcriptional regulation by non-coding 6S small RNAs, which are potentially modified and bound by the EVE domains. In eukaryotes, the EVE domain-containing THYN1-like proteins have been reported to inhibit PCD and regulate the cell cycle, potentially, via binding 5-methylcytosine and its derivatives in DNA and/or RNA. We hypothesize that the link between PCD and cytochrome c was inherited from the α-proteobacterial and proto-mitochondrial endosymbiont and, unexpectedly, could involve modified base recognition by EVE domains. Fast-evolving EVE domains are typically embedded in defense contexts, including toxin-antitoxin modules and type IV restriction systems, suggesting roles in the recognition of modified bases in invading DNA molecules and targeting them for restriction. We additionally identified EVE-like prokaryotic Development and Cell Death (DCD) domains that are also implicated in defense functions including PCD. This function was inherited by eukaryotes, but in animals, the DCD proteins apparently were displaced by the extended Tudor family proteins, whose partnership with Piwi-related Argonautes became the centerpiece of the Piwi-interacting RNA (piRNA) system. CONCLUSIONS Recognition of modified bases in DNA and RNA by EVE-like domains appears to be an important, but until now, under-appreciated, common denominator in a variety of processes including PCD, cell cycle control, antivirus immunity, stress response, and germline development in animals.
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Affiliation(s)
- Ryan T Bell
- National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD, 20894, USA
| | - Yuri I Wolf
- National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD, 20894, USA
| | - Eugene V Koonin
- National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD, 20894, USA.
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31
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Beaupre BA, Moran GR. N5 Is the New C4a: Biochemical Functionalization of Reduced Flavins at the N5 Position. Front Mol Biosci 2020; 7:598912. [PMID: 33195440 PMCID: PMC7662398 DOI: 10.3389/fmolb.2020.598912] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2020] [Accepted: 10/05/2020] [Indexed: 12/31/2022] Open
Abstract
For three decades the C4a-position of reduced flavins was the known site for covalency within flavoenzymes. The reactivity of this position of the reduced isoalloxazine ring with the dioxygen ground-state triplet established the C4a as a site capable of one-electron chemistry. Within the last two decades new types of reduced flavin reactivity have been documented. These studies reveal that the N5 position is also a protean site of reactivity, that is capable of nucleophilic attack to form covalent bonds with substrates. In addition, though the precise mechanism of dioxygen reactivity is yet to be definitively demonstrated, it is clear that the N5 position is directly involved in substrate oxygenation in some enzymes. In this review we document the lineage of discoveries that identified five unique modes of N5 reactivity that collectively illustrate the versatility of this position of the reduced isoalloxazine ring.
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Affiliation(s)
- Brett A Beaupre
- Department of Chemistry and Biochemistry, Loyola University Chicago, Chicago, IL, United States
| | - Graham R Moran
- Department of Chemistry and Biochemistry, Loyola University Chicago, Chicago, IL, United States
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32
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Identification of Genes Associated with Sensitivity to Ultraviolet A (UVA) Irradiation by Transposon Mutagenesis of Vibrio parahaemolyticus. APPLIED SCIENCES-BASEL 2020. [DOI: 10.3390/app10165549] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
Ultraviolet (UV) irradiation is used to disinfect water and food and can be classified as UVA (detected at wavelengths 320–400 nm), UVB (280–320 nm), and UVC (<280 nm). We developed a method for UVA sterilization of equipment with a UVA-light-emitting diode (LED); however, a high rate of fluence was needed to promote pathogen inactivation. The aim of this study was to identify genes associated with UVA sensitivity with the goal of improving UVA-LED-mediated bactericidal activity. We constructed a transposon-mutant library of Vibrio parahaemolyticus and selected six mutants with high sensitivity to UVA irradiation. Genes associated with this phenotype include F-type H+-transporting ATPases (atp), as well as those involved in general secretion (gsp), and ubiquinone and terpenoid-quinone biosynthesis (ubi). Gene complementation resulted in decreased sensitivity to UVA-LED. The atp mutants had lower intracellular adenosine triphosphate (ATP) concentrations than the wild-type treatment, with 20 mM L-serine resulting in elevated ATP concentrations and decreased sensitivity to UVA-LED. The gsp mutants exhibited high levels of extracellular protein transport and the ubi mutants exhibited significantly different intracellular concentrations of ubiquinone-8. Taken together, our results suggest that the protein products of the atp, gsp, and ubi genes may regulate sensitivity to UVA irradiation.
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33
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Cohen DR, Townsend CA. C-N-Coupled Metabolites Yield Insights into Dynemicin A Biosynthesis. Chembiochem 2020; 21:2137-2142. [PMID: 32198800 PMCID: PMC7685002 DOI: 10.1002/cbic.202000177] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2020] [Indexed: 11/08/2022]
Abstract
The biosynthesis of the three structural subclasses of enediyne antitumor antibiotics remains largely unknown beyond a common C16 -hexaene precursor. For the anthraquinone-fused subtype, however, an unexpected iodoanthracene γ-thiolactone was established to be a mid-pathway intermediate to dynemicin A. Having deleted a putative flavin-dependent oxidoreductase from the dynemicin biosynthetic gene cluster, we can now report four metabolites that incorporate the iodoanthracene and reveal the formation of the C-N bond linking the anthraquinone and enediyne halves emblematic of this structural subclass. The coupling of an aryl iodide and an amine is familiar from organometallic chemistry, but has little or no precedent in natural product biosynthesis. These metabolites suggest further that enediyne formation occurs early in the overall biosynthesis, and that even earlier events might convert the C16 -hexaene to a common C15 intermediate that partitions to enediyne and anthraquinone building blocks for the heterodimerization.
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Affiliation(s)
- Douglas R Cohen
- Department of Chemistry, The Johns Hopkins University, 3400 North Charles Street, Baltimore, MD, 21218, USA
| | - Craig A Townsend
- Department of Chemistry, The Johns Hopkins University, 3400 North Charles Street, Baltimore, MD, 21218, USA
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34
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Zhu Y, Plaza N, Kojima Y, Yoshida M, Zhang J, Jellison J, Pingali SV, O’Neill H, Goodell B. Nanostructural Analysis of Enzymatic and Non-enzymatic Brown Rot Fungal Deconstruction of the Lignocellulose Cell Wall †. Front Microbiol 2020; 11:1389. [PMID: 32670241 PMCID: PMC7326796 DOI: 10.3389/fmicb.2020.01389] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2020] [Accepted: 05/29/2020] [Indexed: 12/14/2022] Open
Abstract
Brown rot (BR) decay mechanisms employ carbohydrate-active enzymes (CAZymes) as well as a unique non-enzymatic chelator-mediated Fenton (CMF) chemistry to deconstruct lignocellulosic materials. Unlike white rot fungi, BR fungi lack peroxidases for lignin deconstruction, and also lack some endoglucanase/cellobiohydrolase activities. The role that the CMF mechanism plays in "opening up" the wood cell wall structure in advance of enzymatic action, and any interaction between CMF constituents and the selective CAZyme suite that BRs possess, is still unclear. Expression patterns for CMF redox metabolites and lytic polysaccharide monooxygenase (LPMO-AA9 family) genes showed that some LPMO isozymes were upregulated with genes associated with CMF at early stages of brown rot by Gloeophyllum trabeum. In the structural studies, wood decayed by the G. trabeum was compared to CMF-treated wood, or CMF-treated wood followed by treatment with either the early-upregulated LPMO or a commercial CAZyme cocktail. Structural modification of decayed/treated wood was characterized using small angle neutron scattering. CMF treatment produced neutron scattering patterns similar to that of the BR decay indicating that both systems enlarged the nanopore structure of wood cell walls to permit enzyme access. Enzymatic deconstruction of cellulose or lignin in raw wood samples was not achieved via CAZyme cocktail or LPMO enzyme action alone. CMF treatment resulted in depolymerization of crystalline cellulose as attack progressed from the outer regions of individual crystallites. Multiple pulses of CMF treatment on raw wood showed a progressive increase in the spacing between the cellulose elementary fibrils (EFs), indicating the CMF eroded the matrix outside the EF bundles, leading to less tightly packed EFs. Peracetic acid delignification treatment enhanced subsequent CMF treatment effects, and allowed both enzyme systems to further increase spacing of the EFs. Moreover, even after a single pulse of CMF treatment, both enzymes were apparently able to penetrate the cell wall to further increase EF spacing. The data suggest the potential for the early-upregulated LPMO enzyme to work in association with CMF chemistry, suggesting that G. trabeum may have adopted mechanisms to integrate non-enzymatic and enzymatic chemistries together during early stages of brown rot decay.
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Affiliation(s)
- Yuan Zhu
- School of Materials Science and Engineering, Central South University of Forestry and Technology, Changsha, China
| | - Nayomi Plaza
- Forest Products Laboratory, USDA Forest Service, Madison, WI, United States
| | - Yuka Kojima
- Department of Environmental and Natural Resource Science, Tokyo University of Agriculture and Technology, Fuchu, Japan
| | - Makoto Yoshida
- Department of Environmental and Natural Resource Science, Tokyo University of Agriculture and Technology, Fuchu, Japan
| | - Jiwei Zhang
- Department of Bioproducts and Biosystems Engineering, University of Minnesota, Saint Paul, MN, United States
| | - Jody Jellison
- Center for Agriculture, Food and the Environment, University of Massachusetts, Amherst, MA, United States
| | - Sai Venkatesh Pingali
- Biology and Soft Matter Division, Oak Ridge National Laboratory, Oak Ridge, TN, United States
| | - Hugh O’Neill
- Biology and Soft Matter Division, Oak Ridge National Laboratory, Oak Ridge, TN, United States
| | - Barry Goodell
- Department of Microbiology, Morrill Science Center IV-N, University of Massachusetts, Amherst, MA, United States
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35
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Wang Y, Hekimi S. The Complexity of Making Ubiquinone. Trends Endocrinol Metab 2019; 30:929-943. [PMID: 31601461 DOI: 10.1016/j.tem.2019.08.009] [Citation(s) in RCA: 44] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/03/2019] [Revised: 08/19/2019] [Accepted: 08/20/2019] [Indexed: 12/15/2022]
Abstract
Ubiquinone (UQ, coenzyme Q) is an essential electron transfer lipid in the mitochondrial respiratory chain. It is a main source of mitochondrial reactive oxygen species (ROS) but also has antioxidant properties. This mix of characteristics is why ubiquinone supplementation is considered a potential therapy for many diseases involving mitochondrial dysfunction. Mutations in the ubiquinone biosynthetic pathway are increasingly being identified in patients. Furthermore, secondary ubiquinone deficiency is a common finding associated with mitochondrial disorders and might exacerbate these conditions. Recent developments have suggested that ubiquinone biosynthesis occurs in discrete domains of the mitochondrial inner membrane close to ER-mitochondria contact sites. This spatial requirement for ubiquinone biosynthesis could be the link between secondary ubiquinone deficiency and mitochondrial dysfunction, which commonly results in loss of mitochondrial structural integrity.
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Affiliation(s)
- Ying Wang
- Department of Biology, McGill University, Montreal, Canada
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Li J, Zhou H, Pan X, Li Z, Lu Y, He N, Meng T, Yao C, Chen C, Ling X. The role of fluconazole in the regulation of fatty acid and unsaponifiable matter biosynthesis in Schizochytrium sp. MYA 1381. BMC Microbiol 2019; 19:256. [PMID: 31729956 PMCID: PMC6858700 DOI: 10.1186/s12866-019-1622-4] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2018] [Accepted: 10/23/2019] [Indexed: 12/01/2022] Open
Abstract
Background Schizochytrium has been widely used in industry for synthesizing polyunsaturated fatty acids (PUFAs), especially docosahexaenoic acid (DHA). However, unclear biosynthesis pathway of PUFAs inhibits further production of the Schizochytrium. Unsaponifiable matter (UM) from mevalonate pathway is crucial to cell growth and intracellular metabolism in all higher eukaryotes and microalgae. Therefore, regulation of UM biosynthesis in Schizochytrium may have important effects on fatty acids synthesis. Moreover, it is well known that UMs, such as squalene and β-carotene, are of great commercial value. Thus, regulating UM biosynthesis may also allow for an increased valuation of Schizochytrium. Results To investigate the correlation of UM biosynthesis with fatty acids accumulation in Schizochytrium, fluconazole was used to block the sterols pathway. The addition of 60 mg/L fluconazole at 48 h increased the total lipids (TLs) at 96 h by 16% without affecting cell growth, which was accompanied by remarkable changes in UMs and NADPH. Cholesterol content was reduced by 8%, and the squalene content improved by 45% at 72 h, which demonstrated fluconazole’s role in inhibiting squalene flow to cholesterol. As another typical UM with antioxidant capacity, the β-carotene production was increased by 53% at 96 h. The increase of squalene and β-carotene could boost intracellular oxidation resistance to protect fatty acids from oxidation. The NADPH was found to be 33% higher than that of the control at 96 h, which meant that the cells had more reducing power for fatty acid synthesis. Metabolic analysis further confirmed that regulation of sterols was closely related to glucose absorption, pigment biosynthesis and fatty acid production in Schizochytrium. Conclusion This work first reported the effect of UM biosynthesis on fatty acid accumulation in Schizochytrium. The UM was found to affect fatty acid biosynthesis by changing cell membrane function, intracellular antioxidation and reducing power. We believe that this work provides valuable insights in improving PUFA and other valuable matters in microalgae.
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Affiliation(s)
- Jun Li
- Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China
| | - Hao Zhou
- Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China
| | - Xueshan Pan
- Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China
| | - Zhipeng Li
- Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China
| | - Yinghua Lu
- Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China.,Fujian Collaborative Innovation Center for Exploitation and Utilization of Marine Biological Resources, Xiamen, Fujian, People's Republic of China
| | - Ning He
- Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China.,The Key Lab for Synthetic Biotechnology of Xiamen City, Xiamen University, Xiamen, 361005, People's Republic of China
| | - Tong Meng
- Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China
| | - Chuanyi Yao
- Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China.,The Key Lab for Synthetic Biotechnology of Xiamen City, Xiamen University, Xiamen, 361005, People's Republic of China
| | - Cuixue Chen
- Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China
| | - Xueping Ling
- Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, 361005, People's Republic of China. .,The Key Lab for Synthetic Biotechnology of Xiamen City, Xiamen University, Xiamen, 361005, People's Republic of China.
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Verdaguer IB, Zafra CA, Crispim M, Sussmann RA, Kimura EA, Katzin AM. Prenylquinones in Human Parasitic Protozoa: Biosynthesis, Physiological Functions, and Potential as Chemotherapeutic Targets. Molecules 2019; 24:molecules24203721. [PMID: 31623105 PMCID: PMC6832408 DOI: 10.3390/molecules24203721] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2019] [Revised: 09/25/2019] [Accepted: 10/01/2019] [Indexed: 12/19/2022] Open
Abstract
Human parasitic protozoa cause a large number of diseases worldwide and, for some of these diseases, there are no effective treatments to date, and drug resistance has been observed. For these reasons, the discovery of new etiological treatments is necessary. In this sense, parasitic metabolic pathways that are absent in vertebrate hosts would be interesting research candidates for the identification of new drug targets. Most likely due to the protozoa variability, uncertain phylogenetic origin, endosymbiotic events, and evolutionary pressure for adaptation to adverse environments, a surprising variety of prenylquinones can be found within these organisms. These compounds are involved in essential metabolic reactions in organisms, for example, prevention of lipoperoxidation, participation in the mitochondrial respiratory chain or as enzymatic cofactors. This review will describe several prenylquinones that have been previously characterized in human pathogenic protozoa. Among all existing prenylquinones, this review is focused on ubiquinone, menaquinone, tocopherols, chlorobiumquinone, and thermoplasmaquinone. This review will also discuss the biosynthesis of prenylquinones, starting from the isoprenic side chains to the aromatic head group precursors. The isoprenic side chain biosynthesis maybe come from mevalonate or non-mevalonate pathways as well as leucine dependent pathways for isoprenoid biosynthesis. Finally, the isoprenic chains elongation and prenylquinone aromatic precursors origins from amino acid degradation or the shikimate pathway is reviewed. The phylogenetic distribution and what is known about the biological functions of these compounds among species will be described, as will the therapeutic strategies associated with prenylquinone metabolism in protozoan parasites.
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Affiliation(s)
- Ignasi B. Verdaguer
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
| | - Camila A. Zafra
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
| | - Marcell Crispim
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
| | - Rodrigo A.C. Sussmann
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
- Centro de Formação em Ciências Ambientais, Universidade Federal do Sul da Bahia, Porto Seguro 45810-000 Bahia, Brazil
| | - Emília A. Kimura
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
| | - Alejandro M. Katzin
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
- Correspondence: ; Tel.: +55-11-3091-7330; Fax: +5511-3091-7417
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Díaz-Casado ME, Quiles JL, Barriocanal-Casado E, González-García P, Battino M, López LC, Varela-López A. The Paradox of Coenzyme Q 10 in Aging. Nutrients 2019; 11:nu11092221. [PMID: 31540029 PMCID: PMC6770889 DOI: 10.3390/nu11092221] [Citation(s) in RCA: 50] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2019] [Revised: 09/06/2019] [Accepted: 09/08/2019] [Indexed: 12/14/2022] Open
Abstract
Coenzyme Q (CoQ) is an essential endogenously synthesized molecule that links different metabolic pathways to mitochondrial energy production thanks to its location in the mitochondrial inner membrane and its redox capacity, which also provide it with the capability to work as an antioxidant. Although defects in CoQ biosynthesis in human and mouse models cause CoQ deficiency syndrome, some animals models with particular defects in the CoQ biosynthetic pathway have shown an increase in life span, a fact that has been attributed to the concept of mitohormesis. Paradoxically, CoQ levels decline in some tissues in human and rodents during aging and coenzyme Q10 (CoQ10) supplementation has shown benefits as an anti-aging agent, especially under certain conditions associated with increased oxidative stress. Also, CoQ10 has shown therapeutic benefits in aging-related disorders, particularly in cardiovascular and metabolic diseases. Thus, we discuss the paradox of health benefits due to a defect in the CoQ biosynthetic pathway or exogenous supplementation of CoQ10.
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Affiliation(s)
- M Elena Díaz-Casado
- Institute of Biotechnology, Department of Physiology, Biomedical Research Center, University of Granada, Avda del Conocimiento sn, 18016 Granada, Spain.
- Centro de Investigación Biomédica en Red de Fragilidad y Envejecimiento Saludable (CIBERFES), 18016 Granada, Spain.
| | - José L Quiles
- Institute of Nutrition and Food Technology "José Mataix Verdú", Department of Physiology, Biomedical Research Center, University of Granada, Avda del Conocimiento sn, 18016 Granada, Spain.
| | - Eliana Barriocanal-Casado
- Institute of Biotechnology, Department of Physiology, Biomedical Research Center, University of Granada, Avda del Conocimiento sn, 18016 Granada, Spain.
- Centro de Investigación Biomédica en Red de Fragilidad y Envejecimiento Saludable (CIBERFES), 18016 Granada, Spain.
| | - Pilar González-García
- Institute of Biotechnology, Department of Physiology, Biomedical Research Center, University of Granada, Avda del Conocimiento sn, 18016 Granada, Spain.
- Centro de Investigación Biomédica en Red de Fragilidad y Envejecimiento Saludable (CIBERFES), 18016 Granada, Spain.
| | - Maurizio Battino
- Department of Clinical Sicences, Università Politecnica delle Marche, 60131 Ancona, Italy.
- Nutrition and Food Science Group, Department of Analytical and Food Chemistry, CITACA, CACTI, University of Vigo, 36310 Vigo, Spain.
- International Research Center for Food Nutrition and Safety, Jiangsu University, Zhenjiang 212013, China.
| | - Luis C López
- Institute of Biotechnology, Department of Physiology, Biomedical Research Center, University of Granada, Avda del Conocimiento sn, 18016 Granada, Spain.
- Centro de Investigación Biomédica en Red de Fragilidad y Envejecimiento Saludable (CIBERFES), 18016 Granada, Spain.
| | - Alfonso Varela-López
- Institute of Nutrition and Food Technology "José Mataix Verdú", Department of Physiology, Biomedical Research Center, University of Granada, Avda del Conocimiento sn, 18016 Granada, Spain.
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Choo HJ, Ahn JH. Synthesis of Three Bioactive Aromatic Compounds by Introducing Polyketide Synthase Genes into Engineered Escherichia coli. JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY 2019; 67:8581-8589. [PMID: 31321975 DOI: 10.1021/acs.jafc.9b03439] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/10/2023]
Abstract
Intermediates in aromatic amino acid biosynthesis can serve as substrates for the synthesis of bioactive compounds. In this study we used two intermediates in the shikimate pathway of Escherichia coli, chorismate and anthranilate, to synthesize three bioactive compounds: 4-hydroxycoumarin (4-HC), 2,4-dihydroxyquinoline (DHQ), and 4-hydroxy-1-methyl-2(1H)-quinolone (NMQ). We introduced genes for the synthesis of salicylic acid from chorismate to supply the substrate for 4-HC and the gene encoding N-methyltransferase for the synthesis of N-methylanthranilate from anthranilate. Polyketide synthases and coenzyme (Co)A ligases were tested to determine the optimal combination of genes for the synthesis of each compound. We also tested several constructs and identified the best one for increasing levels of endogenous substrates for chorismate, anthranilate, and malonyl-CoA. With the use of these strategies, 255.4 mg/L 4-HC, 753.7 mg/L DHQ, and 17.5 mg/L NMQ were synthesized. This work provides a basis for the synthesis of diverse coumarin and quinoline derivatives with potential medical applications.
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Affiliation(s)
- Hye Jeong Choo
- Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center , Konkuk University , Seoul 05029 , Republic of Korea
| | - Joong-Hoon Ahn
- Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center , Konkuk University , Seoul 05029 , Republic of Korea
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40
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Payer SE, Faber K, Glueck SM. Non-Oxidative Enzymatic (De)Carboxylation of (Hetero)Aromatics and Acrylic Acid Derivatives. Adv Synth Catal 2019; 361:2402-2420. [PMID: 31379472 PMCID: PMC6644310 DOI: 10.1002/adsc.201900275] [Citation(s) in RCA: 43] [Impact Index Per Article: 7.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2019] [Revised: 04/16/2019] [Indexed: 12/20/2022]
Abstract
The utilization of carbon dioxide as a C1-building block for the production of valuable chemicals has recently attracted much interest. Whereas chemical CO2 fixation is dominated by C-O and C-N bond forming reactions, the development of novel concepts for the carboxylation of C-nucleophiles, which leads to the formation of carboxylic acids, is highly desired. Beside transition metal catalysis, biocatalysis has emerged as an attractive method for the highly regioselective (de)carboxylation of electron-rich (hetero)aromatics, which has been recently further expanded to include conjugated α,β-unsaturated (acrylic) acid derivatives. Depending on the type of substrate, different classes of enzymes have been explored for (i) the ortho-carboxylation of phenols catalyzed by metal-dependent ortho-benzoic acid decarboxylases and (ii) the side-chain carboxylation of para-hydroxystyrenes mediated by metal-independent phenolic acid decarboxylases. Just recently, the portfolio of bio-carboxylation reactions was complemented by (iii) the para-carboxylation of phenols and the decarboxylation of electron-rich heterocyclic and acrylic acid derivatives mediated by prenylated FMN-dependent decarboxylases, which is the main focus of this review. Bio(de)carboxylation processes proceed under physiological reaction conditions employing bicarbonate or (pressurized) CO2 when running in the energetically uphill carboxylation direction. Aiming to facilitate the application of these enzymes in preparative-scale biotransformations, their catalytic mechanism and substrate scope are analyzed in this review.
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Affiliation(s)
- Stefan E. Payer
- Institute of ChemistryUniversity of GrazHeinrichstrasse 288010GrazAustria
| | - Kurt Faber
- Institute of ChemistryUniversity of GrazHeinrichstrasse 288010GrazAustria
| | - Silvia M. Glueck
- Institute of ChemistryUniversity of GrazHeinrichstrasse 288010GrazAustria
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Mahdinia E, Demirci A, Berenjian A. Biofilm reactors as a promising method for vitamin K (menaquinone-7) production. Appl Microbiol Biotechnol 2019; 103:5583-5592. [PMID: 31152205 DOI: 10.1007/s00253-019-09913-w] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/20/2019] [Revised: 05/15/2019] [Accepted: 05/15/2019] [Indexed: 12/17/2022]
Abstract
Menaquinone-7 (MK-7) is the most potent subtype of vitamin K with extraordinarily high half-life in the circulatory system. Therefore, MK-7 plays a critical role in promoting human wellbeing today. Studies on MK-7 every year show more and more magnificent benefits of it in preventing cardiovascular diseases and osteoporosis to battling cancer cells, Alzheimer's and Parkinson's diseases. Thus, it needs to be supplemented to daily diet for accumulative and long-term benefits. Chemical synthesis of MK-7 produces a significant cis-isomer form of it, which has no biological activity. Fortunately, due to its key role in electron transfer in bacteria, trans-MK-7 is biosynthesized by especially Gram-positive strains mainly Bacillus genus. Concordantly, MK-7 could be produced via solid or liquid state fermentation strategies. In either regime, when static fermentation is applied in the absence of agitation and aeration, operational issues arise such as heat and mass transfer inefficiencies. Thus, scaling up the process becomes a challenge. On the other hand, studies have indicated that biofilm and pellicle formation that occur in static fermentations are key characteristics for extracellular MK-7 secretion. Therefore, this review covers the most recent discoveries of the therapeutic properties of MK-7 and optimization attempts at increasing its biosynthesis in different media compositions and effective growth parameters as well as the cutting-edge use of biofilm reactors where B. subtilis cells have the infrastructures to form mature biofilm formations on plastic composite supports. Biofilm reactors therefore can provide robust extracellular MK-7 secretion while simultaneously enduring high agitation and aeration rates, which then address the scale-up and operational issues associated with static fermentation strategies.
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Affiliation(s)
- Ehsan Mahdinia
- Department of Agricultural and Biological Engineering, The Pennsylvania State University, State College, PA, USA
| | - Ali Demirci
- Department of Agricultural and Biological Engineering, The Pennsylvania State University, State College, PA, USA. .,The Huck Institutes of Life Sciences, The Pennsylvania State University, University Park, PA, 16802, USA.
| | - Aydin Berenjian
- Faculty of Science and Engineering, The University of Waikato, Hamilton, 3240, New Zealand
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Huccetogullari D, Luo ZW, Lee SY. Metabolic engineering of microorganisms for production of aromatic compounds. Microb Cell Fact 2019; 18:41. [PMID: 30808357 PMCID: PMC6390333 DOI: 10.1186/s12934-019-1090-4] [Citation(s) in RCA: 117] [Impact Index Per Article: 19.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/15/2019] [Accepted: 02/19/2019] [Indexed: 01/09/2023] Open
Abstract
Metabolic engineering has been enabling development of high performance microbial strains for the efficient production of natural and non-natural compounds from renewable non-food biomass. Even though microbial production of various chemicals has successfully been conducted and commercialized, there are still numerous chemicals and materials that await their efficient bio-based production. Aromatic chemicals, which are typically derived from benzene, toluene and xylene in petroleum industry, have been used in large amounts in various industries. Over the last three decades, many metabolically engineered microorganisms have been developed for the bio-based production of aromatic chemicals, many of which are derived from aromatic amino acid pathways. This review highlights the latest metabolic engineering strategies and tools applied to the biosynthesis of aromatic chemicals, many derived from shikimate and aromatic amino acids, including L-phenylalanine, L-tyrosine and L-tryptophan. It is expected that more and more engineered microorganisms capable of efficiently producing aromatic chemicals will be developed toward their industrial-scale production from renewable biomass.
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Affiliation(s)
- Damla Huccetogullari
- Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus Program) and Institute for the BioCentury, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, 34141, Republic of Korea
- Systems Metabolic Engineering and Systems Healthcare Cross-Generation Collaborative Laboratory, KAIST, Daejeon, 34141, Republic of Korea
| | - Zi Wei Luo
- Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus Program) and Institute for the BioCentury, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, 34141, Republic of Korea
- Systems Metabolic Engineering and Systems Healthcare Cross-Generation Collaborative Laboratory, KAIST, Daejeon, 34141, Republic of Korea
| | - Sang Yup Lee
- Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus Program) and Institute for the BioCentury, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, 34141, Republic of Korea.
- Systems Metabolic Engineering and Systems Healthcare Cross-Generation Collaborative Laboratory, KAIST, Daejeon, 34141, Republic of Korea.
- BioProcess Engineering Research Center and Bioinformatics Research Center, KAIST, Daejeon, 34141, Republic of Korea.
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Using a Chemical Genetic Screen to Enhance Our Understanding of the Antimicrobial Properties of Gallium against Escherichia coli. Genes (Basel) 2019; 10:genes10010034. [PMID: 30634525 PMCID: PMC6356860 DOI: 10.3390/genes10010034] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2018] [Revised: 12/17/2018] [Accepted: 01/04/2019] [Indexed: 12/13/2022] Open
Abstract
The diagnostic and therapeutic agent gallium offers multiple clinical and commercial uses including the treatment of cancer and the localization of tumors, among others. Further, this metal has been proven to be an effective antimicrobial agent against a number of microbes. Despite the latter, the fundamental mechanisms of gallium action have yet to be fully identified and understood. To further the development of this antimicrobial, it is imperative that we understand the mechanisms by which gallium interacts with cells. As a result, we screened the Escherichia coli Keio mutant collection as a means of identifying the genes that are implicated in prolonged gallium toxicity or resistance and mapped their biological processes to their respective cellular system. We discovered that the deletion of genes functioning in response to oxidative stress, DNA or iron–sulfur cluster repair, and nucleotide biosynthesis were sensitive to gallium, while Ga resistance comprised of genes involved in iron/siderophore import, amino acid biosynthesis and cell envelope maintenance. Altogether, our explanations of these findings offer further insight into the mechanisms of gallium toxicity and resistance in E. coli.
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Shukla S, Dubey KK. CoQ10 a super-vitamin: review on application and biosynthesis. 3 Biotech 2018; 8:249. [PMID: 29755918 DOI: 10.1007/s13205-018-1271-6] [Citation(s) in RCA: 37] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2018] [Accepted: 04/30/2018] [Indexed: 12/13/2022] Open
Abstract
Coenzyme Q10 (CoQ) or ubiquinone is found in the biological system which is synthesized by the conjugation of benzoquinone ring with isoprenoid chain of variable length. Coenzyme Q10 supplementation energizes the body and increases body energy production in the form of ATP and helps to treat various human diseases such as cardiomyopathy, muscular dystrophy, periodontal disease, etc. Reports of these potential therapeutic advantages of CoQ10 have resulted in its high market demand, which focus the researchers to work on this molecule and develop better bioprocess methods for commercial level production. At the moment, chemical synthesis, semi-synthetic method as well as bio-production utilizing microbes as biofactory are in use for the synthesis of CoQ10. Chemical synthesis involves use of cheap and easily available precursor molecules such as isoprenol, chloromethylquinone, vinylalane, and solanesol. Chemical synthesis methods due to the use of various solvents and chemicals are less feasible, which limits its application. The microbial production of CoQ10 has added advantages of being produced in optically pure form with high yield using inexpensive medium composition. Several bacteria, e.g., Agrobacterium, Paracoccus, Rhodobacterium, and yeast such as Candida, Rhodotorula are the potent ubiquinone producer. Some alternative biosynthetic pathway for designing of CoQ10 production coupled with metabolic engineering might help to increase CoQ10 production. The most common practiced strategy for strain development for commercial CoQ10 production is through natural isolation and chemical mutagenesis. Here, we have reviewed the chemical, semi-synthetic as well as microbial CoQ10 production in detail.
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Affiliation(s)
- Shraddha Shukla
- Bioprocess Engineering Laboratory, Department of Biotechnology, Central University of Haryana, Mahendergarh, Haryana 123031 India
| | - Kashyap Kumar Dubey
- Bioprocess Engineering Laboratory, Department of Biotechnology, Central University of Haryana, Mahendergarh, Haryana 123031 India
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Wang PH, Khusnutdinova AN, Luo F, Xiao J, Nemr K, Flick R, Brown G, Mahadevan R, Edwards EA, Yakunin AF. Biosynthesis and Activity of Prenylated FMN Cofactors. Cell Chem Biol 2018; 25:560-570.e6. [PMID: 29551348 DOI: 10.1016/j.chembiol.2018.02.007] [Citation(s) in RCA: 39] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2017] [Revised: 12/06/2017] [Accepted: 02/07/2018] [Indexed: 10/17/2022]
Abstract
Prenylated flavin mononucleotide (prFMN) is a recently discovered cofactor required by the UbiD family of reversible decarboxylases involved in ubiquinone biosynthesis, biological decomposition of lignin, and biotransformation of aromatic compounds. This cofactor is synthesized by UbiX-like prenyltransferases catalyzing the transfer of the dimethylallyl moiety of dimethylallyl-monophosphate (DMAP) to FMN. The origin of DMAP for prFMN biosynthesis and the biochemical properties of free prFMN are unknown. We show that in Escherichia coli cells, DMAP can be produced by phosphorylating prenol using ThiM or dephosphorylating DMAPP using Nudix hydrolases. We produced 14 active prenyltransferases whose properties enabled the purification and characterization of protein-free forms of prFMN. In vitro assays revealed that the UbiD-like ferulate decarboxylase (Fdc1) can be activated by free prFMNiminium or C2'-hydroxylated prFMNiminium under both oxidized and reduced conditions. These insights into the biosynthesis and properties of prFMN will facilitate further elucidation of the biochemical diversity of reversible UbiD (de)carboxylases.
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Affiliation(s)
- Po-Hsiang Wang
- Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, ON M5S 3E5, Canada
| | - Anna N Khusnutdinova
- Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, ON M5S 3E5, Canada
| | - Fei Luo
- Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, ON M5S 3E5, Canada
| | - Johnny Xiao
- Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, ON M5S 3E5, Canada
| | - Kayla Nemr
- Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, ON M5S 3E5, Canada
| | - Robert Flick
- Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, ON M5S 3E5, Canada
| | - Greg Brown
- Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, ON M5S 3E5, Canada
| | - Radhakrishnan Mahadevan
- Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, ON M5S 3E5, Canada
| | - Elizabeth A Edwards
- Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, ON M5S 3E5, Canada; Department of Cell and Systems Biology, University of Toronto, 25 Harbord Street, Toronto, ON M5S 3E5, Canada.
| | - Alexander F Yakunin
- Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, ON M5S 3E5, Canada.
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Abstract
Prenylquinones are isoprenoid compounds with a characteristic quinone structure and isoprenyl tail that are ubiquitous in almost all living organisms. There are four major prenylquinone classes: ubiquinone (UQ), menaquinone (MK), plastoquinone (PQ), and rhodoquinone (RQ). The quinone structure and isoprenyl tail length differ among organisms. UQ, PQ, and RQ contain benzoquinone, while MK contains naphthoquinone. UQ, MK, and RQ are involved in oxidative phosphorylation, while PQ functions in photosynthetic electron transfer. Some organisms possess two types of prenylquinones; Escherichia coli has UQ8 and MK8, and Caenorhabditis elegans has UQ9 and RQ9. Crystal structures of most of the enzymes involved in MK synthesis have been solved. Studies on the biosynthesis and functions of quinones have advanced recently, including for phylloquinone (PhQ), which has a phytyl moiety instead of an isoprenyl tail. Herein, the synthesis and applications of prenylquinones are reviewed.
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Affiliation(s)
- Makoto Kawamukai
- a Department of Life Science and Biotechnology, Faculty of Life and Environmental Science , Shimane University , Matsue , Japan
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47
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Liu S, Tun HM, Leung FC, Bennett DC, Zhang H, Cheng KM. Interaction of genotype and diet on small intestine microbiota of Japanese quail fed a cholesterol enriched diet. Sci Rep 2018; 8:2381. [PMID: 29402949 PMCID: PMC5799165 DOI: 10.1038/s41598-018-20508-9] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2017] [Accepted: 01/18/2018] [Indexed: 02/06/2023] Open
Abstract
Our previous study has shown that genetic selection for susceptibility/resistance to diet-induced atherosclerosis has affected the Japanese quail's cecal environment to accommodate distinctly different cecal microbiota. In this study, we fed the Atherosclerosis-resistant (RES) and -susceptable (SUS) quail a regular and a cholesterol enriched diet to examine the interaction of host genotype and diet on the diversity, composition, and metabolic functions of the duodenal and ileal microbiota with relations to atherosclerosis development. In the duodenal content, 9 OTUs (operational taxonomic units) were identified whose abundance had significant positive correlations with plasma total cholesterol, LDL level and/or LDL/HDL ratio. In the ileal content, 7 OTUs have significant correlation with plasma HDL. Cholesterol fed RES hosted significantly less Escherichia and unclassified Enterobacteriaceae (possibly pathogenic) in their duodenum than SUS fed the same diet. Dietary cholesterol significantly decreased the duodenal microbiome of SUS's biosynthesis of Ubiquinone and other terpenoid-quinone. Cholesterol fed RES had significantly more microbiome genes for Vitamin B6, selenocompound, taurine and hypotaurine, and Linoleic acid metabolism; Bisphenol degradation; primary bile acid, and butirosin and neomycin biosynthesis than SUS on the same diet. Microbiome in the ileum and ceca of RES contributed significantly towards the resistance to diet induced atherosclerosis.
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Affiliation(s)
- Shasha Liu
- The State Key Laboratory of Animal Nutrition, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing, China
- Avian Research Centre, Faculty of Land and Food Systems, University of British Columbia, Vancouver, British Columbia, Canada
| | - Hein Min Tun
- School of Biological Sciences, Faculty of Science, University of Hong Kong, Hong Kong SAR, China
- Department of Pediatrics, University of Alberta, Alberta, Canada
| | - Frederick C Leung
- School of Biological Sciences, Faculty of Science, University of Hong Kong, Hong Kong SAR, China
| | - Darin C Bennett
- Avian Research Centre, Faculty of Land and Food Systems, University of British Columbia, Vancouver, British Columbia, Canada
- Animal Science Department, California Polytechnic State University, San Luis Obispo, California, USA
| | - Hongfu Zhang
- The State Key Laboratory of Animal Nutrition, Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing, China.
| | - Kimberly M Cheng
- Avian Research Centre, Faculty of Land and Food Systems, University of British Columbia, Vancouver, British Columbia, Canada.
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48
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Arunrattanamook N, Marsh ENG. Kinetic Characterization of Prenyl-Flavin Synthase from Saccharomyces cerevisiae. Biochemistry 2017; 57:696-700. [PMID: 29232110 DOI: 10.1021/acs.biochem.7b01131] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
We have characterized the kinetics and substrate requirements of prenyl-flavin synthase from yeast. This enzyme catalyzes the addition of an isopentenyl unit to reduced flavin mononucleotide (FMN) to form an additional six-membered ring that bridges N5 and C6 of the flavin nucleus, thereby converting the flavin from a redox cofactor to one that supports the decarboxylation of aryl carboxylic acids. In contrast to bacterial enzymes, the yeast enzyme was found to use dimethylallyl pyrophosphate, rather than dimethylallyl phosphate, as the prenyl donor in the reaction. We developed a coupled assay for prenyl-flavin synthase activity in which turnover was linked to the activation of the prenyl-flavin-dependent enzyme, ferulic acid decarboxylase. The kinetics of the reaction are extremely slow: kcat = 12.2 ± 0.2 h-1, and KM for dimethylallyl pyrophosphate = 9.8 ± 0.7 μM. The KM for reduced FMN was too low to be accurately measured. The kinetics of reduced FMN consumption were studied under pre-steady state conditions. The reaction of FMN was described well by first-order kinetics with a kapp of 17.4 ± 1.1 h-1. These results indicate that a chemical step, most likely formation of the carbon-carbon bond between C6 of the flavin and the isopentenyl moiety, is substantially rate-determining in the reaction.
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Affiliation(s)
- Nattapol Arunrattanamook
- Department of Chemistry, ‡Department of Chemical Engineering, and §Department of Biological Chemistry, University of Michigan , Ann Arbor, Michigan 48109, United States
| | - E Neil G Marsh
- Department of Chemistry, ‡Department of Chemical Engineering, and §Department of Biological Chemistry, University of Michigan , Ann Arbor, Michigan 48109, United States
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49
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Zhang BB, Hu PF, Huang J, Hu YD, Chen L, Xu GR. Current Advances on the Structure, Bioactivity, Synthesis, and Metabolic Regulation of Novel Ubiquinone Derivatives in the Edible and Medicinal Mushroom Antrodia cinnamomea. JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY 2017; 65:10395-10405. [PMID: 29125753 DOI: 10.1021/acs.jafc.7b04206] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/07/2023]
Abstract
In recent years, Antrodia cinnamomea has attracted great attention around the world as an extremely precious edible and medicinal mushroom. Ubiquinone derivatives, which are characteristic metabolites of A. cinnamomea, have shown great bioactivities. Some of them have been regarded as promising therapeutic agents and approved into clinical trial by the U.S. Food and Drug Administration. Although some excellent reviews have been published covering different aspects of A. cinnamomea, this review brings, for the first time, complete information about the structure, bioactivity, chemical synthesis, biosynthesis, and metabolic regulation of ubiquinone derivatives in A. cinnamomea. It not only advances our knowledge on the bioactive metabolites, especially the ubiquinone derivatives, in A. cinnamomea but also provides valuable information for the investigation on other edible and medicinal mushrooms.
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Affiliation(s)
- Bo-Bo Zhang
- Key Laboratory of Carbohydrate Chemistry and Biotechnology, School of Biotechnology, Jiangnan University , Wuxi, Jiangsu 214122, People's Republic of China
| | - Peng-Fei Hu
- Key Laboratory of Carbohydrate Chemistry and Biotechnology, School of Biotechnology, Jiangnan University , Wuxi, Jiangsu 214122, People's Republic of China
| | - Jing Huang
- Key Laboratory of Carbohydrate Chemistry and Biotechnology, School of Biotechnology, Jiangnan University , Wuxi, Jiangsu 214122, People's Republic of China
| | - Yong-Dan Hu
- Yunnan Institute of Food Safety, Kunming University of Science and Technology , Kunming, Yunnan 650500, People's Republic of China
| | - Lei Chen
- Key Laboratory of Carbohydrate Chemistry and Biotechnology, School of Biotechnology, Jiangnan University , Wuxi, Jiangsu 214122, People's Republic of China
| | - Gan-Rong Xu
- Key Laboratory of Carbohydrate Chemistry and Biotechnology, School of Biotechnology, Jiangnan University , Wuxi, Jiangsu 214122, People's Republic of China
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50
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Frutos OD, Barriguín G, Lebrero R, Muñoz R. Assessing the influence of the carbon source on the abatement of industrial N 2O emissions coupled with the synthesis of added-value bioproducts. THE SCIENCE OF THE TOTAL ENVIRONMENT 2017; 598:765-771. [PMID: 28456126 DOI: 10.1016/j.scitotenv.2017.04.161] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/28/2017] [Revised: 04/12/2017] [Accepted: 04/20/2017] [Indexed: 06/07/2023]
Abstract
The continuous abatement of a synthetic N2O emission from a nitric acid plant coupled with the simultaneously production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) copolymer and the coenzyme Q10 (CoQ10) in a bubble column reactor (BCR) was tested using methanol, glycerol and a mixture of sodium acetate-acetic acid (Ac-HAc) as a carbon and electron donor source. The BCRs were inoculated with Paracoccus denitrificans and supplied with the carbon/electron donor at a loading rate of 139gCm-3d-1. High N2O removal efficiencies (81-91%) were achieved, with glycerol supporting the highest abatement. The PHBV cell content ranged from 25 to 53%, with highest accumulation in the culture obtained with methanol and Ac-HAc. However, the greatest PHBV productivities were observed in the BCRs operated with glycerol and Ac-HAc (21.7 and 33.5gPHBVm-3d-1, respectively). Glycerol supply induced the highest molar ratio (23%) of the homopolymer 3-hydroxyvalerate in the composition of PHBV. In addition, the specific cell content of CoQ10 ranged from 0.4 to 1mgg-1. This work constitutes, to the best of our knowledge, the first study combining N2O abatement with the simultaneous production of multiple bioproducts, which pave the way to the development of greenhouse gas biorefineries for climate change mitigation.
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Affiliation(s)
- Osvaldo D Frutos
- Department of Chemical Engineering and Environmental Technology, University of Valladolid, Dr. Mergelina, s/n, 47011 Valladolid, Spain; Facultad de Ciencias Agrarias, Universidad Nacional de Asunción, Campus San Lorenzo, Paraguay
| | - Gonzalo Barriguín
- Department of Chemical Engineering and Environmental Technology, University of Valladolid, Dr. Mergelina, s/n, 47011 Valladolid, Spain
| | - Raquel Lebrero
- Department of Chemical Engineering and Environmental Technology, University of Valladolid, Dr. Mergelina, s/n, 47011 Valladolid, Spain
| | - Raúl Muñoz
- Department of Chemical Engineering and Environmental Technology, University of Valladolid, Dr. Mergelina, s/n, 47011 Valladolid, Spain.
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