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Capdevila DA, Rondón JJ, Edmonds KA, Rocchio JS, Dujovne MV, Giedroc DP. Bacterial Metallostasis: Metal Sensing, Metalloproteome Remodeling, and Metal Trafficking. Chem Rev 2024; 124:13574-13659. [PMID: 39658019 DOI: 10.1021/acs.chemrev.4c00264] [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: 12/12/2024]
Abstract
Transition metals function as structural and catalytic cofactors for a large diversity of proteins and enzymes that collectively comprise the metalloproteome. Metallostasis considers all cellular processes, notably metal sensing, metalloproteome remodeling, and trafficking (or allocation) of metals that collectively ensure the functional integrity and adaptability of the metalloproteome. Bacteria employ both protein and RNA-based mechanisms that sense intracellular transition metal bioavailability and orchestrate systems-level outputs that maintain metallostasis. In this review, we contextualize metallostasis by briefly discussing the metalloproteome and specialized roles that metals play in biology. We then offer a comprehensive perspective on the diversity of metalloregulatory proteins and metal-sensing riboswitches, defining general principles within each sensor superfamily that capture how specificity is encoded in the sequence, and how selectivity can be leveraged in downstream synthetic biology and biotechnology applications. This is followed by a discussion of recent work that highlights selected metalloregulatory outputs, including metalloproteome remodeling and metal allocation by metallochaperones to both client proteins and compartments. We close by briefly discussing places where more work is needed to fill in gaps in our understanding of metallostasis.
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Affiliation(s)
- Daiana A Capdevila
- Fundación Instituto Leloir, Instituto de Investigaciones Bioquímicas de Buenos Aires (IIBBA-CONICET), C1405 BWE Buenos Aires, Argentina
| | - Johnma J Rondón
- Fundación Instituto Leloir, Instituto de Investigaciones Bioquímicas de Buenos Aires (IIBBA-CONICET), C1405 BWE Buenos Aires, Argentina
| | - Katherine A Edmonds
- Department of Chemistry, Indiana University, Bloomington, Indiana 47405-7102, United States
| | - Joseph S Rocchio
- Department of Chemistry, Indiana University, Bloomington, Indiana 47405-7102, United States
| | - Matias Villarruel Dujovne
- Fundación Instituto Leloir, Instituto de Investigaciones Bioquímicas de Buenos Aires (IIBBA-CONICET), C1405 BWE Buenos Aires, Argentina
| | - David P Giedroc
- Department of Chemistry, Indiana University, Bloomington, Indiana 47405-7102, United States
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2
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Huang ZS, Tan XQ, Yang HB, Zeng Y, Chen SJ, Wei ZS, Huang YQ. Mechanistic insights into tris(2-chloroisopropyl) phosphate biomineralization coupled with lead (II) biostabilization driven by denitrifying bacteria. THE SCIENCE OF THE TOTAL ENVIRONMENT 2024; 945:173927. [PMID: 38901584 DOI: 10.1016/j.scitotenv.2024.173927] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/11/2024] [Revised: 05/24/2024] [Accepted: 06/09/2024] [Indexed: 06/22/2024]
Abstract
The ubiquity and persistence of organophosphate esters (OPEs) and heavy metal (HMs) pose global environmental risks. This study explored tris(2-chloroisopropyl)phosphate (TCPP) biomineralization coupled to lead (Pb2+) biostabilization driven by denitrifying bacteria (DNB). The domesticated DNB achieved synergistic bioremoval of TCPP and Pb2+ in the batch bioreactor (efficiency: 98 %).TCPP mineralized into PO43- and Cl-, and Pb2+ precipitated with PO43-. The TCPP-degrading/Pb2+-resistant DNB: Achromobacter, Pseudomonas, Citrobacter, and Stenotrophomonas, dominated the bacterial community, and synergized TCPP biomineralization and Pb2+ biostabilization. Metagenomics and metaproteomics revealed TCPP underwent dechlorination, hydrolysis, the TCA cycle-based dissimilation, and assimilation; Pb2+ was detoxified via bioprecipitation, bacterial membrane biosorption, EPS biocomplexation, and efflux out of cells. TCPP, as an initial donor, along with NO3-, as the terminal acceptor, formed a respiratory redox as the primary energy metabolism. Both TCPP and Pb2+ can stimulate phosphatase expression, which established the mutual enhancements between their bioconversions by catalyzing TCPP dephosphorylation and facilitating Pb2+ bioprecipitation. TCPP may alleviate the Pb2+-induced oxidative stress by aiding protein phosphorylation. 80 % of Pb2+ converted into crystalized pyromorphite. These results provide the mechanistic foundations and help develop greener strategies for synergistic bioremediation of OPEs and HMs.
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Affiliation(s)
- Zhen-Shan Huang
- School of Environment, Guangdong Provincial Key Laboratory of Chemical Pollution and Environmental Safety, MOE Key Laboratory of Theoretical Chemistry of Environment, South China Normal University, Guangzhou 510006, China
| | - Xiu-Qin Tan
- State Environmental Protection Key Laboratory of Water Environmental Simulation and Pollution Control, South China Institute of Environmental Sciences, MEE, Guangzhou 510530, China
| | - Han-Biao Yang
- School of Environment, Guangdong Provincial Key Laboratory of Chemical Pollution and Environmental Safety, MOE Key Laboratory of Theoretical Chemistry of Environment, South China Normal University, Guangzhou 510006, China
| | - Yuan Zeng
- School of Environment, Guangdong Provincial Key Laboratory of Chemical Pollution and Environmental Safety, MOE Key Laboratory of Theoretical Chemistry of Environment, South China Normal University, Guangzhou 510006, China
| | - She-Jun Chen
- School of Environment, Guangdong Provincial Key Laboratory of Chemical Pollution and Environmental Safety, MOE Key Laboratory of Theoretical Chemistry of Environment, South China Normal University, Guangzhou 510006, China.
| | - Zai-Shan Wei
- School of Environmental Science and Engineering, Sun Yat-sen University, Guangdong Provincial Key Laboratory of Environmental Pollution Control and Remediation Technology, Guangzhou 510275, China
| | - Yu-Qi Huang
- School of Environment, Guangdong Provincial Key Laboratory of Chemical Pollution and Environmental Safety, MOE Key Laboratory of Theoretical Chemistry of Environment, South China Normal University, Guangzhou 510006, China
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3
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Cho HL, Gullett KL, Fout AR. Synthesis and characterization of tetrapodal nickel complexes with adaptable ligand binding geometries. Chem Commun (Camb) 2024; 60:10564-10567. [PMID: 39229921 DOI: 10.1039/d4cc03186e] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 09/05/2024]
Abstract
This study explores the versatile binding properties of a tetrapodal ligand framework with nickel, demonstrating significant ligand fluxionality through the interconversions of several complexes. Kinetic studies using UV-vis and NMR techniques underscore the pivotal role of solvent coordination in initiating these dynamic processes. A unique reverse-dative Ni → Ag interaction provides another approach in modifying nickel's geometry.
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Affiliation(s)
- Hsien-Liang Cho
- Department of Chemistry, Texas A&M University, College Station, Texas 77843, USA.
| | - Kelly L Gullett
- School of Chemical Sciences, University of Illinois at Urbana-Champaign, 600 S. Mathews Ave., Urbana, Illinois 61801, USA
| | - Alison R Fout
- Department of Chemistry, Texas A&M University, College Station, Texas 77843, USA.
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Zambelli B. Intracellular phase separation and its role in nickel sensing. Trends Cell Biol 2023; 33:732-733. [PMID: 37433710 DOI: 10.1016/j.tcb.2023.06.008] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/22/2023] [Accepted: 06/22/2023] [Indexed: 07/13/2023]
Abstract
Nickel homeostasis in many bacteria is controlled by the nickel-sensor NikR. A recent study by Cao et al. found that Escherichia coli NikR undergoes phase separation and that this event enhances its function as a nickel-dependent transcriptional repressor. The results suggest that phase separation is functional for bacterial metal homeostasis.
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Affiliation(s)
- Barbara Zambelli
- Laboratory of Bioinorganic Chemistry, Department of Pharmacy and Biotechnology, University of Bologna, Bologna, Italy.
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5
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Rodríguez-Meza O, Palomino-Vizcaino G, Quintanar L, Costas M. Mercury ions impact the kinetic and thermal stabilities of human lens γ-crystallins via direct metal-protein interactions. J Inorg Biochem 2023; 242:112159. [PMID: 36827733 DOI: 10.1016/j.jinorgbio.2023.112159] [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: 10/02/2022] [Revised: 02/06/2023] [Accepted: 02/06/2023] [Indexed: 02/17/2023]
Abstract
Loss of metal homeostasis may be involved in several age-related diseases, such as cataracts. Cataracts are caused by the aggregation of lens proteins into light-scattering high molecular weight complexes that impair vision. Environmental exposure to heavy metals, such as mercury, is a risk factor for cataract development. Indeed, mercury ions induce the non-amyloid aggregation of human γC- and γS crystallins, while human γD-crystallin is not sensitive to this metal. Using Differential Scanning Calorimetry (DSC), we evaluate the impact of mercury ions on the kinetic stability of the three most abundant human γ-crystallins. The metal/crystallin interactions were characterized using Isothermal Titration Calorimetry (ITC). Human γD-crystallins exhibited kinetic stabilization due to the presence of mercury ions, despite its thermal stability being decreased. In contrast, human γC- and γS-crystallins are both, thermally and kinetically destabilized by this metal, consistent with their sensitivity to mercury-induced aggregation. The interaction of human γ-crystallins with mercury ions is highly exothermic and complex, since the protein interacts with the metal at more than three sites. The isolated domains of human γ-D and its variant with the H22Q mutation were also studied, revealing the importance of these regions in the mercury-induced stabilization by a direct metal-protein interaction.
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Affiliation(s)
- Oscar Rodríguez-Meza
- Laboratorio de Biofisicoquímica, Departamento de Fisicoquímica, Facultad de Química, Universidad Nacional Autónoma de México, CdMx 04510, Mexico
| | | | - Liliana Quintanar
- Departamento de Química, Centro de Investigación y Estudios Avanzados (Cinvestav), CdMx 07360, Mexico
| | - Miguel Costas
- Laboratorio de Biofisicoquímica, Departamento de Fisicoquímica, Facultad de Química, Universidad Nacional Autónoma de México, CdMx 04510, Mexico.
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6
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How Theoretical Evaluations Can Generate Guidelines for Designing/Engineering Metalloproteins with Desired Metal Affinity and Selectivity. MOLECULES (BASEL, SWITZERLAND) 2022; 28:molecules28010249. [PMID: 36615442 PMCID: PMC9822464 DOI: 10.3390/molecules28010249] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 11/24/2022] [Revised: 12/23/2022] [Accepted: 12/25/2022] [Indexed: 12/29/2022]
Abstract
Almost half of all known proteins contain metal co-factors. Crucial for the flawless performance of a metalloprotein is the selection with high fidelity of the cognate metal cation from the surrounding biological fluids. Therefore, elucidating the factors controlling the metal binding and selectivity in metalloproteins is of particular significance. The knowledge thus acquired not only contributes to better understanding of the intimate mechanism of these events but, also, significantly enriches the researcher's toolbox that could be used in designing/engineering novel metalloprotein structures with pre-programmed properties. A powerful tool in aid of deciphering the physical principles behind the processes of metal recognition and selectivity is theoretical modeling of metal-containing biological structures. This review summarizes recent findings in the field with an emphasis on elucidating the major factors governing these processes. The results from theoretical evaluations are discussed. It is the hope that the physical principles evaluated can serve as guidelines in designing/engineering of novel metalloproteins of interest to both science and industry.
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Baksh KA, Augustine J, Sljoka A, Prosser RS, Zamble DB. Mechanistic insights into the nickel-dependent allosteric response of the Helicobacter pylori NikR transcription factor. J Biol Chem 2022; 299:102785. [PMID: 36502919 PMCID: PMC9860126 DOI: 10.1016/j.jbc.2022.102785] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2022] [Revised: 11/30/2022] [Accepted: 12/03/2022] [Indexed: 12/13/2022] Open
Abstract
In Helicobacter pylori, the nickel-responsive NikR transcription factor plays a key role in regulating intracellular nickel concentrations, which is an essential process for survival of this pathogen in the acidic human stomach. Nickel binding to H. pylori NikR (HpNikR) allosterically activates DNA binding to target promoters encoding genes involved in nickel homeostasis and acid adaptation, to either activate or repress their transcription. We previously showed that HpNikR adopts an equilibrium between an open conformation and DNA-binding competent cis and trans states. Nickel binding slows down conformational exchange between these states and shifts the equilibrium toward the binding-competent states. The protein then becomes stabilized in a cis conformation upon binding the ureA promoter. Here, we investigate how nickel binding creates this response and how it is transmitted to the DNA-binding domains. Through mutagenesis, DNA-binding studies, and computational methods, the allosteric response to nickel was found to be propagated from the nickel-binding sites to the DNA-binding domains via the β-sheets of the metal-binding domain and a network of residues at the inter-domain interface. Our computational results suggest that nickel binding increases protein rigidity to slow down the conformational exchange. A thymine base in the ureA promoter sequence, known to be critical for high affinity DNA binding by HpNikR, was also found to be important for the allosteric response, while a modified version of this promoter further highlighted the importance of the DNA sequence in modulating the response. Collectively, our results provide insights into regulation of a key protein for H. pylori survival.
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Affiliation(s)
- Karina A. Baksh
- Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada
| | - Jerry Augustine
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada
| | - Adnan Sljoka
- RIKEN Center for Advanced Intelligence Project, RIKEN, Chuo-ku, Tokyo, Japan,For correspondence: R. Scott Prosser; Adnan Sljoka
| | - R. Scott Prosser
- Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada,Department of Chemistry, University of Toronto, Toronto, Ontario, Canada,For correspondence: R. Scott Prosser; Adnan Sljoka
| | - Deborah B. Zamble
- Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada,Department of Chemistry, University of Toronto, Toronto, Ontario, Canada
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Choi WW, Jeong H, Kim Y, Lee HS. Gene nceA encodes a Ni/Co-sensing transcription factor to regulate metal efflux in Corynebacterium glutamicum. METALLOMICS : INTEGRATED BIOMETAL SCIENCE 2022; 14:6865361. [PMID: 36460048 DOI: 10.1093/mtomcs/mfac094] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/04/2022] [Accepted: 11/23/2022] [Indexed: 12/04/2022]
Abstract
The function of Corynebacterium glutamicum open reading frame (ORF) NCgl2684 (named nceA in this study), which was annotated to encode a metalloregulator, was assessed using physiological, genetic, and biochemical approaches. Cells with deleted-nceA (ΔnceA) showed a resistant phenotype to NiSO4 and CoSO4 and showed faster growth in minimal medium containing 20 μM NiSO4 or 10 μM CoSO4 than both the wild-type and nceA-overexpressing (P180-nceA) cells. In the ΔnceA strain, the transcription of the downstream-located ORF NCgl2685 (nceB), annotated to encode efflux protein, was increased approximately 4-fold, whereas gene transcription decreased down to 30% level in the P180-nceA strain. The transcriptions of the nceA and nceB genes were stimulated, even when as little as 5 nM NiSO4 was added to the growth medium. Protein NceA was able to bind DNA comprising the promoter region (from -14 to + 18) of the nceA--nceB operon. The protein-DNA interaction was abolished in the presence of 20 μM NiSO4, 50 μM CoSO4, or 50 μM CdSO4. Although manganese induced the transcription of the nceA and nceB genes, it failed to interrupt protein-DNA interaction. Simultaneously, the P180-nceA cells showed increased sensitivity to oxidants such as menadione, hydrogen peroxide, and cumene hydroperoxide, but not diamide. Collectively, our data show that NceA is a nickel- and cobalt-sensing transcriptional regulator that controls the transcription of the probable efflux protein-encoding nceB. The genes are able to suppress intracellular levels of nickel to prevent reactions, which can cause oxidative damage to cellular components.
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Affiliation(s)
- Won-Woo Choi
- Department of Biotechnology and Bioinformatics, Korea University, Sejong 30019, Republic of Korea.,Interdisciplinary Graduate Program for Artificial Intelligence Smart Convergence Technology, Korea University, Sejong 30019, Republic of Korea
| | - Haeri Jeong
- Department of Biotechnology and Bioinformatics, Korea University, Sejong 30019, Republic of Korea
| | - Younhee Kim
- Department of Korean Medicine, Semyung University, Jecheon, Chungbuk 27136, Republic of Korea
| | - Heung-Shick Lee
- Department of Biotechnology and Bioinformatics, Korea University, Sejong 30019, Republic of Korea.,Interdisciplinary Graduate Program for Artificial Intelligence Smart Convergence Technology, Korea University, Sejong 30019, Republic of Korea
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9
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Choi TS, Tezcan FA. Design of a Flexible, Zn-Selective Protein Scaffold that Displays Anti-Irving-Williams Behavior. J Am Chem Soc 2022; 144:18090-18100. [PMID: 36154053 PMCID: PMC9949983 DOI: 10.1021/jacs.2c08050] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Abstract
Selective metal binding is a key requirement not only for the functions of natural metalloproteins but also for the potential applications of artificial metalloproteins in heterogeneous environments such as cells and environmental samples. The selection of transition-metal ions through protein design can, in principle, be achieved through the appropriate choice and the precise positioning of amino acids that comprise the primary metal coordination sphere. However, this task is made difficult by the intrinsic flexibility of proteins and the fact that protein design approaches generally lack the sub-Å precision required for the steric selection of metal ions. We recently introduced a flexible/probabilistic protein design strategy (MASCoT) that allows metal ions to search for optimal coordination geometry within a flexible, yet covalently constrained dimer interface. In an earlier proof-of-principle study, we used MASCoT to generate an artificial metalloprotein dimer, (AB)2, which selectively bound CoII and NiII over CuII (as well as other first-row transition-metal ions) through the imposition of a rigid octahedral coordination geometry, thus countering the Irving-Williams trend. In this study, we set out to redesign (AB)2 to examine the applicability of MASCoT to the selective binding of other metal ions. We report here the design and characterization of a new flexible protein dimer, B2, which displays ZnII selectivity over all other tested metal ions including CuII both in vitro and in cellulo. Selective, anti-Irving-Williams ZnII binding by B2 is achieved through the formation of a unique trinuclear Zn coordination motif in which His and Glu residues are rigidly placed in a tetrahedral geometry. These results highlight the utility of protein flexibility in the design and discovery of selective binding motifs.
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10
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Matilla MA, Velando F, Martín-Mora D, Monteagudo-Cascales E, Krell T. A catalogue of signal molecules that interact with sensor kinases, chemoreceptors and transcriptional regulators. FEMS Microbiol Rev 2021; 46:6356564. [PMID: 34424339 DOI: 10.1093/femsre/fuab043] [Citation(s) in RCA: 70] [Impact Index Per Article: 17.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2021] [Accepted: 08/10/2021] [Indexed: 12/12/2022] Open
Abstract
Bacteria have evolved many different signal transduction systems that sense signals and generate a variety of responses. Generally, most abundant are transcriptional regulators, sensor histidine kinases and chemoreceptors. Typically, these systems recognize their signal molecules with dedicated ligand-binding domains (LBDs), which, in turn, generate a molecular stimulus that modulates the activity of the output module. There are an enormous number of different LBDs that recognize a similarly diverse set of signals. To give a global perspective of the signals that interact with transcriptional regulators, sensor kinases and chemoreceptors, we manually retrieved information on the protein-ligand interaction from about 1,200 publications and 3D structures. The resulting 811 proteins were classified according to the Pfam family into 127 groups. These data permit a delineation of the signal profiles of individual LBD families as well as distinguishing between families that recognize signals in a promiscuous manner and those that possess a well-defined ligand range. A major bottleneck in the field is the fact that the signal input of many signaling systems is unknown. The signal repertoire reported here will help the scientific community design experimental strategies to identify the signaling molecules for uncharacterised sensor proteins.
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Affiliation(s)
- Miguel A Matilla
- Department of Environmental Protection, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, Prof. Albareda 1, 18008 Granada, Spain
| | - Félix Velando
- Department of Environmental Protection, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, Prof. Albareda 1, 18008 Granada, Spain
| | - David Martín-Mora
- Department of Environmental Protection, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, Prof. Albareda 1, 18008 Granada, Spain
| | - Elizabet Monteagudo-Cascales
- Department of Environmental Protection, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, Prof. Albareda 1, 18008 Granada, Spain
| | - Tino Krell
- Department of Environmental Protection, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, Prof. Albareda 1, 18008 Granada, Spain
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11
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Nong Q, Dong H, Liu Y, Liu L, He B, Huang Y, Jiang J, Luan T, Chen B, Hu L. Characterization of the mercury-binding proteins in tuna and salmon sashimi: Implications for health risk of mercury in food. CHEMOSPHERE 2021; 263:128110. [PMID: 33297103 DOI: 10.1016/j.chemosphere.2020.128110] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/18/2020] [Revised: 08/12/2020] [Accepted: 08/21/2020] [Indexed: 06/12/2023]
Abstract
Fish consumption is one of the major ways through which humans receive exposure to mercury (Hg). The existing forms of Hg in food, particularly Hg bound to proteins, may affect the absorption of Hg by humans and subsequently its potentially toxic effects. However, the knowledge regarding Hg-binding proteins in edible fish muscle is scarce. In the present study, salmon and tuna fish muscles, collected from seven different regions and countries, were analyzed using metallomics- and proteomics-based techniques. The concentration of Hg in sashimi samples ranged from 4.4 to 317.4 ng/g. Size exclusion chromatography (SEC) coupled with inductively coupled plasma mass spectrometer (ICP-MS) showed that beta-actin was a novel Hg-binding protein from the fish muscles, and this protein could also bind bismuth (Bi), silver (Ag), and copper (Cu). Hg bound to beta-actin accounted for approximately 30.2-37.6% of the total Hg in the tuna muscles and was significantly correlated to total Hg in the fish muscles (r = 0.98, p < 0.01) and in the fraction of soluble proteins (r = 0.94, p < 0.01). These findings suggest that proteins act as the main Hg accumulation sites in edible fish; thus, increasing human exposure to Hg following gastrointestinal digestion.
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Affiliation(s)
- Qiying Nong
- School of Life Sciences, Sun Yat-Sen University, Guangzhou, 510275, China; State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, 100085, China.
| | - Hongzhe Dong
- State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, 100085, China
| | - Yingqiu Liu
- State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, 100085, China
| | - Lihong Liu
- State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, 100085, China
| | - Bin He
- State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, 100085, China
| | - Yongshun Huang
- Guangdong Provincial Hospital for Occupational Diseases Prevention and Treatment, Guangzhou, 510300, China
| | - Jie Jiang
- Shenzhen Center for Disease Control and Prevention, Shenzhen, 518055, China
| | - Tiangang Luan
- School of Life Sciences, Sun Yat-Sen University, Guangzhou, 510275, China
| | - Baowei Chen
- Southern Marine Science and Engineering Guangdong Laboratory, School of Marine Sciences, Sun Yat-Sen University, Zhuhai, 519082, China.
| | - Ligang Hu
- State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, 100085, China; Institute of Environment and Health, Jianghan University, Wuhan, 430056, China; School of Environment, Hangzhou Institute for Advanced Study, University of Chinese Academy of Sciences, Hangzhou, 310000, China
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12
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Baksh KA, Pichugin D, Prosser RS, Zamble DB. Allosteric regulation of the nickel-responsive NikR transcription factor from Helicobacter pylori. J Biol Chem 2021; 296:100069. [PMID: 33199369 PMCID: PMC7949043 DOI: 10.1074/jbc.ra120.015459] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/30/2020] [Revised: 11/08/2020] [Accepted: 11/15/2020] [Indexed: 12/20/2022] Open
Abstract
Nickel is essential for the survival of the pathogenic bacteria Helicobacter pylori in the fluctuating pH of the human stomach. Due to its inherent toxicity and limited availability, nickel homeostasis is maintained through a network of pathways that are coordinated by the nickel-responsive transcription factor NikR. Nickel binding to H. pylori NikR (HpNikR) induces an allosteric response favoring a conformation that can bind specific DNA motifs, thereby serving to either activate or repress transcription of specific genes involved in nickel homeostasis and acid adaptation. Here, we examine how nickel induces this response using 19F-NMR, which reveals conformational and dynamic changes associated with nickel-activated DNA complex formation. HpNikR adopts an equilibrium between an open state and DNA-binding competent states regardless of nickel binding, but a higher level of dynamics is observed in the absence of metal. Nickel binding shifts the equilibrium toward the binding-competent states and decreases the mobility of the DNA-binding domains. The nickel-bound protein is then able to adopt a single conformation upon binding a target DNA promoter. Zinc, which does not promote high-affinity DNA binding, is unable to induce the same allosteric response as nickel. We propose that the allosteric mechanism of nickel-activated DNA binding by HpNikR is driven by conformational selection.
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Affiliation(s)
- Karina A Baksh
- Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada
| | - Dmitry Pichugin
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada
| | - Robert Scott Prosser
- Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada; Department of Chemistry, University of Toronto, Toronto, Ontario, Canada.
| | - Deborah B Zamble
- Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada; Department of Chemistry, University of Toronto, Toronto, Ontario, Canada
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13
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Harty ML, Sharma AN, Bearne SL. Catalytic properties of the metal ion variants of mandelate racemase reveal alterations in the apparent electrophilicity of the metal cofactor. Metallomics 2020; 11:707-723. [PMID: 30843025 DOI: 10.1039/c8mt00330k] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
Mandalate racemase (MR) from Pseudomonas putida requires a divalent metal cation, usually Mg2+, to catalyse the interconversion of the enantiomers of mandelate. Although the active site Mg2+ may be replaced by Mn2+, Co2+, or Ni2+, substitution by these metal ions does not markedly (<10-fold) alter the kinetic parameters Kappm, kappcat, and (kcat/Km)app for the substrates (R)- and (S)-mandelate, and the alternative substrate (S)-trifluorolactate. Viscosity variation experiments with Mn2+-MR showed that the metal ion plays a role in the uniform binding of the transition states for enzyme-substrate association, the chemical step, and enzyme-product dissociation. Surprisingly, the competitive inhibition constants (Ki) for inhibition of each metalloenzyme variant by benzohydroxamate did not vary significantly with the identity of the metal ion unlike the marked variation of the stability constants (K1) observed for M2+·BzH complex formation in solution. A similar trend was observed for the inhibition of the metalloenzyme variants by F-, except for Mg2+-MR, which bound F- tighter than would be predicted based on the stability constants for formation of M2+·F- complexes in solution. Thus, the enzyme modifies the enatic state of the bound metal ion cofactor so that the apparent electrophilicity of Mg2+ is enhanced, while that of Ni2+ is attenuated, resulting in a levelling effect relative to the trends observed for the free metals in solution.
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Affiliation(s)
- Matthew L Harty
- Department of Biochemistry and Molecular Biology, Dalhousie University, Halifax, NS B3H 4R2, Canada.
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14
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Baksh KA, Zamble DB. Allosteric control of metal-responsive transcriptional regulators in bacteria. J Biol Chem 2020; 295:1673-1684. [PMID: 31857375 PMCID: PMC7008368 DOI: 10.1074/jbc.rev119.011444] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
Abstract
Many transition metals are essential trace nutrients for living organisms, but they are also cytotoxic in high concentrations. Bacteria maintain the delicate balance between metal starvation and toxicity through a complex network of metal homeostasis pathways. These systems are coordinated by the activities of metal-responsive transcription factors-also known as metal-sensor proteins or metalloregulators-that are tuned to sense the bioavailability of specific metals in the cell in order to regulate the expression of genes encoding proteins that contribute to metal homeostasis. Metal binding to a metalloregulator allosterically influences its ability to bind specific DNA sequences through a variety of intricate mechanisms that lie on a continuum between large conformational changes and subtle changes in internal dynamics. This review summarizes recent advances in our understanding of how metal sensor proteins respond to intracellular metal concentrations. In particular, we highlight the allosteric mechanisms used for metal-responsive regulation of several prokaryotic single-component metalloregulators, and we briefly discuss current open questions of how metalloregulators function in bacterial cells. Understanding the regulation and function of metal-responsive transcription factors is a fundamental aspect of metallobiochemistry and is important for gaining insights into bacterial growth and virulence.
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Affiliation(s)
- Karina A Baksh
- Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada
| | - Deborah B Zamble
- Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada; Department of Chemistry, University of Toronto, Toronto, Ontario M5S 3H6, Canada.
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15
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Ha Y, Hu H, Higgins K, Maroney M, Hedman B, Hodgson K, Solomon E. The Electronic Structure of the Metal Active Site Determines the Geometric Structure and Function of the Metalloregulator NikR. Biochemistry 2019; 58:3585-3591. [PMID: 31339709 DOI: 10.1021/acs.biochem.9b00542] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
NikR is a nickel-responsive metalloregulator protein that controls the level of Ni2+ ions in living cells. Previous studies have shown that NikR can bind a series of first-row transition metal ions but binds to DNA with high affinity only as a Ni2+ complex. To understand this metal selectivity, S K-edge X-ray absorption spectroscopy of NikR bound to different metal ions was used to evaluate the different electronic structures. The experimental results are coupled with density functional theory calculations on relevant models. This study shows that both the Zeff of the metal ion and the donor nature of the ligands determine the electronic structure of the metal site. This impacts the geometric structure of the metal site and thus the conformation of the protein. This contribution of electronic structure to geometric structure can be extended to other metal selective metalloregulators.
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Affiliation(s)
- Yang Ha
- Department of Chemistry , Stanford University , Stanford , California 94035 , United States.,Stanford Synchrotron Radiation Lightsource, SLAC , Stanford University , Menlo Park , California 94025 , United States
| | - Heidi Hu
- Department of Chemistry , University of Massachusetts Amherst , Amherst , Massachusetts 01003 , United States
| | - Khadine Higgins
- Department of Chemistry , University of Massachusetts Amherst , Amherst , Massachusetts 01003 , United States
| | - Michael Maroney
- Department of Chemistry , University of Massachusetts Amherst , Amherst , Massachusetts 01003 , United States
| | - Britt Hedman
- Stanford Synchrotron Radiation Lightsource, SLAC , Stanford University , Menlo Park , California 94025 , United States
| | - Keith Hodgson
- Department of Chemistry , Stanford University , Stanford , California 94035 , United States.,Stanford Synchrotron Radiation Lightsource, SLAC , Stanford University , Menlo Park , California 94025 , United States
| | - Edward Solomon
- Department of Chemistry , Stanford University , Stanford , California 94035 , United States.,Stanford Synchrotron Radiation Lightsource, SLAC , Stanford University , Menlo Park , California 94025 , United States
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16
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Abstract
Nickel is essential for the survival of many pathogenic bacteria. E. coli and H. pylori require nickel for [NiFe]-hydrogenases. H. pylori also requires nickel for urease. At high concentrations nickel can be toxic to the cell, therefore, nickel concentrations are tightly regulated. Metalloregulators help to maintain nickel concentration in the cell by regulating the expression of the genes associated with nickel import and export. Nickel import into the cell, delivery of nickel to target proteins, and export of nickel from the cell is a very intricate and well-choreographed process. The delivery of nickel to [NiFe]-hydrogenase and urease is complex and involves several chaperones and accessory proteins. A combination of biochemical, crystallographic, and spectroscopic techniques has been utilized to study the structures of these proteins, as well as protein-protein interactions resulting in an expansion of our knowledge regarding how these proteins sense and bind nickel. In this review, recent advances in the field will be discussed, focusing on the metal site structures of nickel bound to metalloregulators and chaperones.
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17
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Emergence of metal selectivity and promiscuity in metalloenzymes. J Biol Inorg Chem 2019; 24:517-531. [DOI: 10.1007/s00775-019-01667-0] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2019] [Accepted: 05/13/2019] [Indexed: 01/27/2023]
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18
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Metallochaperones and metalloregulation in bacteria. Essays Biochem 2017; 61:177-200. [PMID: 28487396 DOI: 10.1042/ebc20160076] [Citation(s) in RCA: 87] [Impact Index Per Article: 10.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/26/2016] [Revised: 02/23/2017] [Accepted: 02/27/2017] [Indexed: 12/21/2022]
Abstract
Bacterial transition metal homoeostasis or simply 'metallostasis' describes the process by which cells control the intracellular availability of functionally required metal cofactors, from manganese (Mn) to zinc (Zn), avoiding both metal deprivation and toxicity. Metallostasis is an emerging aspect of the vertebrate host-pathogen interface that is defined by a 'tug-of-war' for biologically essential metals and provides the motivation for much recent work in this area. The host employs a number of strategies to starve the microbial pathogen of essential metals, while for others attempts to limit bacterial infections by leveraging highly competitive metals. Bacteria must be capable of adapting to these efforts to remodel the transition metal landscape and employ highly specialized metal sensing transcriptional regulators, termed metalloregulatory proteins,and metallochaperones, that allocate metals to specific destinations, to mediate this adaptive response. In this essay, we discuss recent progress in our understanding of the structural mechanisms and metal specificity of this adaptive response, focusing on energy-requiring metallochaperones that play roles in the metallocofactor active site assembly in metalloenzymes and metallosensors, which govern the systems-level response to metal limitation and intoxication.
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19
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Yoon Y, Kim S, Chae Y, Kang Y, Lee Y, Jeong SW, An YJ. Use of Tunable Whole-Cell Bioreporters to Assess Bioavailable Cadmium and Remediation Performance in Soils. PLoS One 2016; 11:e0154506. [PMID: 27171374 PMCID: PMC4865175 DOI: 10.1371/journal.pone.0154506] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2015] [Accepted: 04/14/2016] [Indexed: 11/18/2022] Open
Abstract
It is important to have tools to measure the bioavailability to assess the risks of pollutants because the bioavailability is defined as the portions of pollutants showing the biological effects on living organisms. This study described the construction of tunable Escherichia coli whole-cell bioreporter (WCB) using the promoter region of zinc-inducible operon and its application on contaminated soils. It was verified that this WCB system showed specific and sensitive responses to cadmium rather than zinc in the experimental conditions. It was inferred that Cd(II) associates stronger with ZntR, a regulatory protein of zinc-inducible operon, than other metal ions. Moreover, the expression of reporter genes, egfp and mcherry, were proportional to the concentration of cadmium, thereby being a quantitative sensor to monitor bioavailable cadmium. The capability to determine bioavailable cadmium was verified with Cd(II) amended LUFA soils, and then the applicability on environmental systems was investigated with field soils collected from smelter area in Korea before and after soil-washing. The total amount of cadmium was decreased after soil washing, while the bioavailability was increased. Consequently, it would be valuable to have tools to assess bioavailability and the effectiveness of soil remediation should be evaluated in the aspect of bioavailability as well as removal efficiency.
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Affiliation(s)
- Youngdae Yoon
- Department of Environmental Health Science, Konkuk University, Seoul 05029, Korea
| | - Sunghoon Kim
- Department of Environmental Health Science, Konkuk University, Seoul 05029, Korea
| | - Yooeun Chae
- Department of Environmental Health Science, Konkuk University, Seoul 05029, Korea
| | - Yerin Kang
- Department of Environmental Health Science, Konkuk University, Seoul 05029, Korea
| | - Youngshim Lee
- Division of Bioscience and Biotechnology, BMIC, Konkuk University, Seoul 05029, Korea
| | - Seung-Woo Jeong
- Department of Environmental Engineering, Kunsan National University, Kunsan 54150, Korea
| | - Youn-Joo An
- Department of Environmental Health Science, Konkuk University, Seoul 05029, Korea
- * E-mail:
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20
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Abstract
This chapter focuses on transition metals. All transition metal cations are toxic-those that are essential for Escherichia coli and belong to the first transition period of the periodic system of the element and also the "toxic-only" metals with higher atomic numbers. Common themes are visible in the metabolism of these ions. First, there is transport. High-rate but low-affinity uptake systems provide a variety of cations and anions to the cells. Control of the respective systems seems to be mainly through regulation of transport activity (flux control), with control of gene expression playing only a minor role. If these systems do not provide sufficient amounts of a needed ion to the cell, genes for ATP-hydrolyzing high-affinity but low-rate uptake systems are induced, e.g., ABC transport systems or P-type ATPases. On the other hand, if the amount of an ion is in surplus, genes for efflux systems are induced. By combining different kinds of uptake and efflux systems with regulation at the levels of gene expression and transport activity, the concentration of a single ion in the cytoplasm and the composition of the cellular ion "bouquet" can be rapidly adjusted and carefully controlled. The toxicity threshold of an ion is defined by its ability to produce radicals (copper, iron, chromate), to bind to sulfide and thiol groups (copper, zinc, all cations of the second and third transition period), or to interfere with the metabolism of other ions. Iron poses an exceptional metabolic problem due its metabolic importance and the low solubility of Fe(III) compounds, combined with the ability to cause dangerous Fenton reactions. This dilemma for the cells led to the evolution of sophisticated multi-channel iron uptake and storage pathways to prevent the occurrence of unbound iron in the cytoplasm. Toxic metals like Cd2+ bind to thiols and sulfide, preventing assembly of iron complexes and releasing the metal from iron-sulfur clusters. In the unique case of mercury, the cation can be reduced to the volatile metallic form. Interference of nickel and cobalt with iron is prevented by the low abundance of these metals in the cytoplasm and their sequestration by metal chaperones, in the case of nickel, or by B12 and its derivatives, in the case of cobalt. The most dangerous metal, copper, catalyzes Fenton-like reactions, binds to thiol groups, and interferes with iron metabolism. E. coli solves this problem probably by preventing copper uptake, combined with rapid efflux if the metal happens to enter the cytoplasm.
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21
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Musiani F, Zambelli B, Bazzani M, Mazzei L, Ciurli S. Nickel-responsive transcriptional regulators. Metallomics 2015; 7:1305-18. [DOI: 10.1039/c5mt00072f] [Citation(s) in RCA: 28] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
The structural features, metal coordination modes and metal binding thermodynamics of known Ni(ii)-dependent transcriptional regulators are highlighted and discussed.
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Affiliation(s)
- Francesco Musiani
- Laboratory of Bioinorganic Chemistry
- Department of Pharmacy and Biotechnology
- University of Bologna
- 40127 Bologna, Italy
| | - Barbara Zambelli
- Laboratory of Bioinorganic Chemistry
- Department of Pharmacy and Biotechnology
- University of Bologna
- 40127 Bologna, Italy
| | - Micaela Bazzani
- Laboratory of Bioinorganic Chemistry
- Department of Pharmacy and Biotechnology
- University of Bologna
- 40127 Bologna, Italy
| | - Luca Mazzei
- Laboratory of Bioinorganic Chemistry
- Department of Pharmacy and Biotechnology
- University of Bologna
- 40127 Bologna, Italy
| | - Stefano Ciurli
- Laboratory of Bioinorganic Chemistry
- Department of Pharmacy and Biotechnology
- University of Bologna
- 40127 Bologna, Italy
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22
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Kim HM, Ahn BE, Lee JH, Roe JH. Regulation of a nickel–cobalt efflux system and nickel homeostasis in a soil actinobacterium Streptomyces coelicolor. Metallomics 2015; 7:702-9. [DOI: 10.1039/c4mt00318g] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/04/2023]
Abstract
In nickel-tolerantStreptomyces coelicolor, a highly nickel-sensitive regulator (Nur) for nickel uptake systems and an extremely insensitive regulator (NmtR) for a nickel efflux pump constitute the nickel homeostasis system.
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Affiliation(s)
- Hae Mi Kim
- School of Biological Sciences, and Institute of Microbiology
- Seoul National University
- Seoul 151-742, Korea
| | - Bo-Eun Ahn
- School of Biological Sciences, and Institute of Microbiology
- Seoul National University
- Seoul 151-742, Korea
| | - Ju-Hyung Lee
- School of Biological Sciences, and Institute of Microbiology
- Seoul National University
- Seoul 151-742, Korea
| | - Jung-Hye Roe
- School of Biological Sciences, and Institute of Microbiology
- Seoul National University
- Seoul 151-742, Korea
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23
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Chivers PT. Cobalt and Nickel. BINDING, TRANSPORT AND STORAGE OF METAL IONS IN BIOLOGICAL CELLS 2014. [DOI: 10.1039/9781849739979-00381] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/30/2023]
Abstract
Cobalt and nickel play key roles in biological systems as cofactors in a small number of important enzymes. The majority of these are found in microbes. Evidence for direct roles for Ni(II) and Co(II) enzymes in higher organisms is limited, with the exception of the well-known requirement for the cobalt-containing vitamin B12 cofactor and the Ni-dependent urease in plants. Nonetheless, nickel in particular plays a key role in human health because of its essential role in microbes that inhabit various growth niches within the body. These roles can be beneficial, as can be seen with the anaerobic production and consumption of H2 in the digestive tract by bacteria and archaea that results in increased yields of short-chain fatty acids. In other cases, nickel has an established role in the establishment of pathogenic infection (Helicobacter pylori urease and colonization of the stomach). The synthesis of Co- and Ni-containing enzymes requires metal import from the extracellular milieu followed by the targeting of these metals to the appropriate protein and enzymes involved in metallocluster or cofactor biosynthesis. These metals are toxic in excess so their levels must be regulated carefully. This complex pathway of metalloenzyme synthesis and intracellular homeostasis requires proteins that can specifically recognize these metals in a hierarchical manner. This chapter focuses on quantitative and structural details of the cobalt and nickel binding sites in transport, trafficking and regulatory proteins involved in cobalt and nickel metabolism in microbes.
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Affiliation(s)
- Peter T. Chivers
- Department of Chemistry, School of Biological and Biomedical Sciences, and Biophysical Sciences Institute, Durham University Durham UK
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24
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Liu Y, Li W, Wei Y, Jiang Y, Tan X. Efficient preparation and metal specificity of the regulatory protein TroR from the human pathogen Treponema pallidum. Metallomics 2014; 5:1448-57. [PMID: 23945957 DOI: 10.1039/c3mt00163f] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
TroR is a putative metal-dependent regulatory protein that has been linked to the virulence of the human pathogen Treponema pallidum. It shares high homology with the well-known iron-dependent regulatory protein DtxR from Corynebacterium diphtheriae, as well as the manganese-dependent MntR from Bacillus subtilis. However, it has been uncertain whether manganese or zinc is the natural cofactor of TroR to date. Herein, we established an efficient method named "double-fusion tagging" to obtain soluble TroR for the first time. A series of studies, including ICP, CD, fluorescence, ITC, and electrophoresis mobility shift assay (EMSA), were performed to resolve the discrepancies in its metal-binding specificity. In addition, bioinformatic analysis as well as mutation studies were carried out to find the genetic relationships of TroR with its homology proteins. In conclusion, our findings indicate that TroR is a manganese-dependent rather than a zinc-dependent regulatory protein.
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Affiliation(s)
- Yi Liu
- Institutes of Biomedical Science, Fudan University, Shanghai 200433, China.
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25
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Foster AW, Pernil R, Patterson CJ, Robinson NJ. Metal specificity of cyanobacterial nickel-responsive repressor InrS: cells maintain zinc and copper below the detection threshold for InrS. Mol Microbiol 2014; 92:797-812. [PMID: 24666373 PMCID: PMC4235346 DOI: 10.1111/mmi.12594] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 03/24/2014] [Indexed: 12/25/2022]
Abstract
InrS is a Ni(II)-responsive, CsoR/RcnR-like, DNA-binding transcriptional repressor of the nrsD gene, but the Ni(II) co-ordination sphere of InrS is unlike Ni(II)-RcnR. We show that copper and Zn(II) also bind tightly to InrS and in vitro these ions also impair InrS binding to the nrsD operator-promoter. InrS does not respond to Zn(II) (or copper) in vivo after 48 h, when Zn(II) sensor ZiaR responds, but InrS transiently responds (1 h) to both metals. InrS conserves only one (of two) second co-ordination shell residues of CsoR (Glu98 in InrS). The allosteric mechanism of InrS is distinct from Cu(I)-CsoR and conservation of deduced second shell residues better predicts metal specificity than do the metal ligands. The allosteric mechanism of InrS permits greater promiscuity in vitro than CsoR. The factors dictating metal-selectivity in vivo are that KNi(II) and ΔGCNi(II)-InrS·DNA are sufficiently high, relative to other metal sensors, for InrS to detect Ni(II), while the equivalent parameters for copper may be insufficient for copper-sensing in S ynechocystis (at 48 h). InrS KZn(II) (5.6 × 10−13 M) is comparable to the sensory sites of ZiaR (and Zur), but ΔGCZn(II)-InrS·DNA is less than ΔGCZn(II)-ZiaR·DNA implying that relative to other sensors, ΔGCZn(II)-Sensor·DNA rather than KZn(II) determines the final detection threshold for Zn(II).
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Affiliation(s)
- Andrew W Foster
- Department of Chemistry, School of Biological and Biomedical Sciences, Durham University, Durham, DH1 3LE, UK
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26
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Kuroda K, Ebisutani K, Iida K, Nishitani T, Ueda M. Enhanced adsorption and recovery of uranyl ions by NikR mutant-displaying yeast. Biomolecules 2014; 4:390-401. [PMID: 24970221 PMCID: PMC4101488 DOI: 10.3390/biom4020390] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/04/2014] [Revised: 03/12/2014] [Accepted: 03/14/2014] [Indexed: 11/25/2022] Open
Abstract
Uranium is one of the most important metal resources, and the technology for the recovery of uranyl ions (UO22+) from aqueous solutions is required to ensure a semi-permanent supply of uranium. The NikR protein is a Ni2+-dependent transcriptional repressor of the nickel-ion uptake system in Escherichia coli, but its mutant protein (NikRm) is able to selectively bind uranyl ions in the interface of the two monomers. In this study, NikRm protein with ability to adsorb uranyl ions was displayed on the cell surface of Saccharomyces cerevisiae. To perform the binding of metal ions in the interface of the two monomers, two metal-binding domains (MBDs) of NikRm were tandemly fused via linker peptides and displayed on the yeast cell surface by fusion with the cell wall-anchoring domain of yeast α-agglutinin. The NikRm-MBD-displaying yeast cells with particular linker lengths showed the enhanced adsorption of uranyl ions in comparison to the control strain. By treating cells with citrate buffer (pH 4.3), the uranyl ions adsorbed on the cell surface were recovered. Our results indicate that the adsorption system by yeast cells displaying tandemly fused MBDs of NikRm is effective for simple and concentrated recovery of uranyl ions, as well as adsorption of uranyl ions.
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Affiliation(s)
- Kouichi Kuroda
- Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
| | - Kazuki Ebisutani
- Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
| | - Katsuya Iida
- Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
| | - Takashi Nishitani
- Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
| | - Mitsuyoshi Ueda
- Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
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27
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Ge RG, Wang DX, Hao MC, Sun XS. Nickel trafficking system responsible for urease maturation in Helicobacter pylori. World J Gastroenterol 2013; 19:8211-8218. [PMID: 24363511 PMCID: PMC3857443 DOI: 10.3748/wjg.v19.i45.8211] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/06/2013] [Revised: 10/17/2013] [Accepted: 11/03/2013] [Indexed: 02/06/2023] Open
Abstract
Helicobacter pylori (H. pylori) is a common human pathogen responsible for various gastric diseases. This bacterium relies on the production of urease and hydrogenase to inhabit the acidic environment of the stomach. Nickel is an essential cofactor for urease and hydrogenase. H. pylori has to uptake sufficient nickel ions for the maturation of urease, and on the other way, to prevent the toxic effects of excessive nickel ions. Therefore, H. pylori has to strike a delicate balance between the import of nickel ions, its efficient intracellular storage, and delivery to nickel-dependent metalloenzymes when required. The assembly and maturation of the urease enzyme is a complex and timely ordered process, requiring various regulatory, uptake, chaperone and accessory proteins. In this review, we focus on several nickel trafficking proteins involved in urease maturation: NikR, NixA, HypAB, UreEFGH, HspA, Hpn and Hpnl. The work will deepen our understanding of how this pathogenic bacterium adapts to severe habitant environments in the host.
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28
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Dudev T, Lim C. Competition among metal ions for protein binding sites: determinants of metal ion selectivity in proteins. Chem Rev 2013; 114:538-56. [PMID: 24040963 DOI: 10.1021/cr4004665] [Citation(s) in RCA: 305] [Impact Index Per Article: 25.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/24/2022]
Affiliation(s)
- Todor Dudev
- Institute of Biomedical Sciences, Academia Sinica , Taipei 11529, Taiwan
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29
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30
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Higgins KA, Carr CE, Maroney MJ. Specific metal recognition in nickel trafficking. Biochemistry 2012; 51:7816-32. [PMID: 22970729 DOI: 10.1021/bi300981m] [Citation(s) in RCA: 46] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Abstract
Nickel is an essential metal for a number of bacterial species that have developed systems for acquiring, delivering, and incorporating the metal into target enzymes and controlling the levels of nickel in cells to prevent toxic effects. As with other transition metals, these trafficking systems must be able to distinguish between the desired metal and other transition metal ions with similar physical and chemical properties. Because there are few enzymes (targets) that require nickel for activity (e.g., Escherichia coli transports nickel for hydrogenases made under anaerobic conditions, and Helicobacter pylori requires nickel for hydrogenase and urease that are essential for acid viability), the "traffic pattern" for nickel is relatively simple, and nickel trafficking therefore presents an opportunity to examine a system for the mechanisms that are used to distinguish nickel from other metals. In this review, we describe the details known for examples of uptake permeases, metallochaperones and proteins involved in metallocenter assembly, and nickel metalloregulators. We also illustrate a variety of mechanisms, including molecular recognition in the case of NikA protein and examples of allosteric regulation for HypA, NikR, and RcnR, employed to generate specific biological responses to nickel ions.
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Affiliation(s)
- Khadine A Higgins
- Department of Chemistry, University of Massachusetts, Amherst, Massachusetts 01003, USA
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31
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Krecisz S, Jones MD, Zamble DB. Nonspecific interactions between Escherichia coli NikR and DNA are critical for nickel-activated DNA binding. Biochemistry 2012; 51:7873-9. [PMID: 22971172 DOI: 10.1021/bi300510z] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
The Escherichia coli transcription factor NikR is responsible for nickel-mediated repression of the operon encoding the Nik uptake transporter. The crystal structure of Ni(II)-NikR bound to the nik operator sequence revealed that residues in the loop preceding helix α3 in the metal-binding domain, which becomes structurally ordered upon stoichiometric nickel binding, interact with the DNA backbone. Here, we show that mutating both of these residues that make the nonspecific contacts, K64 and R65, abolishes DNA binding in vitro and nickel-responsive transcriptional repression of the nik promoter in vivo. In contrast, mutation of Q118, which forms a bridge between R65 and a potassium site, does not impact the activities of NikR. These data support the model that the nonspecific interactions between the metal-binding domain of the protein and the DNA phosphodiester backbone are critical for the Ni(II)-responsive activity of E. coli NikR.
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Affiliation(s)
- Sandra Krecisz
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada M5S 3H6
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32
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Higgins KA, Chivers PT, Maroney MJ. Role of the N-terminus in determining metal-specific responses in the E. coli Ni- and Co-responsive metalloregulator, RcnR. J Am Chem Soc 2012; 134:7081-93. [PMID: 22471551 PMCID: PMC3375346 DOI: 10.1021/ja300834b] [Citation(s) in RCA: 31] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/28/2022]
Abstract
RcnR (resistance to cobalt and nickel regulator) is a 40-kDa homotetrameric protein and metalloregulator that controls the transcription of the Co(II) and Ni(II) exporter, RcnAB, by binding to DNA as an apoprotein and releasing DNA in response to specifically binding Co(II) and Ni(II) ions. Using X-ray absorption spectroscopy (XAS) to examine the structure of metals bound and lacZ reporter assays of the transcription of RcnA in response to metal binding, in WT and mutant proteins, the roles of coordination number, ligand selection, and residues in the N-terminus of the protein were examined as determinants in metal ion recognition. The studies show that the cognate metal ions, Co(II) and Ni(II), which bind in (N/O)(5)S six-coordinate sites, are distinguished from non-cognate metal ions (Cu(I) and Zn(II)), which bind only three protein ligands and one anion from the buffer, by coordination number and ligand selection. Using mutations of residues near the N-terminus, the N-terminal amine is shown to be a ligand of the cognate metal ions that is missing in the complexes with non-cognate metal ions. The side chain of His3 is also shown to play an important role in distinguishing metal ions. The imidazole group is shown to be a ligand in the Co(II) RcnR complex, but not in the Zn(II) complex. Further, His3 does not appear to bind to Ni(II), providing a structural basis for the differential regulation of RcnAB by the two cognate ions. The Zn(II) complexes change coordination number in response to the residue in position three. In H3C-RcnR, the Zn(II) complex is five-coordinate, and in H3E-RcnR the Zn(II) ion is bound to six protein ligands. The metric parameters of this unusual Zn(II) structure resemble those of the WT-Ni(II) complex, and the mutant protein is able to regulate expression of RcnAB in response to binding the non-cognate ion. The results are discussed within a protein allosteric model for gene regulation by metalloregulators.
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Affiliation(s)
- Khadine A. Higgins
- Department of Chemistry , University of Massachusetts, Amherst, Massachusetts 01003
| | - Peter T. Chivers
- Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis MO 63110
| | - Michael J. Maroney
- Department of Chemistry , University of Massachusetts, Amherst, Massachusetts 01003
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33
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Guerra AJ, Giedroc DP. Metal site occupancy and allosteric switching in bacterial metal sensor proteins. Arch Biochem Biophys 2012; 519:210-22. [PMID: 22178748 PMCID: PMC3312040 DOI: 10.1016/j.abb.2011.11.021] [Citation(s) in RCA: 49] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2011] [Revised: 11/23/2011] [Accepted: 11/29/2011] [Indexed: 12/22/2022]
Abstract
All prokaryotes encode a panel of metal sensor or metalloregulatory proteins that govern the expression of genes that allows an organism to quickly adapt to toxicity or deprivation of both biologically essential transition metal ions, e.g., Zn, Cu, Fe, and heavy metal pollutants. As such, metal sensor proteins can be considered arbiters of intracellular transition metal bioavailability and thus potentially control the metallation state of the metalloproteins in the cell. Metal sensor proteins are specialized allosteric proteins that regulate transcription as a result direct binding of one or two cognate metal ions, to the exclusion of all others. In most cases, the binding of the cognate metal ion induces a structural change in a protein oligomer that either activates or inhibits operator DNA binding. A quantitative measure of the degree to which a particular metal drives metalloregulation of operator DNA-binding is the allosteric coupling free energy, ΔGc. In this review, we summarize recent work directed toward understanding metal occupancy and metal selectivity of these allosteric switches in selected families of metal sensor proteins and examine the structural origins of ΔGc in the functional context a thermodynamic "set-point" model of intracellular metal homeostasis.
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Affiliation(s)
- Alfredo J. Guerra
- Department of Chemistry, Indiana University, 212 S. Hawthorne Drive, Bloomington, IN USA 47405-7102
| | - David P. Giedroc
- Department of Chemistry, Indiana University, 212 S. Hawthorne Drive, Bloomington, IN USA 47405-7102
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34
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Foster AW, Patterson CJ, Pernil R, Hess CR, Robinson NJ. Cytosolic Ni(II) sensor in cyanobacterium: nickel detection follows nickel affinity across four families of metal sensors. J Biol Chem 2012; 287:12142-51. [PMID: 22356910 PMCID: PMC3320959 DOI: 10.1074/jbc.m111.338301] [Citation(s) in RCA: 40] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/18/2023] Open
Abstract
Efflux of surplus Ni(II) across the outer and inner membranes of Synechocystis PCC 6803 is mediated by the Nrs system under the control of a sensor of periplasmic Ni(II), NrsS. Here, we show that the product of ORF sll0176, which encodes a CsoR/RcnR-like protein now designated InrS (for internal nickel-responsive sensor), represses nrsD (NrsD is deduced to efflux Ni(II) across the inner membrane) from a cryptic promoter between the final two ORFs in the nrs operon. Transcripts initiated from the newly identified nrsD promoter accumulate in response to nickel or cobalt but not copper, and recombinant InrS forms specific, Ni(II)-inhibited complexes with the nrsD promoter region. Metal-dependent difference spectra of Ni(II)- and Cu(I)-InrS are similar to Cu(I)-sensing CsoR and dissimilar to Ni(II)/Co(II)-sensing RcnR, consistent with factors beyond the primary coordination sphere switching metal selectivity. Competition with chelators mag-fura-2, nitrilotriacetic acid, EDTA, and EGTA estimate KD Ni(II) for the tightest site of InrS as 2.05 (±1.5) × 10−14m, and weaker KD Ni(II) for the cells' metal sensors of other types: Zn(II) co-repressor Zur, Co(II) activator CoaR, and Zn(II) derepressor ZiaR. Ni(II) transfer to InrS occurs upon addition to Ni(II) forms of each other sensor. InrS binds Ni(II) sufficiently tightly to derepress Ni(II) export at concentrations below KD Ni(II) of the other sensors.
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Affiliation(s)
- Andrew W Foster
- Biophysical Sciences Institute, Department of Chemistry, School of Biological and Biomedical Sciences, University of Durham, Durham DH1 3LE, United Kingdom
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35
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Kaluarachchi H, Siebel JF, Kaluarachchi-Duffy S, Krecisz S, Sutherland DEK, Stillman MJ, Zamble DB. Metal selectivity of the Escherichia coli nickel metallochaperone, SlyD. Biochemistry 2011; 50:10666-77. [PMID: 22047179 DOI: 10.1021/bi2014882] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
SlyD is a Ni(II)-binding protein that contributes to nickel homeostasis in Escherichia coli. The C-terminal domain of SlyD contains a rich variety of metal-binding amino acids, suggesting broader metal binding capabilities, and previous work demonstrated that the protein can coordinate several types of first-row transition metals. However, the binding of SlyD to metals other than Ni(II) has not been previously characterized. To improve our understanding of the in vitro metal-binding activity of SlyD and how it correlates with the in vivo function of this protein, the interactions between SlyD and the series of biologically relevant transition metals [Mn(II), Fe(II), Co(II), Cu(I), and Zn(II)] were examined by using a combination of optical spectroscopy and mass spectrometry. Binding of SlyD to Mn(II) or Fe(II) ions was not detected, but the protein coordinates multiple ions of Co(II), Zn(II), and Cu(I) with appreciable affinity (K(D) values in or below the nanomolar range), highlighting the promiscuous nature of this protein. The order of affinities of SlyD for the metals examined is as follows: Mn(II) and Fe(II) < Co(II) < Ni(II) ~ Zn(II) ≪ Cu(I). Although the purified protein is unable to overcome the large thermodynamic preference for Cu(I) and exclude Zn(II) chelation in the presence of Ni(II), in vivo studies reveal a Ni(II)-specific function for the protein. Furthermore, these latter experiments support a specific role for SlyD as a [NiFe]-hydrogenase enzyme maturation factor. The implications of the divergence between the metal selectivity of SlyD in vitro and the specific activity in vivo are discussed.
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Affiliation(s)
- Harini Kaluarachchi
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada M5S 3H6
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36
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Benini S, Cianci M, Ciurli S. Holo-Ni2+ Helicobacter pylori NikR contains four square-planar nickel-binding sites at physiological pH. Dalton Trans 2011; 40:7831-3. [PMID: 21725560 DOI: 10.1039/c1dt11107h] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
The crystal structure of Helicobacter pylori holo-NikR, a Ni(2+)-dependent transcription factor, determined at pH 7.3, shows four square-planar nickel-binding sites, involving one cysteinate and three histidine ligands. This observation reconciles previous inconsistencies among calorimetric data, structural information at non-physiological pH, and computational studies.
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Affiliation(s)
- Stefano Benini
- Faculty of Science and Technology, Free University of Bolzano, 39100, Bolzano, Italy.
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37
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Zeng YB, Yang N, Sun H. Metal-Binding Properties of an Hpn-Like Histidine-Rich Protein. Chemistry 2011; 17:5852-60. [DOI: 10.1002/chem.201100279] [Citation(s) in RCA: 32] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2011] [Indexed: 01/22/2023]
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38
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Reyes-Caballero H, Campanello GC, Giedroc DP. Metalloregulatory proteins: metal selectivity and allosteric switching. Biophys Chem 2011; 156:103-14. [PMID: 21511390 DOI: 10.1016/j.bpc.2011.03.010] [Citation(s) in RCA: 130] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2011] [Revised: 03/29/2011] [Accepted: 03/29/2011] [Indexed: 01/13/2023]
Abstract
Prokaryotic organisms have evolved the capacity to quickly adapt to a changing and challenging microenvironment in which the availability of both biologically required and non-essential transition metal ions can vary dramatically. In all bacteria, a panel of metalloregulatory proteins controls the expression of genes encoding membrane transporters and metal trafficking proteins that collectively manage metal homeostasis and resistance. These "metal sensors" are specialized allosteric proteins, in which the direct binding of a specific or small number of "cognate" metal ion(s) drives a conformational change in the regulator that allosterically activates or inhibits operator DNA binding, or alternatively, distorts the promoter structure thereby converting a poor promoter to a strong one. In this review, we discuss our current understanding of the features that control metal specificity of the allosteric response in these systems, and the role that structure, thermodynamics and conformational dynamics play in mediating allosteric activation or inhibition of DNA binding.
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39
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Segura-Cabrera A, Guo X, Rojo-Domínguez A, Rodríguez-Pérez MA. Integrative computational protocol for the discovery of inhibitors of the Helicobacter pylori nickel response regulator (NikR). J Mol Model 2011; 17:3075-84. [PMID: 21360181 DOI: 10.1007/s00894-011-0962-2] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2010] [Accepted: 01/05/2011] [Indexed: 12/21/2022]
Abstract
In order to identify novel inhibitors of the Helicobacter pylori nickel response regulator (HpNikR) an integrative protocol was performed for half a million compounds retrieved from the ZINC database. We firstly implement a structure-based virtual screening to build a library of potential inhibitors against the HpNikR using a docking analysis (AutoDock Vina). The library was then used to perform a hierarchical clustering of docking poses, based on protein-contact footprints calculation from the multiple conformations given by the AutoDock Vina software, and the drug-protein interaction analyses to identify and remove potential promiscuous compounds likely interacting with human proteins, hence causing drug side effects. 250 drug-like compounds were finally proposed as non-promicuous potential inhibitors for HpNikR. These compounds target the DNA-binding sites of HpNikR so that HpNikR-compound binding could be able to mimic key interactions in the DNA-protein recognition process. HpNikR inhibitors with promising potential against H. pylori could also act against other human bacterial pathogens due to the conservation of targeting motif of NikR involved in DNA-protein interaction.
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Affiliation(s)
- Aldo Segura-Cabrera
- Laboratorio de Bioinformática, Centro de Biotecnología Genómica, Instituto Politécnico Nacional, Boulevard del Maestro esquina Elías Piña, Colonia Narciso Mendoza, 88710 Ciudad Reynosa, Tamaulipas, Mexico.
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40
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Phillips CM, Stultz CM, Drennan CL. Searching for the Nik operon: how a ligand-responsive transcription factor hunts for its DNA binding site. Biochemistry 2010; 49:7757-63. [PMID: 20712334 PMCID: PMC2934762 DOI: 10.1021/bi100947k] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/17/2023]
Abstract
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Transcription factors regulate a wide variety of genes in the cell and play a crucial role in maintaining cellular homeostasis. A major unresolved issue is how transcription factors find their specific DNA binding sequence in the vast expanse of the cell and how they do so at rates that appear faster than the diffusion limit. Here, we relate an atomic-detail model that has been developed to describe the transcription factor NikR’s mechanism of DNA binding to the broader theories of how transcription factors find their binding sites on DNA. NikR is the nickel regulatory transcription factor for many bacteria, and NikR from Escherichia coli is one of the best studied ligand-mediated transcription factors. For the E. coli NikR protein, there is a wide variety of structural, biochemical, and computational studies that provide significant insight into the NikR−DNA binding mechanism. We find that the two models, the atomic-level model for E. coli NikR and the cellular model for transcription factors in general, are in agreement, and the details laid out by the NikR system may lend additional credence to the current models for transcription factors searching for DNA.
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Affiliation(s)
- Christine M Phillips
- Department of Chemistry, Massachusetts Institute of Technology,Cambridge, Massachusetts 02139, USA
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41
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Phillips CM, Schreiter ER, Stultz CM, Drennan CL. Structural basis of low-affinity nickel binding to the nickel-responsive transcription factor NikR from Escherichia coli. Biochemistry 2010; 49:7830-8. [PMID: 20704276 PMCID: PMC2934763 DOI: 10.1021/bi100923j] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/02/2023]
Abstract
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Escherichia coli NikR regulates cellular nickel uptake by binding to the nik operon in the presence of nickel and blocking transcription of genes encoding the nickel uptake transporter. NikR has two binding affinities for the nik operon: a nanomolar dissociation constant with stoichiometric nickel and a picomolar dissociation constant with excess nickel [Bloom, S. L., and Zamble, D. B. (2004) Biochemistry 43, 10029−10038; Chivers, P. T., and Sauer, R. T. (2002) Chem. Biol. 9, 1141−1148]. While it is known that the stoichiometric nickel ions bind at the NikR tetrameric interface [Schreiter, E. R., et al. (2003) Nat. Struct. Biol. 10, 794−799; Schreiter, E. R., et al. (2006) Proc. Natl. Acad. Sci. U.S.A. 103, 13676−13681], the binding sites for excess nickel ions have not been fully described. Here we have determined the crystal structure of NikR in the presence of excess nickel to 2.6 Å resolution and have obtained nickel anomalous data (1.4845 Å) in the presence of excess nickel for both NikR alone and NikR cocrystallized with a 30-nucleotide piece of double-stranded DNA containing the nik operon. These anomalous data show that excess nickel ions do not bind to a single location on NikR but instead reveal a total of 22 possible low-affinity nickel sites on the NikR tetramer. These sites, for which there are six different types, are all on the surface of NikR, and most are found in both the NikR alone and NikR−DNA structures. Using a combination of crystallographic data and molecular dynamics simulations, the nickel sites can be described as preferring octahedral geometry, utilizing one to three protein ligands (typically histidine) and at least two water molecules.
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Affiliation(s)
- Christine M Phillips
- Department of Chemistry, Massachusetts Institute of Technology,Cambridge, Massachusetts 02139, USA
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42
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Wang SC, Li Y, Ho M, Bernal ME, Sydor AM, Kagzi WR, Zamble DB. The response of Escherichia coli NikR to nickel: a second nickel-binding site. Biochemistry 2010; 49:6635-45. [PMID: 20583753 DOI: 10.1021/bi100685k] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
The Escherichia coli transcription factor NikR mediates two levels of regulatory control of Ni(II) uptake in response to changes in the levels of available nickel. Despite the evidence that metal binding to two distinct sites on NikR, referred to as the high- and low-affinity Ni(II) sites, is required for Ni(II)-selective DNA binding by the protein, the role of the latter set of Ni(II) ions in the activation of NikR remains controversial, and the position of the putative low-affinity Ni(II)-binding site(s) on NikR has not been determined. In this study we confirm that NikR has a high-affinity Ni(II)-binding site that is maintained upon DNA binding. The ligands of the low-affinity Ni(II)-binding site were examined by using selective chemical modification and mass spectrometry performed in the presence of excess Ni(II) and DNA. We localized this Ni(II) site to a region at the interface between the metal- and DNA-binding domains and identified His48 and His110 as residues that participate in the low-affinity Ni(II)-binding response. Mutation of His48 and His110 to asparagines reduces significantly both NikR's tendency to precipitate in the presence of excess Ni(II) and the affinity of the DNA-bound complex in the presence of excess Ni(II). A complete scheme involving all of the metal-binding sites that contribute to the regulatory function of E. coli NikR in nickel homeostasis is described.
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Affiliation(s)
- Sheila C Wang
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada M5S 3H6
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43
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Iwig JS, Chivers PT. Coordinating intracellular nickel-metal-site structure-function relationships and the NikR and RcnR repressors. Nat Prod Rep 2010; 27:658-67. [PMID: 20442957 DOI: 10.1039/b906683g] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
Metalloregulator function requires both sensitivity and selectivity to ensure metal-specific activity without interfering with intracellular metal trafficking pathways. Here, we examine the role of metal coordination geometry in the function of NikR and RcnR, two widely conserved nickel-responsive regulators that are both present in E. coli. The available data suggest an emerging trend in which coordination number is linked to metal-binding affinity, and thus regulatory function. The differences in coordination geometry also suggest that the kinetic mechanisms of metal-association and dissociation will contribute to metalloregulator function. We also discuss ways in which the ligand binding properties of metalloregulators may be tuned to alter the regulatory response.
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Affiliation(s)
- Jeffrey S Iwig
- Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, USA
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44
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Bahlawane C, Dian C, Muller C, Round A, Fauquant C, Schauer K, de Reuse H, Terradot L, Michaud-Soret I. Structural and mechanistic insights into Helicobacter pylori NikR activation. Nucleic Acids Res 2010; 38:3106-18. [PMID: 20089510 PMCID: PMC2875016 DOI: 10.1093/nar/gkp1216] [Citation(s) in RCA: 33] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023] Open
Abstract
NikR is a transcriptional metalloregulator central in the mandatory response to acidity of Helicobacter pylori that controls the expression of numerous genes by binding to specific promoter regions. NikR/DNA interactions were proposed to rely on protein activation by Ni(II) binding to high-affinity (HA) and possibly secondary external (X) sites. We describe a biochemical characterization of HpNikR mutants that shows that the HA sites are essential but not sufficient for DNA binding, while the secondary external (X) sites and residues from the HpNikR dimer–dimer interface are important for DNA binding. We show that a second metal is necessary for HpNikR/DNA binding, but only to some promoters. Small-angle X-ray scattering shows that HpNikR adopts a defined conformation in solution, resembling the cis-conformation and suggests that nickel does not trigger large conformational changes in HpNikR. The crystal structures of selected mutants identify the effects of each mutation on HpNikR structure. This study unravels key structural features from which we derive a model for HpNikR activation where: (i) HA sites and an hydrogen bond network are required for DNA binding and (ii) metallation of a unique secondary external site (X) modulates HpNikR DNA binding to low-affinity promoters by disruption of a salt bridge.
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Affiliation(s)
- C Bahlawane
- CNRS UMR 5249 Laboratoire de Chimie et Biologie des Métaux, France
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45
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Kaluarachchi H, Sutherland DEK, Young A, Pickering IJ, Stillman MJ, Zamble DB. The Ni(II)-Binding Properties of the Metallochaperone SlyD. J Am Chem Soc 2009; 131:18489-500. [DOI: 10.1021/ja9081765] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Affiliation(s)
- Harini Kaluarachchi
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada M5S 3H6, Department of Chemistry, University of Western Ontario, London, Ontario, Canada N6A 5B7, and Department of Geological Sciences, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E2
| | - Duncan E. K. Sutherland
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada M5S 3H6, Department of Chemistry, University of Western Ontario, London, Ontario, Canada N6A 5B7, and Department of Geological Sciences, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E2
| | - Alex Young
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada M5S 3H6, Department of Chemistry, University of Western Ontario, London, Ontario, Canada N6A 5B7, and Department of Geological Sciences, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E2
| | - Ingrid J. Pickering
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada M5S 3H6, Department of Chemistry, University of Western Ontario, London, Ontario, Canada N6A 5B7, and Department of Geological Sciences, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E2
| | - Martin J. Stillman
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada M5S 3H6, Department of Chemistry, University of Western Ontario, London, Ontario, Canada N6A 5B7, and Department of Geological Sciences, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E2
| | - Deborah B. Zamble
- Department of Chemistry, University of Toronto, Toronto, Ontario, Canada M5S 3H6, Department of Chemistry, University of Western Ontario, London, Ontario, Canada N6A 5B7, and Department of Geological Sciences, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E2
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46
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Phillips CM, Nerenberg PS, Drennan CL, Stultz CM. Physical basis of metal-binding specificity in Escherichia coli NikR. J Am Chem Soc 2009; 131:10220-8. [PMID: 19621966 DOI: 10.1021/ja9026314] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Abstract
In Escherichia coli and other bacteria, nickel uptake is regulated by the transcription factor NikR. Nickel binding at high-affinity sites in E. coli NikR (EcNikR) facilitates EcNikR binding to the nik operon, where it then suppresses transcription of genes encoding the nickel uptake transporter, NikABCDE. A structure of the EcNikR-DNA complex suggests that a second metal-binding site is also present when NikR binds to the nik operon. Moreover, this co-crystal structure raises the question of what metal occupies the second site under physiological conditions: K(+), which is present in the crystal structure, or Ni(2+), which has been proposed to bind to low- as well as high-affinity sites on EcNikR. To determine which ion is preferred at the second metal-binding site and the physical basis for any preference of one ion over another in both the second metal-binding site and the high-affinity sites, we conducted a series of detailed molecular simulations on the EcNikR structure. Simulations that place Ni(2+) at high-affinity sites lead to stable trajectories with realistic ion-ligand distances and geometries, while simulations that place K(+) at these sites lead to conformational changes in the protein that are likely unfavorable for ion binding. By contrast, simulations on the second metal site in the EcNikR-DNA complex lead to stable trajectories with realistic geometries regardless of whether K(+) or Ni(2+) occupies this site. Electrostatic binding free energy calculations, however, suggest that EcNikR binding to DNA is more favorable when the second metal-binding site contains K(+). An analysis of the energetic contributions to the electrostatic binding free energy suggests that, while the interaction between EcNikR and DNA is more favorable when the second site contains Ni(2+), the large desolvation penalty associated with moving Ni(2+) from solution to the relatively buried second site offsets this favorable interaction term. Additional free energy simulations that account for both electrostatic and non-electrostatic effects argue that EcNikR binding to DNA is most favorable when the second site contains a monovalent ion the size of K(+). Taken together, these data suggest that the EcNikR structure is most stable when Ni(2+) occupies high-affinity sites and that EcNikR binding to DNA is more favorable when the second site contains K(+).
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Affiliation(s)
- Christine M Phillips
- Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA
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47
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Ma Z, Jacobsen FE, Giedroc DP. Coordination chemistry of bacterial metal transport and sensing. Chem Rev 2009; 109:4644-81. [PMID: 19788177 PMCID: PMC2783614 DOI: 10.1021/cr900077w] [Citation(s) in RCA: 434] [Impact Index Per Article: 27.1] [Reference Citation Analysis] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Affiliation(s)
- Zhen Ma
- Department of Chemistry, Indiana University, Bloomington, IN 47401-7005 USA
- Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843-2128 USA
| | - Faith E. Jacobsen
- Department of Chemistry, Indiana University, Bloomington, IN 47401-7005 USA
| | - David P. Giedroc
- Department of Chemistry, Indiana University, Bloomington, IN 47401-7005 USA
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48
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Affiliation(s)
- Yanjie Li
- Department of Chemistry, University of Toronto, 80 St. George Street, Toronto, ON M5S 3H6, Canada
| | - Deborah B. Zamble
- Department of Chemistry, University of Toronto, 80 St. George Street, Toronto, ON M5S 3H6, Canada
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49
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Jenkins RM, Singleton ML, Almaraz E, Reibenspies JH, Darensbourg MY. Imidazole-containing (N3S)-Ni(II) complexes relating to nickel containing biomolecules. Inorg Chem 2009; 48:7280-93. [PMID: 19572492 PMCID: PMC2908898 DOI: 10.1021/ic900778k] [Citation(s) in RCA: 39] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
Dimeric (N(2)S)Ni complexes and the monomeric N(2)S(2) bismercaptodiazacycloheptane nickel complex, (bme-dach)Ni, serve as precursors to two N(2)-, N'-/ S- complexes where N(2) = diazacycloheptane, N' = imidazole and S = thiolate. As rare examples of nickel complexes containing a mixed thiolate/imidazole ligand set, these complexes are characterized by X-ray diffraction, UV/vis, and variable temperature (1)H NMR spectroscopies, and electrochemistry. Density functional theory computations relate the orientation of the imidazole with respect to the N(2)N'SNi square plane to the VT NMR observed fluxionality and activation parameters. The superoxide dismutase activity of the imidazole complexes was investigated by the nitroblue tetrazolium assay.
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Affiliation(s)
- Roxanne M. Jenkins
- Department of Chemistry, Texas A&M University, College Station, Texas 77843
| | | | - Elky Almaraz
- Department of Chemistry, Texas A&M University, College Station, Texas 77843
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Ma Z, Cowart DM, Scott RA, Giedroc DP. Molecular insights into the metal selectivity of the copper(I)-sensing repressor CsoR from Bacillus subtilis. Biochemistry 2009; 48:3325-34. [PMID: 19249860 DOI: 10.1021/bi900115w] [Citation(s) in RCA: 89] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
Bacillus subtilis CsoR (Bsu CsoR) is a copper-sensing transcriptional repressor that regulates the expression of the copZA operon encoding a copper chaperone and a Cu efflux P-type ATPase, respectively. Bsu CsoR is a homologue of Mycobacterium tuberculosis CsoR (Mtb CsoR), representative of a large Cu(I)-sensing regulatory protein family. We show here that Bsu CsoR binds approximately 1 mol equiv of Cu(I) per monomer in vitro with an affinity >or=10(21) M(-1). X-ray absorption spectroscopy shows Cu(I) adopts a trigonal S(2)N coordination like Mtb CsoR. Both apo and Cu(I)-bound Bsu CsoR are stable tetramers in the low micromolar monomer concentration range by sedimentation velocity and equilibrium ultracentrifugation. Apo-Bsu CsoR binds to a pseudopalindromic 30 bp copZA operator-promoter DNA with a stoichiometry of two tetramers per DNA and stepwise affinities of K(1)(apo) = 3.1(+/-0.8) x 10(7) M(-1) and K(2)(apo) = 8.3 (+/-2.2) x 10(7) M(-1) (0.4 M NaCl, 25 degrees C, pH 6.5). Cu(I) Bsu CsoR binds to the same DNA with greatly reduced affinities, K(1)(Cu) = 2.9(+/-0.4) x 10(6) M(-1) and K(2)(Cu) <or= 1.0 x 10(5) M(-1) consistent with a copper-dependent derepression model. This Cu-dependent regulation is abrogated by a "second shell" Glu90-to-Ala substitution. Bsu CsoR binds Ni(II) with very high affinity but forms a non-native coordination geometry, as does Co(II) and likely Zn(II); none of these metals strongly regulates copZA operator DNA binding in vitro. The implications of these findings on the specificity of metal-sensing sites in CsoR/RcnR proteins are discussed.
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Affiliation(s)
- Zhen Ma
- Department of Chemistry, Indiana University, Bloomington, Indiana 47405-7102, USA
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