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1
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Zacharopoulou M, Seetaloo N, Ross J, Stephens AD, Fusco G, McCoy TM, Dai W, Mela I, Fernandez-Villegas A, Martel A, Routh AF, De Simone A, Phillips JJ, Kaminski Schierle GS. Local Ionic Conditions Modulate the Aggregation Propensity and Influence the Structural Polymorphism of α-Synuclein. J Am Chem Soc 2025. [PMID: 40207671 DOI: 10.1021/jacs.4c13473] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 04/11/2025]
Abstract
Parkinson's disease (PD) is linked to the aggregation of the intrinsically disordered protein α-synuclein (aSyn), but the precise triggers and mechanisms driving this process remain unclear. Local environmental factors, such as ion concentrations, can influence aSyn's conformational ensemble and its tendency to aggregate. In this study, we explore how physiologically relevant ions, mainly Ca2+ and Na+, affect aSyn aggregation, monomer structural dynamics, and fibril polymorphism. ThT fluorescence assays show that all ions speed up aggregation, with Ca2+ having the strongest effect. Using heteronuclear single quantum correlation nuclear magnetic resonance (1H-15N HSQC NMR) spectroscopy, we validate that Ca2+ binds at the C-terminus while Na+ interacts nonspecifically across the sequence. Small-angle neutron scattering (SANS) and hydrogen-deuterium exchange mass spectrometry (HDX-MS) show that Na+ leads to more extended aSyn structures, while Ca2+ results in moderate extension. Molecular dynamics (MD) simulations support this, showing Na+ increases extension between the NAC region and C-terminus, whereas Ca2+ biases the ensemble toward a moderately elongated structure. MD also shows that Ca2+ increases water persistence times in the hydration shell, indicating that aSyn aggregation propensity is due to a combination of conformational bias of the monomer and solvent mobility. Atomic force microscopy (AFM) points toward the formation of distinct fibril polymorphs under different ionic conditions, suggesting ion-induced monomer changes contribute to the diversity of fibril structures. These findings underscore the pivotal influence of the local ionic milieu in shaping the structure and aggregation propensity of aSyn, offering insights into the molecular underpinnings of PD and potential therapeutic strategies targeting aSyn dynamics.
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Affiliation(s)
- Maria Zacharopoulou
- Department of Chemical Engineering and Biotechnology, University of Cambridge, Philippa Fawcett Drive, Cambridge CB3 0AS, U.K
| | - Neeleema Seetaloo
- Living Systems Institute, University of Exeter, Stocker Road, Exeter EX4 4QD, U.K
| | - James Ross
- School of Molecular and Cellular Biology and Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds LS2 9JT, U.K
| | - Amberley D Stephens
- Department of Chemical Engineering and Biotechnology, University of Cambridge, Philippa Fawcett Drive, Cambridge CB3 0AS, U.K
| | - Giuliana Fusco
- Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, U.K
- Department of Life Sciences, Imperial College London, London SW7 2AZ, U.K
| | - Thomas M McCoy
- Department of Chemical Engineering and Biotechnology, University of Cambridge, Philippa Fawcett Drive, Cambridge CB3 0AS, U.K
| | - Wenyue Dai
- Department of Chemical Engineering and Biotechnology, University of Cambridge, Philippa Fawcett Drive, Cambridge CB3 0AS, U.K
| | - Ioanna Mela
- Department of Chemical Engineering and Biotechnology, University of Cambridge, Philippa Fawcett Drive, Cambridge CB3 0AS, U.K
| | - Ana Fernandez-Villegas
- Department of Chemical Engineering and Biotechnology, University of Cambridge, Philippa Fawcett Drive, Cambridge CB3 0AS, U.K
| | - Anne Martel
- Institut Laue Langevin, 71 Avenue des Martyrs, Grenoble CS 20156 38042, France
| | - Alexander F Routh
- Department of Chemical Engineering and Biotechnology, University of Cambridge, Philippa Fawcett Drive, Cambridge CB3 0AS, U.K
| | - Alfonso De Simone
- Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, U.K
- Department of Life Sciences, Imperial College London, London SW7 2AZ, U.K
| | - Jonathan J Phillips
- Living Systems Institute, University of Exeter, Stocker Road, Exeter EX4 4QD, U.K
| | - Gabriele S Kaminski Schierle
- Department of Chemical Engineering and Biotechnology, University of Cambridge, Philippa Fawcett Drive, Cambridge CB3 0AS, U.K
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2
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Yang J, Cheng WX, Zhang P, Wu G, Sheng ST, Yang J, Zhao S, Hu Q, Ji W, Shi Q. Conformational ensembles for protein structure prediction. Sci Rep 2025; 15:8513. [PMID: 40074747 PMCID: PMC11904239 DOI: 10.1038/s41598-024-84066-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2024] [Accepted: 12/19/2024] [Indexed: 03/14/2025] Open
Abstract
Acquisition of conformational ensembles for a protein is a challenging task, which is actually involving to the solution for protein folding problem and the study of intrinsically disordered protein. Despite AlphaFold with artificial intelligence acquired unprecedented accuracy to predict structures, its result is limited to a single state of conformation and it cannot provide multiple conformations to display protein intrinsic disorder. To overcome the barrier, a FiveFold approach was developed with a single sequence method. It applied the protein folding shape code (PFSC) uniformly to expose local folds of five amino acid residues, formed the protein folding variation matrix (PFVM) to reveal local folding variations along sequence, obtained a massive number of folding conformations in PFSC strings, and then an ensemble of multiple conformational protein structures is constructed. The P53_HUMAN as a well-known protein and LEF1_HUMAN and Q8GT36_SPIOL as typical disordered proteins are token as the benchmark to evaluate the predicted outcomes. The results demonstrated an effective algorithm and biological meaningful process well to predict protein multiple conformation structures.
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Affiliation(s)
- Jiaan Yang
- Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, Guangdong, China.
- Micro Biotech, Ltd., Shanghai, 200123, China.
| | - Wen Xiang Cheng
- Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, Guangdong, China
| | - Peng Zhang
- Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, Guangdong, China
- Biomedical Engineering, Shenzhen University of Advanced Technology, Shenzhen, 518060, China
| | - Gang Wu
- School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, Hubei, China
| | - Si Tong Sheng
- HYK High-Throughput Biotechnology Institute, Shenzhen, 518057, Guangdong, China
| | - Junjie Yang
- Wuhan International Biohub Cooperation, Wuhan, 430075, Hubei, China
| | - Suwen Zhao
- iHuman Institute, ShanghaiTech University, Shanghai, 201210, China
| | - Qiyue Hu
- Beyang Therapeutics Co. Ltd, Shanghai, 201210, China
| | - Wenxin Ji
- National Facility for Protein Science in Shanghai, Shanghai Advanced Research Institute, Chinese Academy of Sciences, Shanghai, 201210, China
| | - Qiong Shi
- Laboratory of Aquatic Genomics, College of Life Sciences and Oceanography, Shenzhen University, Shenzhen, 518057, China
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3
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Erboz A, Kesekler E, Gentili PL, Uversky VN, Coskuner-Weber O. Electromagnetic radiation and biophoton emission in neuronal communication and neurodegenerative diseases. PROGRESS IN BIOPHYSICS AND MOLECULAR BIOLOGY 2025; 195:87-99. [PMID: 39732343 DOI: 10.1016/j.pbiomolbio.2024.12.004] [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: 11/15/2024] [Revised: 12/08/2024] [Accepted: 12/24/2024] [Indexed: 12/30/2024]
Abstract
The intersection of electromagnetic radiation and neuronal communication, focusing on the potential role of biophoton emission in brain function and neurodegenerative diseases is an emerging research area. Traditionally, it is believed that neurons encode and communicate information via electrochemical impulses, generating electromagnetic fields detectable by EEG and MEG. Recent discoveries indicate that neurons may also emit biophotons, suggesting an additional communication channel alongside the regular synaptic interactions. This dual signaling system is analyzed for its potential in synchronizing neuronal activity and improving information transfer, with implications for brain-like computing systems. The clinical relevance is explored through the lens of neurodegenerative diseases and intrinsically disordered proteins, where oxidative stress may alter biophoton emission, offering clues for pathological conditions, such as Alzheimer's and Parkinson's diseases. The potential therapeutic use of Low-Level Laser Therapy (LLLT) is also examined for its ability to modulate biophoton activity and mitigate oxidative stress, presenting new opportunities for treatment. Here, we invite further exploration into the intricate roles the electromagnetic phenomena play in brain function, potentially leading to breakthroughs in computational neuroscience and medical therapies for neurodegenerative diseases.
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Affiliation(s)
- Aysin Erboz
- Molecular Biotechnology, Turkish-German University, Sahinkaya Caddesi No. 106, Beykoz, Istanbul, 34820, Turkey
| | - Elif Kesekler
- Molecular Biotechnology, Turkish-German University, Sahinkaya Caddesi No. 106, Beykoz, Istanbul, 34820, Turkey
| | - Pier Luigi Gentili
- Department of Chemistry, Biology, and Biotechnology, Università degli Studi di Perugia, 06123, Perugia, Italy.
| | - Vladimir N Uversky
- Department of Molecular Medicine and USF Health Byrd Alzheimer's Institute, Morsani College of Medicine, University of South Florida, 12901 Bruce B. Downs Blvd., MDC07, Tampa, FL 33612, USA.
| | - Orkid Coskuner-Weber
- Molecular Biotechnology, Turkish-German University, Sahinkaya Caddesi No. 106, Beykoz, Istanbul, 34820, Turkey.
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4
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Kotowski K, Roterman I, Stapor K. DisorderUnetLM: Validating ProteinUnet for efficient protein intrinsic disorder prediction. Comput Biol Med 2025; 185:109586. [PMID: 39708500 DOI: 10.1016/j.compbiomed.2024.109586] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2024] [Revised: 12/03/2024] [Accepted: 12/14/2024] [Indexed: 12/23/2024]
Abstract
The prediction of intrinsic disorder regions has significant implications for understanding protein functions and dynamics. It can help to discover novel protein-protein interactions essential for designing new drugs and enzymes. Recently, a new generation of predictors based on protein language models (pLMs) is emerging. These algorithms reach state-of-the-art accuracy without calculating time-consuming multiple sequence alignments (MSAs). This article introduces the new DisorderUnetLM disorder predictor, which builds upon the idea of ProteinUnet. It uses the Attention U-Net convolutional network and incorporates features from the ProtTrans pLM. DisorderUnetLM achieves top results in the direct comparison with recent predictors exploiting MSAs and pLMs. Moreover, among 43 predictors on the latest CAID-2 benchmark, it ranks 1st for the NOX subset in terms of the ROC-AUC metric (0.844) and 2nd for the AP metric (0.596). For the CAID-2 PDB subset, it ranks in the top 10 (ROC-AUC of 0.924 and AP of 0.862). The code and model are publicly available and fully reproducible at doi.org/10.24433/CO.7350682.v1.
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Affiliation(s)
- Krzysztof Kotowski
- Department of Applied Informatics, Silesian University of Technology, Akademicka 16, 44-100, Gliwice, Poland
| | - Irena Roterman
- Department of Bioinformatics and Telemedicine, Jagiellonian University Medical College, Medyczna 7, 30-688, Kraków, Poland
| | - Katarzyna Stapor
- Department of Applied Informatics, Silesian University of Technology, Akademicka 16, 44-100, Gliwice, Poland.
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5
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Gollapalli S, Sooram B, Sugandh H, Saudagar P. The landscape of intrinsically disordered proteins in Leishmania parasite: Implications for drug discovery. Int J Biol Macromol 2024; 283:137290. [PMID: 39537071 DOI: 10.1016/j.ijbiomac.2024.137290] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/29/2024] [Revised: 11/01/2024] [Accepted: 11/04/2024] [Indexed: 11/16/2024]
Abstract
Proteins that lack three-dimensional structures are known as Intrinsically disordered proteins (IDPs). In this study, we aimed to identify intrinsically disordered proteins in the Leishmania donovani proteome using various predictors that can identify IDPs based on amino acid residues and charge hydropathy. Top identified IDPs are analyzed using STRING, PSP-Hunter, Deep Loc-2.0, and Alpha fold to understand the protein-protein interaction, phase separation, localization, and structural assessment of those proteins. From this study, we found that >50 % of Leishmania donovani proteome has proteins or regions of proteins that are intrinsically disordered with VSL2 score >0.5; most proteins interact with many other proteins with PPI enrichment p-value <1.0e-16. Few proteins, such as Protein phosphatase inhibitor, UMSBP, and Zinc knuckle, have redox-sensitive regions. Functional disorder profiles of identified IDPs showed MoRFs and predicted protein domains. HASPB, UTP11, Nucleolar protein 12, and UMSBP have a high probability of undergoing phase separation. Localization studies showed that most of these proteins are in the cytoplasm and nucleus. Our present study of identifying IDPs in Leishmania proteome yields significant information on druggable targets and can serve as a basis for further studies to identify unexplored pathways.
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Affiliation(s)
- Seshaveena Gollapalli
- Department of Biotechnology, National Institute of Technology-Warangal, Warangal 506004, Telangana, India
| | - Banesh Sooram
- Division of Neurogeriatrics, Karolinska Institutet, Solna, 17 165, Solnavagen, Sweden
| | - Hitesh Sugandh
- Department of Biotechnology, National Institute of Technology-Warangal, Warangal 506004, Telangana, India
| | - Prakash Saudagar
- Department of Biotechnology, National Institute of Technology-Warangal, Warangal 506004, Telangana, India.
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6
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Kim S, Lee NK, Jung Y, Johner A. Kinetics of Polyampholyte Dimerization: Influence of Charge Sequences. Polymers (Basel) 2024; 16:2928. [PMID: 39458755 PMCID: PMC11510756 DOI: 10.3390/polym16202928] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2024] [Revised: 10/09/2024] [Accepted: 10/10/2024] [Indexed: 10/28/2024] Open
Abstract
Polyampholytes (PAs) exhibit complex behaviors in various environments influenced by their charge distribution. This study focuses on the kinetics of dimerization of PAs, aiming to elucidate the underlying mechanisms and clarify relevant characteristics of the charge sequence. We focus on PAs with non-zero net charges, employing molecular dynamics simulations and theoretical analyses to examine how charge sequences influence the rates of dimer formation and dissociation. Our findings reveal that the charge sequence of tails and the blockiness of the minority charge group markedly influence the kinetics of dimerization: large blockiness and tails with a high number of majority-type charges slow down the dissociation of dimers. Additionally, the presence of an extended (central) block of the majority charge promotes structural diversity. Within dimer states, blocks alternate between intra- and inter-chain contacts. The duration times in the dimer states are significantly longer than the typical dwell times of block inter-contacts, with a notable extension when multiple blocks are engaged. Intrinsically disordered proteins (IDPs) play crucial roles in cellular functions, primarily due to their ability to undergo rapid conformational changes and form transient complexes. These properties largely depend on the sequence of charged residues. We provide insights into the fundamental principles governing the structural and dynamical properties of polyampholytic IDP, emphasizing the importance of sequence-specific effects on both aggregation and dissociation.
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Affiliation(s)
- Seowon Kim
- Department of Physics and Astronomy, Sejong University, Seoul 05006, Republic of Korea
| | - Nam-Kyung Lee
- Department of Physics and Astronomy, Sejong University, Seoul 05006, Republic of Korea
| | - Youngkyun Jung
- Supercomputing Center, Korea Institute of Science and Technology Information, Daejeon 34141, Republic of Korea;
| | - Albert Johner
- Institut Charles Sadron CNRS-Unistra, 6 rue Boussingault, 67083 Strasbourg, CEDEX, France
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7
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Manav N, Jit BP, Kataria B, Sharma A. Cellular and epigenetic perspective of protein stability and its implications in the biological system. Epigenomics 2024; 16:879-900. [PMID: 38884355 PMCID: PMC11370918 DOI: 10.1080/17501911.2024.2351788] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2023] [Accepted: 04/30/2024] [Indexed: 06/18/2024] Open
Abstract
Protein stability is a fundamental prerequisite in both experimental and therapeutic applications. Current advancements in high throughput experimental techniques and functional ontology approaches have elucidated that impairment in the structure and stability of proteins is intricately associated with the cause and cure of several diseases. Therefore, it is paramount to deeply understand the physical and molecular confounding factors governing the stability of proteins. In this review article, we comprehensively investigated the evolution of protein stability, examining its emergence over time, its relationship with organizational aspects and the experimental methods used to understand it. Furthermore, we have also emphasized the role of Epigenetics and its interplay with post-translational modifications (PTMs) in regulating the stability of proteins.
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Affiliation(s)
- Nisha Manav
- Department of Biochemistry, All India Institute of Medical Sciences New Delhi, Ansari Nagar, 110029, India
| | - Bimal Prasad Jit
- Department of Biochemistry, All India Institute of Medical Sciences New Delhi, Ansari Nagar, 110029, India
| | - Babita Kataria
- Department of Medical Oncology, National Cancer Institute, All India Institute of Medical Sciences, Jhajjar, 124105, India
| | - Ashok Sharma
- Department of Biochemistry, All India Institute of Medical Sciences New Delhi, Ansari Nagar, 110029, India
- Department of Biochemistry, National Cancer Institute, All India Institute of Medical Sciences, Jhajjar, 124105, India
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8
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Uversky VN, Madeira PP, Zaslavsky BY. What Can Be Learned from the Partitioning Behavior of Proteins in Aqueous Two-Phase Systems? Int J Mol Sci 2024; 25:6339. [PMID: 38928046 PMCID: PMC11203663 DOI: 10.3390/ijms25126339] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2024] [Revised: 05/31/2024] [Accepted: 06/03/2024] [Indexed: 06/28/2024] Open
Abstract
This review covers the analytical applications of protein partitioning in aqueous two-phase systems (ATPSs). We review the advancements in the analytical application of protein partitioning in ATPSs that have been achieved over the last two decades. Multiple examples of different applications, such as the quality control of recombinant proteins, analysis of protein misfolding, characterization of structural changes as small as a single-point mutation, conformational changes upon binding of different ligands, detection of protein-protein interactions, and analysis of structurally different isoforms of a protein are presented. The new approach to discovering new drugs for a known target (e.g., a receptor) is described when one or more previous drugs are already available with well-characterized biological efficacy profiles.
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Affiliation(s)
- Vladimir N. Uversky
- Department of Molecular Medicine, Byrd Alzheimer’s Research Institute, Morsani College of Medicine, University of South Florida, Tampa, FL 33612, USA;
| | - Pedro P. Madeira
- Centro de Investigacao em Materiais Ceramicos e Compositos, Department of Chemistry, 3810-193 Aveiro, Portugal;
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9
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Viegas RG, Martins IBS, Leite VBP. Understanding the Energy Landscape of Intrinsically Disordered Protein Ensembles. J Chem Inf Model 2024; 64:4149-4157. [PMID: 38713459 DOI: 10.1021/acs.jcim.4c00080] [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: 05/08/2024]
Abstract
A substantial portion of various organisms' proteomes comprises intrinsically disordered proteins (IDPs) that lack a defined three-dimensional structure. These IDPs exhibit a diverse array of conformations, displaying remarkable spatiotemporal heterogeneity and exceptional conformational flexibility. Characterizing the structure or structural ensemble of IDPs presents significant conceptual and methodological challenges owing to the absence of a well-defined native structure. While databases such as the Protein Ensemble Database (PED) provide IDP ensembles obtained through a combination of experimental data and molecular modeling, the absence of reaction coordinates poses challenges in comprehensively understanding pertinent aspects of the system. In this study, we leverage the energy landscape visualization method (JCTC, 6482, 2019) to scrutinize four IDP ensembles sourced from PED. ELViM, a methodology that circumvents the need for a priori reaction coordinates, aids in analyzing the ensembles. The specific IDP ensembles investigated are as follows: two fragments of nucleoporin (NUL: 884-993 and NUS: 1313-1390), yeast sic 1 N-terminal (1-90), and the N-terminal SH3 domain of Drk (1-59). Utilizing ELViM enables the comprehensive validation of ensembles, facilitating the detection of potential inconsistencies in the sampling process. Additionally, it allows for identifying and characterizing the most prevalent conformations within an ensemble. Moreover, ELViM facilitates the comparative analysis of ensembles obtained under diverse conditions, thereby providing a powerful tool for investigating the functional mechanisms of IDPs.
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Affiliation(s)
- Rafael G Viegas
- Federal Institute of Education, Science and Technology of São Paulo (IFSP), Catanduva, São Paulo 15.808-305, Brazil
- Department of Physics, São Paulo State University (UNESP), Institute of Biosciences, Humanities and Exact Sciences, São José do Rio Preto, São Paulo 15054-000, Brazil
| | - Ingrid B S Martins
- Department of Physics, São Paulo State University (UNESP), Institute of Biosciences, Humanities and Exact Sciences, São José do Rio Preto, São Paulo 15054-000, Brazil
| | - Vitor B P Leite
- Department of Physics, São Paulo State University (UNESP), Institute of Biosciences, Humanities and Exact Sciences, São José do Rio Preto, São Paulo 15054-000, Brazil
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10
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Tavili E, Aziziyan F, Dabirmanesh B. Pathways of amyloid fibril formation and protein aggregation. PROGRESS IN MOLECULAR BIOLOGY AND TRANSLATIONAL SCIENCE 2024; 206:11-54. [PMID: 38811078 DOI: 10.1016/bs.pmbts.2024.03.010] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/31/2024]
Abstract
The main cause of many neurodegenerative diseases and systemic amyloidoses is protein and peptide aggregation and the formation of amyloid fibrils. The study of aggregation mechanisms, the discovery and description of aggregate structures, and a comprehensive understanding of the molecular mechanisms of amyloid formation are of great importance for the diagnostic processes at the molecular level and for the development of therapeutic strategies to counter aggregation-associated disorders. Given that understanding protein misfolding phenomena is directly related to the protein folding process, we will briefly explain the protein folding mechanism and then discuss the important factors involved in protein aggregation. In the following, we review different mechanisms of amyloid formation and finally represent the current knowledge on how amyloid fibrils are formed based on kinetic and thermodynamic factors.
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Affiliation(s)
- Elaheh Tavili
- Department of Biochemistry, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran
| | - Fatemeh Aziziyan
- Department of Biochemistry, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran
| | - Bahareh Dabirmanesh
- Department of Biochemistry, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
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11
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Praveen M. Characterizing the West Nile Virus's polyprotein from nucleotide sequence to protein structure - Computational tools. J Taibah Univ Med Sci 2024; 19:338-350. [PMID: 38304694 PMCID: PMC10831166 DOI: 10.1016/j.jtumed.2024.01.001] [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: 07/11/2023] [Revised: 11/27/2023] [Accepted: 01/08/2024] [Indexed: 02/03/2024] Open
Abstract
Objectives West Nile virus (WNV) belongs to the Flaviviridae family and causes West Nile fever. The mechanism of transmission involves the culex mosquito species. Infected individuals are primarily asymptomatic, and few exhibit common symptoms. Moreover, 10 % of neuronal infection caused by this virus cause death. The proteins encoded by these genes had been uncharacterized, although understanding their function and structure is important for formulating antiviral drugs. Methods Herein, we used in silico approaches, including various bioinformatic tools and databases, to analyse the proteins from the WNV polyprotein individually. The characterization included GC content, physicochemical properties, conserved domains, soluble and transmembrane regions, signal localization, protein disorder, and secondary structure features and their respective 3D protein structures. Results Among 11 proteins, eight had >50 % GC content, eight proteins had basic pI values, three proteins were unstable under in vitro conditions, four were thermostable according to >100 AI values and some had negative GRAVY values in physicochemical analyses. All protein-conserved domains were shared among Flaviviridae family members. Five proteins were soluble and lacked transmembrane regions. Two proteins had signals for localization in the host endoplasmic reticulum. Non-structural (NS) 2A showed low protein disorder. The secondary structural features and tertiary structure models provide a valuable biochemical resource for designing selective substrates and synthetic inhibitors. Conclusions WNV proteins NS2A, NS2B, PM, NS3 and NS5 can be used as drug targets for the pharmacological design of lead antiviral compounds.
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Affiliation(s)
- Mallari Praveen
- Department of Zoology, Indira Gandhi National Tribal University, Amarkantak, Madhya Pradesh, India
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12
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Maiti S, Singh A, Maji T, Saibo NV, De S. Experimental methods to study the structure and dynamics of intrinsically disordered regions in proteins. Curr Res Struct Biol 2024; 7:100138. [PMID: 38707546 PMCID: PMC11068507 DOI: 10.1016/j.crstbi.2024.100138] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2023] [Revised: 03/12/2024] [Accepted: 03/15/2024] [Indexed: 05/07/2024] Open
Abstract
Eukaryotic proteins often feature long stretches of amino acids that lack a well-defined three-dimensional structure and are referred to as intrinsically disordered proteins (IDPs) or regions (IDRs). Although these proteins challenge conventional structure-function paradigms, they play vital roles in cellular processes. Recent progress in experimental techniques, such as NMR spectroscopy, single molecule FRET, high speed AFM and SAXS, have provided valuable insights into the biophysical basis of IDP function. This review discusses the advancements made in these techniques particularly for the study of disordered regions in proteins. In NMR spectroscopy new strategies such as 13C detection, non-uniform sampling, segmental isotope labeling, and rapid data acquisition methods address the challenges posed by spectral overcrowding and low stability of IDPs. The importance of various NMR parameters, including chemical shifts, hydrogen exchange rates, and relaxation measurements, to reveal transient secondary structures within IDRs and IDPs are presented. Given the high flexibility of IDPs, the review outlines NMR methods for assessing their dynamics at both fast (ps-ns) and slow (μs-ms) timescales. IDPs exert their functions through interactions with other molecules such as proteins, DNA, or RNA. NMR-based titration experiments yield insights into the thermodynamics and kinetics of these interactions. Detailed study of IDPs requires multiple experimental techniques, and thus, several methods are described for studying disordered proteins, highlighting their respective advantages and limitations. The potential for integrating these complementary techniques, each offering unique perspectives, is explored to achieve a comprehensive understanding of IDPs.
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Affiliation(s)
| | - Aakanksha Singh
- School of Bioscience, Indian Institute of Technology Kharagpur, Kharagpur, WB, 721302, India
| | - Tanisha Maji
- School of Bioscience, Indian Institute of Technology Kharagpur, Kharagpur, WB, 721302, India
| | - Nikita V. Saibo
- School of Bioscience, Indian Institute of Technology Kharagpur, Kharagpur, WB, 721302, India
| | - Soumya De
- School of Bioscience, Indian Institute of Technology Kharagpur, Kharagpur, WB, 721302, India
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13
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Papadimitropoulou A, Makri M, Zoidis G. MYC the oncogene from hell: Novel opportunities for cancer therapy. Eur J Med Chem 2024; 267:116194. [PMID: 38340508 DOI: 10.1016/j.ejmech.2024.116194] [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: 10/30/2023] [Revised: 01/24/2024] [Accepted: 01/25/2024] [Indexed: 02/12/2024]
Abstract
Cancer comprises a heterogeneous disease, characterized by diverse features such as constitutive expression of oncogenes and/or downregulation of tumor suppressor genes. MYC constitutes a master transcriptional regulator, involved in many cellular functions and is aberrantly expressed in more than 70 % of human cancers. The Myc protein belongs to a family of transcription factors whose structural pattern is referred to as basic helix-loop-helix-leucine zipper. Myc binds to its partner, a smaller protein called Max, forming an Myc:Max heterodimeric complex that interacts with specific DNA recognition sequences (E-boxes) and regulates the expression of downstream target genes. Myc protein plays a fundamental role for the life of a cell, as it is involved in many physiological functions such as proliferation, growth and development since it controls the expression of a very large percentage of genes (∼15 %). However, despite the strict control of MYC expression in normal cells, MYC is often deregulated in cancer, exhibiting a key role in stimulating oncogenic process affecting features such as aberrant proliferation, differentiation, angiogenesis, genomic instability and oncogenic transformation. In this review we aim to meticulously describe the fundamental role of MYC in tumorigenesis and highlight its importance as an anticancer drug target. We focus mainly on the different categories of novel small molecules that act as inhibitors of Myc function in diverse ways hence offering great opportunities for an efficient cancer therapy. This knowledge will provide significant information for the development of novel Myc inhibitors and assist to the design of treatments that would effectively act against Myc-dependent cancers.
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Affiliation(s)
- Adriana Papadimitropoulou
- Center for Basic Research, Biomedical Research Foundation of the Academy of Athens, Athens, 11527, Greece
| | - Maria Makri
- Division of Pharmaceutical Chemistry, Department of Pharmacy, School of Health Sciences, National and Kapodistrian University of Athens, Panepistimiopolis-Zografou, GR-15771, Athens, Greece
| | - Grigoris Zoidis
- Division of Pharmaceutical Chemistry, Department of Pharmacy, School of Health Sciences, National and Kapodistrian University of Athens, Panepistimiopolis-Zografou, GR-15771, Athens, Greece.
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14
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Truong HP, Saleh OA. Magnetic tweezers characterization of the entropic elasticity of intrinsically disordered proteins and peptoids. Methods Enzymol 2024; 694:209-236. [PMID: 38492952 DOI: 10.1016/bs.mie.2023.12.011] [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] [Indexed: 03/18/2024]
Abstract
Understanding the conformational behavior of biopolymers is essential to unlocking knowledge of their biophysical mechanisms and functional roles. Single-molecule force spectroscopy can provide a unique perspective on this by exploiting entropic elasticity to uncover key biopolymer structural parameters. A particularly powerful approach involves the use of magnetic tweezers, which can easily generate lower stretching forces (0.1-20 pN). For forces at the low end of this range, the elastic response of biopolymers is sensitive to excluded volume effects, and they can be described by Pincus blob elasticity model that allow robust extraction of the Flory polymer scaling exponent. Here, we detail protocols for the use of magnetic tweezers for force-extension measurements of intrinsically disordered proteins and peptoids. We also discuss procedures for fitting low-force elastic curves to the predictions of polymer physics models to extract key conformational parameters.
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Affiliation(s)
- Hoang P Truong
- Materials Department, University of California, Santa Barbara, CA, United States
| | - Omar A Saleh
- Materials Department, University of California, Santa Barbara, CA, United States; Biomolecular Sciences and Engineering Program, University of California, Santa Barbara, CA, United States; Physics Department, University of California, Santa Barbara, CA, United States.
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15
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Gunasinghe KJ, Rahman T, Chee Wezen X. Unraveling the Behavior of Intrinsically Disordered Protein c-Myc: A Study Utilizing Gaussian-Accelerated Molecular Dynamics. ACS OMEGA 2024; 9:2250-2262. [PMID: 38250404 PMCID: PMC10795134 DOI: 10.1021/acsomega.3c05822] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 08/08/2023] [Revised: 11/02/2023] [Accepted: 11/21/2023] [Indexed: 01/23/2024]
Abstract
The protein c-Myc is a transcription factor that remains largely intrinsically disordered and is known to be involved in various biological processes and is overexpressed in various cancers, making it an attractive drug target. However, intrinsically disordered proteins such as c-Myc do not show funnel-like basins in their free-energy landscapes; this makes their druggability a challenge. For the first time, we propose a heterodimer model of c-Myc/Max in full length in this work. We used Gaussian-accelerated molecular dynamics (GaMD) simulations to explore the behavior of c-Myc and its various regions, including the transactivation domain (TAD) and the basic helix-loop-helix-leucine-zipper (bHLH-Zipper) motif in three different conformational states: (a) monomeric c-Myc, (b) c-Myc when bound to its partner protein, Max, and (c) when Max was removed after binding. We analyzed the GaMD trajectories using root-mean-square deviation (RMSD), radius of gyration, root-mean-square fluctuation, and free-energy landscape (FEL) calculations to elaborate the behaviors of these regions. The results showed that the monomeric c-Myc structure showed a higher RMSD fluctuation as compared with the c-Myc/Max heterodimer in the bHLH-Zipper motif. This indicated that the bHLH-Zipper motif of c-Myc is more stable when it is bound to Max. The TAD region in both monomeric and Max-bound states showed similar plasticity in terms of RMSD. We also conducted residue decomposition calculations and showed that the c-Myc and Max interaction could be driven mainly by electrostatic interactions and the residues Arg299, Ile403, and Leu420 seemed to play important roles in the interaction. Our work provides insights into the behavior of c-Myc and its regions that could support the development of drugs that target c-Myc and other intrinsically disordered proteins.
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Affiliation(s)
| | - Taufiq Rahman
- Department
of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge CB2 1PD, United Kingdom
| | - Xavier Chee Wezen
- Faculty
of Engineering, Computing and Science, Swinburne
University of Technology Sarawak, Kuching 93350, Malaysia
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16
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Lee NK, Chae MK, Jung Y, Johner A, Joanny JF. Polyelectrolytes: From Seminal Works to the Influence of the Charge Sequence. Polymers (Basel) 2023; 15:4593. [PMID: 38232020 PMCID: PMC10708673 DOI: 10.3390/polym15234593] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2023] [Revised: 11/19/2023] [Accepted: 11/27/2023] [Indexed: 01/19/2024] Open
Abstract
We propose a selected tour of the physics of polyelectrolytes (PE) following the line initiated by de Gennes and coworkers in their seminal 1976 paper. The early works which used uniform charge distributions along the PE backbone achieved tremendous progress and set most milestones in the field. Recently, the focus has shifted to the role of the charge sequence. Revisited topics include PE complexation and polyampholytes (PA). We develop the example of a random PE in poor solvent forming pearl-necklace structures. It is shown that the pearls typically adopt very asymmetric mass and charge distributions. Individual sequences do not necessarily reflect the ensemble statistics and a rich variety of behaviors emerges (specially for PA). Pearl necklaces are dynamic structures and switch between various types of pearl-necklace structures, as described for both PE and PA.
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Affiliation(s)
- Nam-Kyung Lee
- Department of Physics and Astronomy, Sejong University, Seoul 05006, Republic of Korea;
| | - Min-Kyung Chae
- National Institute for Mathematical Sciences, Daejeon 34047, Republic of Korea;
| | - Youngkyun Jung
- Supercomputing Center, Korea Institute of Science and Technology Information, Daejeon 34141, Republic of Korea;
| | - Albert Johner
- Institut Charles Sadron CNRS-Unistra, 6 rue Boussingault, 67083 Strasbourg, France
| | - Jean-Francois Joanny
- Institut Curie, Physique des cellules et Cancer, Collège de France Soft Matter and Biophysics Chair, 11, PSL University, Place Marcelin-Berthelot, 75231 Paris, France;
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17
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Yang J, Cheng WX, Wu G, Sheng S, Zhang P. Prediction of folding patterns for intrinsic disordered protein. Sci Rep 2023; 13:20343. [PMID: 37990040 PMCID: PMC10663623 DOI: 10.1038/s41598-023-45969-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2023] [Accepted: 10/26/2023] [Indexed: 11/23/2023] Open
Abstract
The conformation flexibility of natural protein causes both complexity and difficulty to understand the relationship between structure and function. The prediction of intrinsically disordered protein primarily is focusing on to disclose the regions with structural flexibility involving relevant biological functions and various diseases. The order of amino acids in protein sequence determines possible conformations, folding flexibility and biological function. Although many methods provided the information of intrinsically disordered protein (IDP), but the results are mainly limited to determine the locations of regions without knowledge of possible folding conformations. Here, the developed protein folding fingerprint adopted the protein folding variation matrix (PFVM) to reveal all possible folding patterns for the intrinsically disordered protein along its sequence. The PFVM integrally exhibited the intrinsically disordered protein with disordering regions, degree of disorder as well as folding pattern. The advantage of PFVM will not only provide rich information for IDP, but also may promote the study of protein folding problem.
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Affiliation(s)
- Jiaan Yang
- Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, Guangdong, China.
- Micro Biotech, Ltd., Shanghai, 200123, China.
| | - Wen-Xiang Cheng
- Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, Guangdong, China
| | - Gang Wu
- School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China
| | - Sitong Sheng
- HYK High-throughput Biotechnology Institute, Shenzhen, 518057, Guangdong, China
| | - Peng Zhang
- Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, Guangdong, China
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18
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Zhang Y, Zhang X, Zhu L, Wang L, Zhang H, Zhang X, Xu S, Xue J. Identification of the Maize LEA Gene Family and Its Relationship with Kernel Dehydration. PLANTS (BASEL, SWITZERLAND) 2023; 12:3674. [PMID: 37960031 PMCID: PMC10647770 DOI: 10.3390/plants12213674] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/07/2023] [Revised: 10/21/2023] [Accepted: 10/22/2023] [Indexed: 11/15/2023]
Abstract
Maize, the most widely planted and highest yielding of the three major crops in the world, requires the development and breeding of new varieties to accommodate the shift towards mechanized harvesting. However, the moisture content of kernels during harvest poses a significant challenge to mechanized harvesting, leading to seed breakage and increased storage costs. Previous studies highlighted the importance of LEA (Late Embryogenesis Abundant) members in regulating kernel dehydration. In this study, we aimed to gain a better understanding of the relationship between the LEA family and grain dehydration in maize. Through expression pattern analysis of maize, we identified 52 LEA genes (ZmLEAs) distributed across 10 chromosomes, organized into seven subgroups based on phylogenetic analysis, gene structure, and conserved motifs. Evolutionary and selective pressure analysis revealed that the amplification of ZmLEA genes primarily resulted from whole-genome or fragment replication events, with strong purifying selection effects during evolution. Furthermore, the transcriptome data of kernels of two maize inbred lines with varying dehydration rates at different developmental stages showed that 14 ZmLEA genes were expressed differentially in the two inbreds. This suggested that the ZmLEA genes might participate in regulating the kernel dehydration rate (KDR) in maize. Overall, this study enhances our understanding of the ZmLEA family and provides a foundation for further research into its role in regulating genes associated with grain dehydration in maize.
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Affiliation(s)
| | | | | | | | | | | | - Shutu Xu
- Key Laboratory of Biology and Genetic Improvement of Maize in Arid Area of Northwest Region, College of Agronomy, Northwest A&F University, Yangling 712100, China; (Y.Z.); (X.Z.); (L.Z.); (L.W.); (H.Z.); (X.Z.)
| | - Jiquan Xue
- Key Laboratory of Biology and Genetic Improvement of Maize in Arid Area of Northwest Region, College of Agronomy, Northwest A&F University, Yangling 712100, China; (Y.Z.); (X.Z.); (L.Z.); (L.W.); (H.Z.); (X.Z.)
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19
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Ehlert C, Poorinmohammad N, Mohammaei S, Zhang L, Salavati R. Structure-Function Analysis of RBP7910: An Editosome Z-Binding Protein in Trypanosomatids. Molecules 2023; 28:6963. [PMID: 37836806 PMCID: PMC10574248 DOI: 10.3390/molecules28196963] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/13/2023] [Revised: 10/01/2023] [Accepted: 10/05/2023] [Indexed: 10/15/2023] Open
Abstract
RNA editing, a unique post-transcriptional modification, is observed in trypanosomatid parasites as a crucial procedure for the maturation of mitochondrial mRNAs. The editosome protein complex, involving multiple protein components, plays a key role in this process. In Trypanosoma brucei, a putative Z-DNA binding protein known as RBP7910 is associated with the editosome. However, the specific Z-DNA/Z-RNA binding activity and the interacting interface of RBP7910 have yet to be determined. In this study, we conducted a comparative analysis of the binding behavior of RBP7910 with different potential ligands using microscale thermophoresis (MST). Additionally, we generated a 3D model of the protein, revealing potential Z-α and Z-β nucleic acid-binding domains of RBP7910. RBP7910 belongs to the winged-helix-turn-helix (HTH) superfamily of proteins with an α1α2α3β1β2 topology. Finally, using docking techniques, potential interacting surface regions of RBP7910 with notable oligonucleotide ligands were identified. Our findings indicate that RBP7910 exhibits a notable affinity for (CG)n Z-DNA, both in single-stranded and double-stranded forms. Moreover, we observed a broader interacting interface across its Z-α domain when bound to Z-DNA/Z-RNA compared to when bound to non-Z-form nucleic acid ligands.
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Affiliation(s)
- Curtis Ehlert
- Institute of Parasitology, McGill University, Montreal, QC H9X 3V9, Canada; (C.E.); (N.P.); (S.M.); (L.Z.)
| | - Naghmeh Poorinmohammad
- Institute of Parasitology, McGill University, Montreal, QC H9X 3V9, Canada; (C.E.); (N.P.); (S.M.); (L.Z.)
| | - Saba Mohammaei
- Institute of Parasitology, McGill University, Montreal, QC H9X 3V9, Canada; (C.E.); (N.P.); (S.M.); (L.Z.)
| | - Linhua Zhang
- Institute of Parasitology, McGill University, Montreal, QC H9X 3V9, Canada; (C.E.); (N.P.); (S.M.); (L.Z.)
| | - Reza Salavati
- Institute of Parasitology, McGill University, Montreal, QC H9X 3V9, Canada; (C.E.); (N.P.); (S.M.); (L.Z.)
- Department of Biochemistry, McGill University, Montreal, QC H3A 1A3, Canada
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20
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Xavier JAM, Fuentes I, Nuez-Martínez M, Viñas C, Teixidor F. Single stop analysis of a protein surface using molecular probe electrochemistry. J Mater Chem B 2023; 11:8422-8432. [PMID: 37563960 DOI: 10.1039/d3tb00816a] [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: 08/12/2023]
Abstract
Visualization of a protein in its native form and environment without any interference has always been a challenging task. Contrary to the assumption that protein surfaces are smooth, they are in fact highly irregular with undulating surfaces. Hence, in this study, we have tackled this ambiguous nature of the 'surface' of a protein by considering the 'effective' protein surface (EPS) with respect to its interaction with the geometrically well-defined and structurally inert anionic molecule [3,3'-Co(1,2-C2B9H11)2]-, abbreviated as [o-COSAN]-, whose stability, propensity for amine residues, and self-assembling abilities are well reported. This study demonstrates the intricacies of protein surfaces exploiting simple electrochemical measurements using a 'small molecule' redox-active probe. This technique offers the advantage of not utilizing any harsh experimental conditions that could alter the native structure of the protein and hence the protein integrity is retained. Identification of the amino acid residues which are most involved in the interactions with [3,3'-Co(1,2-C2B9H11)2]- and how a protein's environment affects these interactions can help in gaining insights into how to modify proteins to optimize their interactions particularly in the fields of drug design and biotechnology. In this research, we have demonstrated that [3,3'-Co(1,2-C2B9H11)2]- anionic small molecules are excellent candidates for studying and visualizing protein surfaces in their natural environment and allow proteins to be classified according to the surface composition, which imparts their properties. [3,3'-Co(1,2-C2B9H11)2]- 'viewed' each protein surface differently and hence has the potential to act as a simple and easy to handle cantilever for measuring and picturing protein surfaces.
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Affiliation(s)
- Jewel Ann Maria Xavier
- Institut de Ciencia de Materials de Barcelona (ICMAB-CSIC), Campus de la UAB, Bellaterra, Spain.
| | - Isabel Fuentes
- Institut de Ciencia de Materials de Barcelona (ICMAB-CSIC), Campus de la UAB, Bellaterra, Spain.
| | - Miquel Nuez-Martínez
- Institut de Ciencia de Materials de Barcelona (ICMAB-CSIC), Campus de la UAB, Bellaterra, Spain.
| | - Clara Viñas
- Institut de Ciencia de Materials de Barcelona (ICMAB-CSIC), Campus de la UAB, Bellaterra, Spain.
| | - Francesc Teixidor
- Institut de Ciencia de Materials de Barcelona (ICMAB-CSIC), Campus de la UAB, Bellaterra, Spain.
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21
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Mani H, Chang CC, Hsu HJ, Yang CH, Yen JH, Liou JW. Comparison, Analysis, and Molecular Dynamics Simulations of Structures of a Viral Protein Modeled Using Various Computational Tools. Bioengineering (Basel) 2023; 10:1004. [PMID: 37760106 PMCID: PMC10525864 DOI: 10.3390/bioengineering10091004] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2023] [Revised: 08/16/2023] [Accepted: 08/22/2023] [Indexed: 09/29/2023] Open
Abstract
The structural analysis of proteins is a major domain of biomedical research. Such analysis requires resolved three-dimensional structures of proteins. Advancements in computer technology have led to progress in biomedical research. In silico prediction and modeling approaches have facilitated the construction of protein structures, with or without structural templates. In this study, we used three neural network-based de novo modeling approaches-AlphaFold2 (AF2), Robetta-RoseTTAFold (Robetta), and transform-restrained Rosetta (trRosetta)-and two template-based tools-the Molecular Operating Environment (MOE) and iterative threading assembly refinement (I-TASSER)-to construct the structure of a viral capsid protein, hepatitis C virus core protein (HCVcp), whose structure have not been fully resolved by laboratory techniques. Templates with sufficient sequence identity for the homology modeling of complete HCVcp are currently unavailable. Therefore, we performed domain-based homology modeling for MOE simulations. The templates for each domain were obtained through sequence-based searches on NCBI and the Protein Data Bank. Then, the modeled domains were assembled to construct the complete structure of HCVcp. The full-length structure and two truncated forms modeled using various computational tools were compared. Molecular dynamics (MD) simulations were performed to refine the structures. The root mean square deviation of backbone atoms, root mean square fluctuation of Cα atoms, and radius of gyration were calculated to monitor structural changes and convergence in the simulations. The model quality was evaluated through ERRAT and phi-psi plot analysis. In terms of the initial prediction for protein modeling, Robetta and trRosetta outperformed AF2. Regarding template-based tools, MOE outperformed I-TASSER. MD simulations resulted in compactly folded protein structures, which were of good quality and theoretically accurate. Thus, the predicted structures of certain proteins must be refined to obtain reliable structural models. MD simulation is a promising tool for this purpose.
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Affiliation(s)
- Hemalatha Mani
- Institute of Medical Sciences, Tzu Chi University, Hualien 97004, Taiwan
| | - Chun-Chun Chang
- Department of Laboratory Medicine, Hualien Tzu Chi Hospital, Buddhist Tzu Chi Medical Foundation, Hualien 97004, Taiwan
- Department of Laboratory Medicine and Biotechnology, Tzu Chi University, Hualien 97004, Taiwan
| | - Hao-Jen Hsu
- Department of Biomedical Sciences and Engineering, Tzu Chi University, Hualien 97004, Taiwan
| | - Chin-Hao Yang
- Department of Biochemistry, School of Medicine, Tzu Chi University, Hualien 97004, Taiwan
| | - Jui-Hung Yen
- Department of Molecular Biology and Human Genetics, Tzu Chi University, Hualien 97004, Taiwan
| | - Je-Wen Liou
- Institute of Medical Sciences, Tzu Chi University, Hualien 97004, Taiwan
- Department of Laboratory Medicine and Biotechnology, Tzu Chi University, Hualien 97004, Taiwan
- Department of Biochemistry, School of Medicine, Tzu Chi University, Hualien 97004, Taiwan
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22
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Roterman I, Stapor K, Konieczny L. Engagement of intrinsic disordered proteins in protein-protein interaction. Front Mol Biosci 2023; 10:1230922. [PMID: 37583961 PMCID: PMC10423874 DOI: 10.3389/fmolb.2023.1230922] [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: 05/29/2023] [Accepted: 07/11/2023] [Indexed: 08/17/2023] Open
Abstract
Proteins from the intrinsically disordered group (IDP) focus the attention of many researchers engaged in protein structure analysis. The main criteria used in their identification are lack of secondary structure and significant structural variability. This variability takes forms that cannot be identified in the X-ray technique. In the present study, different criteria were used to assess the status of IDP proteins and their fragments recognized as intrinsically disordered regions (IDRs). The status of the hydrophobic core in proteins identified as IDPs and in their complexes was assessed. The status of IDRs as components of the ordering structure resulting from the construction of the hydrophobic core was also assessed. The hydrophobic core is understood as a structure encompassing the entire molecule in the form of a centrally located high concentration of hydrophobicity and a shell with a gradually decreasing level of hydrophobicity until it reaches a level close to zero on the protein surface. It is a model assuming that the protein folding process follows a micellization pattern aiming at exposing polar residues on the surface, with the simultaneous isolation of hydrophobic amino acids from the polar aquatic environment. The use of the model of hydrophobicity distribution in proteins in the form of the 3D Gaussian distribution described on the protein particle introduces the possibility of assessing the degree of similarity to the assumed micelle-like distribution and also enables the identification of deviations and mismatch between the actual distribution and the idealized distribution. The FOD (fuzzy oil drop) model and its modified FOD-M version allow for the quantitative assessment of these differences and the assessment of the relationship of these areas to the protein function. In the present work, the sections of IDRs in protein complexes classified as IDPs are analyzed. The classification "disordered" in the structural sense (lack of secondary structure or high flexibility) does not always entail a mismatch with the structure of the hydrophobic core. Particularly, the interface area, often consisting of IDRs, in many analyzed complexes shows the compliance of the hydrophobicity distribution with the idealized distribution, which proves that matching to the structure of the hydrophobic core does not require secondary structure ordering.
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Affiliation(s)
- Irena Roterman
- Department of Bioinformatics and Telemedicine, Jagiellonian University—Medical College, Kraków, Poland
| | - Katarzyna Stapor
- Department of Applied Informatics, Faculty of Automatic, Electronics and Computer Science, Silesian University of Technology, Gliwice, Poland
| | - Leszek Konieczny
- Chair of Medical Biochemistry, Medical College, Jagiellonian University, Kraków, Poland
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23
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Yuce-Erarslan E, Domb AAJ, Kasem H, Uversky VN, Coskuner-Weber O. Intrinsically Disordered Synthetic Polymers in Biomedical Applications. Polymers (Basel) 2023; 15:polym15102406. [PMID: 37242981 DOI: 10.3390/polym15102406] [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: 03/16/2023] [Revised: 04/29/2023] [Accepted: 05/18/2023] [Indexed: 05/28/2023] Open
Abstract
In biology and medicine, intrinsically disordered synthetic polymers bio-mimicking intrinsically disordered proteins, which lack stable three-dimensional structures, possess high structural/conformational flexibility. They are prone to self-organization and can be extremely useful in various biomedical applications. Among such applications, intrinsically disordered synthetic polymers can have potential usage in drug delivery, organ transplantation, artificial organ design, and immune compatibility. The designing of new syntheses and characterization mechanisms is currently required to provide the lacking intrinsically disordered synthetic polymers for biomedical applications bio-mimicked using intrinsically disordered proteins. Here, we present our strategies for designing intrinsically disordered synthetic polymers for biomedical applications based on bio-mimicking intrinsically disordered proteins.
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Affiliation(s)
- Elif Yuce-Erarslan
- Chemical Engineering, Istanbul University-Cerrahpasa, Avcilar, Istanbul 34320, Turkey
| | - Abraham Avi J Domb
- School of Pharmacy, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem 91120, Israel
| | - Haytam Kasem
- Azrieli College of Engineering, 26 Ya'akov Schreiboim Street, Jerusalem 9103501, Israel
| | - Vladimir N Uversky
- Department of Molecular Medicine, Morsani College of Medicine, University of South Florida, Tampa, FL 33612, USA
| | - Orkid Coskuner-Weber
- Molecular Biotechnology, Turkish-German University, Sahinkaya Caddesi, No. 106, Beykoz, Istanbul 34820, Turkey
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24
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Priyadarshi A, Devi HM, Swaminathan R. Disruption of Spatial Proximities among Charged Groups in Equilibrium-Denatured States of Proteins Tracked Using Protein Charge Transfer Spectra. Biochemistry 2023. [PMID: 37162303 DOI: 10.1021/acs.biochem.3c00006] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 05/11/2023]
Abstract
The absorption and luminescence originating from protein charge transfer spectra (ProCharTS) depend on the proximity between multiple charged groups in a protein. This makes ProCharTS absorbance/luminescence intensity a sensitive probe for detecting changes in the protein structure, which alter the proximity among charged groups in the protein. In this work, ProCharTS absorbance of charge-rich proteins like human serum albumin (HSA), α3C, and α3W was used to monitor structural changes upon chemical denaturant-induced protein unfolding under equilibrium conditions. The denaturation midpoints were estimated using nonlinear regression analysis. For HSA, absorbance at 325 and 340 nm estimated the GdnHCl-induced denaturation midpoints to be 0.80 and 0.61 M, respectively. A similar analysis of α3C and α3W ProCharTS absorbance yielded denaturation midpoints of 0.88 and 0.86 M at 325 nm and 0.96 and 0.66 M at 340 nm, respectively. A previously reported molten globule-like state in the GdnHCl-induced HSA unfolding pathway was detected by the increase in HSA ProCharTS absorbance at 0.5 M GdnHCl. To validate the above results, protein unfolding was additionally monitored using conventional methods like circular dichroism (CD), Trp, and dansyl fluorescence. Our results suggest that disruption of charged amino acid sidechain contacts as revealed by ProCharTS occurs at lower denaturant concentrations compared to the loss of secondary/folded structure monitored by CD and fluorescence. Further, HSA ProCharTS absorbance at 315-340 nm revealed that tertiary contacts among charged residues were disrupted at lower GdnHCl concentrations compared to sequence adjacent contacts. Our data underscore the utility of ProCharTS as a novel label-free tool to track unfolding in charge-rich proteins.
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Affiliation(s)
- Anurag Priyadarshi
- Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati 781 039, Assam, India
| | - Himanshi Maniram Devi
- Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati 781 039, Assam, India
| | - Rajaram Swaminathan
- Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati 781 039, Assam, India
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25
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Pelham JF, Mosier AE, Altshuler SC, Rhodes ML, Kirchhoff CL, Fall WB, Mann C, Baik LS, Chiu JC, Hurley JM. Conformational changes in the negative arm of the circadian clock correlate with dynamic interactomes involved in post-transcriptional regulation. Cell Rep 2023; 42:112376. [PMID: 37043358 PMCID: PMC10562519 DOI: 10.1016/j.celrep.2023.112376] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/07/2021] [Revised: 09/16/2022] [Accepted: 03/24/2023] [Indexed: 04/13/2023] Open
Abstract
Biology is tuned to the Earth's diurnal cycle by the circadian clock, a transcriptional/translational negative feedback loop that regulates physiology via transcriptional activation and other post-transcriptional mechanisms. We hypothesize that circadian post-transcriptional regulation might stem from conformational shifts in the intrinsically disordered proteins that comprise the negative arm of the feedback loop to coordinate variation in negative-arm-centered macromolecular complexes. This work demonstrates temporal conformational fluidity in the negative arm that correlates with 24-h variation in physiologically diverse macromolecular complex components in eukaryotic clock proteins. Short linear motifs on the negative-arm proteins that correspond with the interactors localized to disordered regions and known temporal phosphorylation sites suggesting changes in these macromolecular complexes could be due to conformational changes imparted by the temporal phospho-state. Interactors that oscillate in the macromolecular complexes over circadian time correlate with post-transcriptionally regulated proteins, highlighting how time-of-day variation in the negative-arm protein complexes may tune cellular physiology.
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Affiliation(s)
- Jacqueline F Pelham
- Department of Biological Sciences, Rensselaer Polytechnic Institute, Troy, NY 12180, USA
| | - Alexander E Mosier
- Department of Biological Sciences, Rensselaer Polytechnic Institute, Troy, NY 12180, USA
| | - Samuel C Altshuler
- Department of Biological Sciences, Rensselaer Polytechnic Institute, Troy, NY 12180, USA
| | - Morgan L Rhodes
- Department of Biological Sciences, Rensselaer Polytechnic Institute, Troy, NY 12180, USA
| | | | - William B Fall
- Department of Biological Sciences, Rensselaer Polytechnic Institute, Troy, NY 12180, USA
| | - Catherine Mann
- Department of Biological Sciences, Rensselaer Polytechnic Institute, Troy, NY 12180, USA
| | - Lisa S Baik
- Department of Entomology and Nematology, University of California, Davis, Davis, CA 95616, USA
| | - Joanna C Chiu
- Department of Entomology and Nematology, University of California, Davis, Davis, CA 95616, USA
| | - Jennifer M Hurley
- Department of Biological Sciences, Rensselaer Polytechnic Institute, Troy, NY 12180, USA; Center for Biotechnology and Interdisciplinary Sciences, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
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26
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Zhu JJ, Zhang NJ, Wei T, Chen HF. Enhancing Conformational Sampling for Intrinsically Disordered and Ordered Proteins by Variational Autoencoder. Int J Mol Sci 2023; 24:ijms24086896. [PMID: 37108059 PMCID: PMC10138423 DOI: 10.3390/ijms24086896] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/03/2023] [Revised: 03/26/2023] [Accepted: 03/27/2023] [Indexed: 04/29/2023] Open
Abstract
Intrinsically disordered proteins (IDPs) account for more than 50% of the human proteome and are closely associated with tumors, cardiovascular diseases, and neurodegeneration, which have no fixed three-dimensional structure under physiological conditions. Due to the characteristic of conformational diversity, conventional experimental methods of structural biology, such as NMR, X-ray diffraction, and CryoEM, are unable to capture conformational ensembles. Molecular dynamics (MD) simulation can sample the dynamic conformations at the atomic level, which has become an effective method for studying the structure and function of IDPs. However, the high computational cost prevents MD simulations from being widely used for IDPs conformational sampling. In recent years, significant progress has been made in artificial intelligence, which makes it possible to solve the conformational reconstruction problem of IDP with fewer computational resources. Here, based on short MD simulations of different IDPs systems, we use variational autoencoders (VAEs) to achieve the generative reconstruction of IDPs structures and include a wider range of sampled conformations from longer simulations. Compared with the generative autoencoder (AEs), VAEs add an inference layer between the encoder and decoder in the latent space, which can cover the conformational landscape of IDPs more comprehensively and achieve the effect of enhanced sampling. Through experimental verification, the Cα RMSD between VAE-generated and MD simulation sampling conformations in the 5 IDPs test systems was significantly lower than that of AE. The Spearman correlation coefficient on the structure was higher than that of AE. VAE can also achieve excellent performance regarding structured proteins. In summary, VAEs can be used to effectively sample protein structures.
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Affiliation(s)
- Jun-Jie Zhu
- State Key Laboratory of Microbial Metabolism, Joint International Research Laboratory of Metabolic & Developmental Sciences, Department of Bioinformatics and Biostatistics, National Experimental Teaching Center for Life Sciences and Biotechnology, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China
| | - Ning-Jie Zhang
- State Key Laboratory of Microbial Metabolism, Joint International Research Laboratory of Metabolic & Developmental Sciences, Department of Bioinformatics and Biostatistics, National Experimental Teaching Center for Life Sciences and Biotechnology, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China
| | - Ting Wei
- State Key Laboratory of Microbial Metabolism, Joint International Research Laboratory of Metabolic & Developmental Sciences, Department of Bioinformatics and Biostatistics, National Experimental Teaching Center for Life Sciences and Biotechnology, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China
| | - Hai-Feng Chen
- State Key Laboratory of Microbial Metabolism, Joint International Research Laboratory of Metabolic & Developmental Sciences, Department of Bioinformatics and Biostatistics, National Experimental Teaching Center for Life Sciences and Biotechnology, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China
- Shanghai Center for Bioinformation Technology, Shanghai 200240, China
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27
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Chae MK, Lee NK, Jung Y, Johner A. Shape Fluctuations of Random Polyampholyte and Intrinsically Disordered Protein Sequences. Macromolecules 2023. [DOI: 10.1021/acs.macromol.2c02164] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/01/2023]
Affiliation(s)
- Min-Kyung Chae
- Department of Physics and Astronomy, Sejong University, Seoul05006, Korea
- National Institute for Mathematical Sciences, Daejeon34047, South Korea
| | - Nam-Kyung Lee
- Department of Physics and Astronomy, Sejong University, Seoul05006, Korea
| | - Youngkyun Jung
- Supercomputing Center, Korea Institute of Science and Technology Information, Daejeon34141, Korea
| | - Albert Johner
- Institut Charles Sadron CNRS─Unistra, Universite de Strasbourg, F-67000Strasbourg, France
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28
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Vovk A, Zilman A. Effects of Sequence Composition, Patterning and Hydrodynamics on the Conformation and Dynamics of Intrinsically Disordered Proteins. Int J Mol Sci 2023; 24:1444. [PMID: 36674958 PMCID: PMC9867189 DOI: 10.3390/ijms24021444] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2022] [Revised: 12/24/2022] [Accepted: 12/25/2022] [Indexed: 01/13/2023] Open
Abstract
Intrinsically disordered proteins (IDPs) and intrinsically disordered regions (IDRs) perform diverse functions in cellular organization, transport and signaling. Unlike the well-defined structures of the classical natively folded proteins, IDPs and IDRs dynamically span large conformational and structural ensembles. This dynamic disorder impedes the study of the relationship between the amino acid sequences of the IDPs and their spatial structures and dynamics, with different experimental techniques often offering seemingly contradictory results. Although experimental and theoretical evidence indicates that some IDP properties can be understood based on their average biophysical properties and amino acid composition, other aspects of IDP function are dictated by the specifics of the amino acid sequence. We investigate the effects of several key variables on the dimensions and the dynamics of IDPs using coarse-grained polymer models. We focus on the sequence "patchiness" informed by the sequence and biophysical properties of different classes of IDPs-and in particular FG nucleoporins of the nuclear pore complex (NPC). We show that the sequence composition and patterning are well reflected in the global conformational variables such as the radius of gyration and hydrodynamic radius, while the end-to-end distance and dynamics are highly sequence-specific. We find that in good solvent conditions highly heterogeneous sequences of IDPs can be well mapped onto averaged minimal polymer models for the purpose of prediction of the IDPs dimensions and dynamic relaxation times. The coarse-grained simulations are in a good agreement with the results of atomistic MD. We discuss the implications of these results for the interpretation of the recent experimental measurements, and for the further applications of mesoscopic models of FG nucleoporins and IDPs more broadly.
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Affiliation(s)
- Andrei Vovk
- Department of Physics, University of Toronto, 60 St George Street, Toronto, ON M1M 2P7, Canada
| | - Anton Zilman
- Department of Physics, University of Toronto, 60 St George Street, Toronto, ON M1M 2P7, Canada
- Institute for Biomedical Engineering, University of Toronto, 164 College Street, Toronto, ON M5S 3G9, Canada
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29
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Ehm T, Shinar H, Jacoby G, Meir S, Koren G, Segal Asher M, Korpanty J, Thompson MP, Gianneschi NC, Kozlov MM, Azoulay-Ginsburg S, Amir RJ, Rädler JO, Beck R. Self-Assembly of Tunable Intrinsically Disordered Peptide Amphiphiles. Biomacromolecules 2023; 24:98-108. [PMID: 36469950 PMCID: PMC9832477 DOI: 10.1021/acs.biomac.2c00866] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Intrinsically disordered peptide amphiphiles (IDPAs) present a novel class of synthetic conjugates that consist of short hydrophilic polypeptides anchored to hydrocarbon chains. These hybrid polymer-lipid block constructs spontaneously self-assemble into dispersed nanoscopic aggregates or ordered mesophases in aqueous solution due to hydrophobic interactions. Yet, the possible sequence variations and their influence on the self-assembly structures are vast and have hardly been explored. Here, we measure the nanoscopic self-assembled structures of four IDPA systems that differ by their amino acid sequence. We show that permutations in the charge pattern along the sequence remarkably alter the headgroup conformation and consequently alter the pH-triggered phase transitions between spherical, cylindrical micelles and hexagonal condensed phases. We demonstrate that even a single amino acid mutation is sufficient to tune structural transitions in the condensed IDPA mesophases, while peptide conformations remain unfolded and disordered. Furthermore, alteration of the peptide sequence can render IDPAs to become susceptible to enzymatic cleavage and induce enzymatically activated phase transitions. These results hold great potential for embedding multiple functionalities into lipid nanoparticle delivery systems by incorporating IDPAs with the desired properties.
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Affiliation(s)
- Tamara Ehm
- Raymond
& Beverly Sackler School of Physics & Astronomy, Tel Aviv University, Tel Aviv 6997801, Israel,Faculty
of Physics and Center for NanoScience, Ludwig-Maximilians-Universität, MünchenD-80539, Germany,The
Center for Physics & Chemistry of Living Systems, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for NanoTechnology & NanoScience, Tel Aviv University, Tel Aviv 6997801, Israel
| | - Hila Shinar
- Raymond
& Beverly Sackler School of Physics & Astronomy, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for Physics & Chemistry of Living Systems, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for NanoTechnology & NanoScience, Tel Aviv University, Tel Aviv 6997801, Israel
| | - Guy Jacoby
- Raymond
& Beverly Sackler School of Physics & Astronomy, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for Physics & Chemistry of Living Systems, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for NanoTechnology & NanoScience, Tel Aviv University, Tel Aviv 6997801, Israel
| | - Sagi Meir
- Raymond
& Beverly Sackler School of Physics & Astronomy, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for Physics & Chemistry of Living Systems, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for NanoTechnology & NanoScience, Tel Aviv University, Tel Aviv 6997801, Israel
| | - Gil Koren
- Raymond
& Beverly Sackler School of Physics & Astronomy, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for Physics & Chemistry of Living Systems, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for NanoTechnology & NanoScience, Tel Aviv University, Tel Aviv 6997801, Israel
| | - Merav Segal Asher
- The
Center for NanoTechnology & NanoScience, Tel Aviv University, Tel Aviv 6997801, Israel,Raymond
& Beverly Sackler School of Chemistry, Tel Aviv University, Tel Aviv 6997801, Israel
| | - Joanna Korpanty
- Department
of Chemistry, International Institute for Nanotechnology, Chemistry
of Life Processes Institute, Simpson Querrey Institute, Northwestern University, Evanston, Illinois 60208, United States
| | - Matthew P. Thompson
- Department
of Chemistry, International Institute for Nanotechnology, Chemistry
of Life Processes Institute, Simpson Querrey Institute, Northwestern University, Evanston, Illinois 60208, United States
| | - Nathan C. Gianneschi
- Department
of Chemistry, International Institute for Nanotechnology, Chemistry
of Life Processes Institute, Simpson Querrey Institute, Northwestern University, Evanston, Illinois 60208, United States,Department
of Materials Science & Engineering, Department of Biomedical Engineering
and Department of Pharmacology, Northwestern
University, Evanston, Illinois 60208, United States
| | - Michael M. Kozlov
- The
Center for Physics & Chemistry of Living Systems, Tel Aviv University, Tel Aviv 6997801, Israel,Raymond
& Beverly Sackler School of Medicine, Tel Aviv University, Tel Aviv 6997801, Israel
| | - Salome Azoulay-Ginsburg
- Raymond
& Beverly Sackler School of Chemistry, Tel Aviv University, Tel Aviv 6997801, Israel
| | - Roey J. Amir
- The
Center for Physics & Chemistry of Living Systems, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for NanoTechnology & NanoScience, Tel Aviv University, Tel Aviv 6997801, Israel,Raymond
& Beverly Sackler School of Chemistry, Tel Aviv University, Tel Aviv 6997801, Israel,The
ADAMA Center for Novel Delivery Systems in Crop Protection, Tel Aviv University, Tel Aviv 6997801, Israel,Email
| | - Joachim O. Rädler
- Faculty
of Physics and Center for NanoScience, Ludwig-Maximilians-Universität, MünchenD-80539, Germany,
| | - Roy Beck
- Raymond
& Beverly Sackler School of Physics & Astronomy, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for Physics & Chemistry of Living Systems, Tel Aviv University, Tel Aviv 6997801, Israel,The
Center for NanoTechnology & NanoScience, Tel Aviv University, Tel Aviv 6997801, Israel,
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30
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Rapid Scan Electron Paramagnetic Resonance Spectroscopy Is a Suitable Tool to Study Intermolecular Interactions of Intrinsically Disordered Protein. BIOLOGY 2023; 12:biology12010079. [PMID: 36671771 PMCID: PMC9856040 DOI: 10.3390/biology12010079] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 12/05/2022] [Revised: 12/21/2022] [Accepted: 12/22/2022] [Indexed: 01/06/2023]
Abstract
Intrinsically disordered proteins (IDPs) are involved in most crucial cellular processes. However, they lack a well-defined fold hampering the investigation of their structural ensemble and interactions. Suitable biophysical methods able to manage their inherent flexibility and broad conformational ensemble are scarce. Here, we used rapid scan (RS) electron paramagnetic resonance (EPR) spectroscopy to study the intermolecular interactions of the IDP α-synuclein (aS). aS aggregation and fibril deposition is the hallmark of Parkinson's disease, and specific point mutations, among them A30P and A53T, were linked to the early onset of the disease. To understand the pathological processes, research intensively investigates aS aggregation kinetics, which was reported to be accelerated in the presence of ethanol. Conventional techniques fail to capture these fast processes due to their limited time resolution and, thus, lose kinetic information. We have demonstrated that RS EPR spectroscopy is suitable for studying aS aggregation by resolving underlying kinetics and highlighting differences in fibrillization behavior. RS EPR spectroscopy outperforms traditional EPR methods in terms of sensitivity by a factor of 5 in our case while significantly reducing data acquisition time. Thus, we were able to sample short time intervals capturing single events taking place during the aggregation process. Further studies will therefore be able to shed light on biological processes proceeding on fast time scales.
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31
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Liu AY, Minetti CA, Remeta DP, Breslauer KJ, Chen KY. HSF1, Aging, and Neurodegeneration. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2023; 1409:23-49. [PMID: 35995906 DOI: 10.1007/5584_2022_733] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
Heat shock factor 1 (HSF1) is a master transcription regulator that mediates the induction of heat shock protein chaperones for quality control (QC) of the proteome and maintenance of proteostasis as a protective mechanism in response to stress. Research in this particular area has accelerated dramatically over the past three decades following successful isolation, cloning, and characterization of HSF1. The intricate multi-protein complexes and transcriptional activation orchestrated by HSF1 are fundamental processes within the cellular QC machinery. Our primary focus is on the regulation and function of HSF1 in aging and neurodegenerative diseases (ND) which represent physiological and pathological states of dysfunction in protein QC. This chapter presents an overview of HSF1 structural, functional, and energetic properties in healthy cells while addressing the deterioration of HSF1 function viz-à-viz age-dependent and neuron-specific vulnerability to ND. We discuss the structural domains of HSF1 with emphasis on the intrinsically disordered regions and note that disease proteins associated with ND are often structurally disordered and exquisitely sensitive to changes in cellular environment as may occur during aging. We propose a hypothesis that age-dependent changes of the intrinsically disordered proteome likely hold answers to understand many of the functional, structural, and organizational changes of proteins and signaling pathways in aging - dysfunction of HSF1 and accumulation of disease protein aggregates in ND included.Structured AbstractsIntroduction: Heat shock factor 1 (HSF1) is a master transcription regulator that mediates the induction of heat shock protein chaperones for quality control (QC) of the proteome as a cyto-protective mechanism in response to stress. There is cumulative evidence of age-related deterioration of this QC mechanism that contributes to disease vulnerability. OBJECTIVES Herein we discuss the regulation and function of HSF1 as they relate to the pathophysiological changes of protein quality control in aging and neurodegenerative diseases (ND). METHODS We present an overview of HSF1 structural, functional, and energetic properties in healthy cells while addressing the deterioration of HSF1 function vis-à-vis age-dependent and neuron-specific vulnerability to neurodegenerative diseases. RESULTS We examine the impact of intrinsically disordered regions on the function of HSF1 and note that proteins associated with neurodegeneration are natively unstructured and exquisitely sensitive to changes in cellular environment as may occur during aging. CONCLUSIONS We put forth a hypothesis that age-dependent changes of the intrinsically disordered proteome hold answers to understanding many of the functional, structural, and organizational changes of proteins - dysfunction of HSF1 in aging and appearance of disease protein aggregates in neurodegenerative diseases included.
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Affiliation(s)
- Alice Y Liu
- Department of Cell Biology and Neuroscience, Rutgers The State University of New Jersey, Piscataway, NJ, USA.
- Rutgers Cancer Institute of New Jersey, Rutgers University, New Brunswick, NJ, USA.
| | - Conceição A Minetti
- Department of Chemistry and Chemical Biology, Rutgers The State University of New Jersey, Piscataway, NJ, USA
| | - David P Remeta
- Department of Chemistry and Chemical Biology, Rutgers The State University of New Jersey, Piscataway, NJ, USA
| | - Kenneth J Breslauer
- Rutgers Cancer Institute of New Jersey, Rutgers University, New Brunswick, NJ, USA
- Department of Chemistry and Chemical Biology, Rutgers The State University of New Jersey, Piscataway, NJ, USA
| | - Kuang Yu Chen
- Department of Chemistry and Chemical Biology, Rutgers The State University of New Jersey, Piscataway, NJ, USA
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32
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Intrinsically Disordered Proteins: An Overview. Int J Mol Sci 2022; 23:ijms232214050. [PMID: 36430530 PMCID: PMC9693201 DOI: 10.3390/ijms232214050] [Citation(s) in RCA: 51] [Impact Index Per Article: 17.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/04/2022] [Revised: 11/07/2022] [Accepted: 11/08/2022] [Indexed: 11/16/2022] Open
Abstract
Many proteins and protein segments cannot attain a single stable three-dimensional structure under physiological conditions; instead, they adopt multiple interconverting conformational states. Such intrinsically disordered proteins or protein segments are highly abundant across proteomes, and are involved in various effector functions. This review focuses on different aspects of disordered proteins and disordered protein regions, which form the basis of the so-called "Disorder-function paradigm" of proteins. Additionally, various experimental approaches and computational tools used for characterizing disordered regions in proteins are discussed. Finally, the role of disordered proteins in diseases and their utility as potential drug targets are explored.
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33
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Hou Y, Kumar P, Aggarwal M, Sarkari F, Wolcott KM, Chattoraj DK, Crooke E, Saxena R. The linker domain of the initiator DnaA contributes to its ATP binding and membrane association in E. coli chromosomal replication. SCIENCE ADVANCES 2022; 8:eabq6657. [PMID: 36197974 PMCID: PMC9534497 DOI: 10.1126/sciadv.abq6657] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/22/2022] [Accepted: 08/18/2022] [Indexed: 06/16/2023]
Abstract
DnaA, the initiator of Escherichia coli chromosomal replication, has in its adenosine triphosphatase (ATPase) domain residues required for adenosine 5'-triphosphate (ATP) binding and membrane attachment. Here, we show that D118Q substitution in the DnaA linker domain, a domain known to be without major regulatory functions, influences ATP binding of DnaA and replication initiation in vivo. Although initiation defective by itself, overexpression of DnaA(D118Q) caused overinitiation of replication in dnaA46ts cells and prevented cell growth. The growth defect was rescued by overexpressing the initiation inhibitor, SeqA, indicating that the growth inhibition resulted from overinitiation. Small deletions within the linker showed another unexpected phenotype: cellular growth without requiring normal levels of anionic membrane lipids, a property found in DnaA mutated in its ATPase domain. The deleted proteins were defective in association with anionic membrane vesicles. These results show that changes in the linker domain can alter DnaA functions similarly to the previously shown changes in the ATPase domain.
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Affiliation(s)
- Yanqi Hou
- Department of Biochemistry and Molecular and Cellular Biology, Georgetown University Medical Center, Washington, DC 20007, USA
| | - Pankaj Kumar
- Center for Tuberculosis Research, Department of Medicine, Johns Hopkins University, Baltimore, MD 21287, USA
| | - Monika Aggarwal
- Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, Washington, DC 20007, USA
| | - Farzad Sarkari
- Department of Biochemistry and Molecular and Cellular Biology, Georgetown University Medical Center, Washington, DC 20007, USA
| | - Karen M. Wolcott
- Laboratory of Genome Integrity, Flow Cytometry Core Facility, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Dhruba K. Chattoraj
- Basic Research Laboratory, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Elliott Crooke
- Department of Biochemistry and Molecular and Cellular Biology, Georgetown University Medical Center, Washington, DC 20007, USA
- Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, Washington, DC 20007, USA
| | - Rahul Saxena
- Department of Biochemistry and Molecular and Cellular Biology, Georgetown University Medical Center, Washington, DC 20007, USA
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34
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Chaudhary A, Chaurasia PK, Kushwaha S, Chauhan P, Chawade A, Mani A. Correlating multi-functional role of cold shock domain proteins with intrinsically disordered regions. Int J Biol Macromol 2022; 220:743-753. [PMID: 35987358 DOI: 10.1016/j.ijbiomac.2022.08.100] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/09/2022] [Revised: 07/26/2022] [Accepted: 08/14/2022] [Indexed: 11/05/2022]
Abstract
Cold shock proteins (CSPs) are an ancient and conserved family of proteins. They are renowned for their role in response to low-temperature stress in bacteria and nucleic acid binding activities. In prokaryotes, cold and non-cold inducible CSPs are involved in various cellular and metabolic processes such as growth and development, osmotic oxidation, starvation, stress tolerance, and host cell invasion. In prokaryotes, cold shock condition reduces cell transcription and translation efficiency. Eukaryotic cold shock domain (CSD) proteins are evolved form of prokaryotic CSPs where CSD is flanked by N- and C-terminal domains. Eukaryotic CSPs are multi-functional proteins. CSPs also act as nucleic acid chaperons by preventing the formation of secondary structures in mRNA at low temperatures. In human, CSD proteins play a crucial role in the progression of breast cancer, colon cancer, lung cancer, and Alzheimer's disease. A well-defined three-dimensional structure of intrinsically disordered regions of CSPs family members is still undetermined. In this article, intrinsic disorder regions of CSPs have been explored systematically to understand the pleiotropic role of the cold shock family of proteins.
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Affiliation(s)
- Amit Chaudhary
- Department of Metallurgical Engineering & Materials Science, Indian Institute of Technology Bombay
| | - Pankaj Kumar Chaurasia
- PG Department of Chemistry, L.S. College, Babasaheb Bhimrao Ambedkar Bihar University, Muzaffarpur, Bihar 842001, India
| | - Sandeep Kushwaha
- National Institute of Animal Biotechnology, Hyderabad 500032, India.
| | | | - Aakash Chawade
- Department of Plant Breeding, Swedish University of Agricultural Sciences, 230 53 Alnarp, Sweden.
| | - Ashutosh Mani
- Department of Biotechnology, Motilal Nehru National Institute of Technology Allahabad, Prayagraj 211004, India.
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35
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Smyth S, Zhang Z, Bah A, Tsangaris TE, Dawson J, Forman-Kay JD, Gradinaru CC. Multisite phosphorylation and binding alter conformational dynamics of the 4E-BP2 protein. Biophys J 2022; 121:3049-3060. [PMID: 35841142 PMCID: PMC9463650 DOI: 10.1016/j.bpj.2022.07.015] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/15/2022] [Revised: 05/19/2022] [Accepted: 07/11/2022] [Indexed: 11/02/2022] Open
Abstract
Intrinsically disordered proteins (IDPs) play critical roles in regulatory protein interactions, but detailed structural/dynamic characterization of their ensembles remain challenging, both in isolation and when they form dynamic "fuzzy" complexes. Such is the case for mRNA cap-dependent translation initiation, which is regulated by the interaction of the predominantly folded eukaryotic initiation factor 4E (eIF4E) with the intrinsically disordered eIF4E binding proteins (4E-BPs) in a phosphorylation-dependent manner. Single-molecule Förster resonance energy transfer showed that the conformational changes of 4E-BP2 induced by binding to eIF4E are non-uniform along the sequence; while a central region containing both motifs that bind to eIF4E expands and becomes stiffer, the C-terminal region is less affected. Fluorescence anisotropy decay revealed a non-uniform segmental flexibility around six different labeling sites along the chain. Dynamic quenching of these fluorescent probes by intrinsic aromatic residues measured via fluorescence correlation spectroscopy report on transient intra- and inter-molecular contacts on nanosecond-to-microsecond timescales. Upon hyperphosphorylation, which induces folding of ∼40 residues in 4E-BP2, the quenching rates decreased at most labeling sites. The chain dynamics around sites in the C-terminal region far away from the two binding motifs significantly increased upon binding to eIF4E, suggesting that this region is also involved in the highly dynamic 4E-BP2:eIF4E complex. Our time-resolved fluorescence data paint a sequence-level rigidity map of three states of 4E-BP2 differing in phosphorylation or binding status and distinguish regions that form contacts with eIF4E. This study adds complementary structural and dynamics information to recent studies of 4E-BP2, and it constitutes an important step toward a mechanistic understanding of this important IDP via integrative modeling.
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Affiliation(s)
- Spencer Smyth
- Department of Physics, University of Toronto, Toronto, Ontario, Canada; Department of Chemical & Physical Sciences, University of Toronto Mississauga, Mississauga, Ontario, Canada
| | - Zhenfu Zhang
- Department of Physics, University of Toronto, Toronto, Ontario, Canada; Department of Chemical & Physical Sciences, University of Toronto Mississauga, Mississauga, Ontario, Canada
| | - Alaji Bah
- Program in Molecular Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Thomas E Tsangaris
- Department of Chemical & Physical Sciences, University of Toronto Mississauga, Mississauga, Ontario, Canada
| | - Jennifer Dawson
- Program in Molecular Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Julie D Forman-Kay
- Program in Molecular Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada; Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada
| | - Claudiu C Gradinaru
- Department of Physics, University of Toronto, Toronto, Ontario, Canada; Department of Chemical & Physical Sciences, University of Toronto Mississauga, Mississauga, Ontario, Canada.
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McFadden WM, Yanowitz JL. idpr: A package for profiling and analyzing Intrinsically Disordered Proteins in R. PLoS One 2022; 17:e0266929. [PMID: 35436286 PMCID: PMC9015136 DOI: 10.1371/journal.pone.0266929] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/06/2021] [Accepted: 03/29/2022] [Indexed: 12/23/2022] Open
Abstract
Intrinsically disordered proteins (IDPs) and intrinsically disordered regions (IDRs) are proteins or protein-domains that do not have a single native structure, rather, they are a class of flexible peptides that can rapidly adopt multiple conformations. IDPs are quite abundant, and their dynamic characteristics provide unique advantages for various biological processes. The field of “unstructured biology” has emerged, in part, because of numerous computational studies that had identified the unique characteristics of IDPs and IDRs. The package ‘idpr’, short for Intrinsically Disordered Proteins in R, implements several R functions that match the established characteristics of IDPs to protein sequences of interest. This includes calculations of residue composition, charge-hydropathy relationships, and predictions of intrinsic disorder. Additionally, idpr integrates several amino acid substitution matrices and calculators to supplement IDP-based workflows. Overall, idpr aims to integrate tools for the computational analysis of IDPs within R, facilitating the analysis of these important, yet under-characterized, proteins. The idpr package can be downloaded from Bioconductor (https://bioconductor.org/packages/idpr/).
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Affiliation(s)
| | - Judith L. Yanowitz
- Magee-Womens Research Institute, Pittsburgh, PA, United States of America
- Department of Obstetrics, Gynecology, and Reproductive Sciences, University of Pittsburgh School of Medicine, Pittsburgh, PA, United States of America
- * E-mail:
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Singh A, Kumar P, Sarvagalla S, Bharadwaj T, Nayak N, Coumar MS, Giri R, Garg N. Functional inhibition of c-Myc using novel inhibitors identified through “hot spot” targeting. J Biol Chem 2022; 298:101898. [PMID: 35378126 PMCID: PMC9065629 DOI: 10.1016/j.jbc.2022.101898] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2021] [Revised: 03/18/2022] [Accepted: 03/28/2022] [Indexed: 12/14/2022] Open
Abstract
Protein–protein interactions drive various biological processes in healthy as well as disease states. The transcription factor c-Myc plays a crucial role in maintaining cellular homeostasis, and its deregulated expression is linked to various human cancers; therefore, it can be considered a viable target for cancer therapeutics. However, the structural heterogeneity of c-Myc due to its disordered nature poses a major challenge to drug discovery. In the present study, we used an in silico alanine scanning mutagenesis approach to identify “hot spot” residues within the c-Myc/Myc-associated factor X interface, which is highly disordered and has not yet been systematically analyzed for potential small molecule binding sites. We then used the information gained from this analysis to screen potential inhibitors using a conformation ensemble approach. The fluorescence-based biophysical experiments showed that the identified hit molecules displayed noncovalent interactions with these hot spot residues, and further cell-based experiments showed substantial in vitro potency against diverse c-Myc-expressing cancer/stem cells by deregulating c-Myc activity. These biophysical and computational studies demonstrated stable binding of the hit compounds with the disordered c-Myc protein. Collectively, our data indicated effective drug targeting of the disordered c-Myc protein via the determination of hot spot residues in the c-Myc/Myc-associated factor X heterodimer.
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Poboinev VV, Khrustalev VV, Khrustaleva TA, Kasko TE, Popkov VD. The PentUnFOLD algorithm as a tool to distinguish the dark and the light sides of the structural instability of proteins. Amino Acids 2022; 54:1155-1171. [PMID: 35294674 PMCID: PMC8924573 DOI: 10.1007/s00726-022-03153-5] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2021] [Accepted: 02/14/2022] [Indexed: 12/12/2022]
Abstract
Intrinsically disordered proteins are frequently involved in important regulatory processes in the cell thanks to their ability to bind several different targets performing sometimes even opposite functions. The PentUnFOLD algorithm is a physicochemical method that is based on new propensity scales for disordered, nonstable and stable elements of secondary structure and on the counting of stabilizing and destabilizing intraprotein contacts. Unlike other methods, it works with a PDB file, and it can determine not only those fragments of alpha helices, beta strands, and random coils that can turn into disordered state (the “dark” side of the disorder), but also nonstable regions of alpha helices and beta strands which are able to turn into random coils (the “light” side), and vice versa (H ↔ C, E ↔ C). The scales have been obtained from structural data on disordered regions from the middle parts of amino acid sequences only, and not on their expectedly disordered N- and C-termini. Among other tendencies we have found that regions of both alpha helices and beta strands that can turn into the disordered state are relatively enriched in residues of Ala, Met, Asp, and Lys, while regions of both alpha helices and beta strands that can turn into random coil are relatively enriched in hydrophilic residues, and Cys, Pro, and Gly. Moreover, PentUnFOLD has the option to determine the effect of secondary structure transitions on the stability of a given region of a protein. The PentUnFOLD algorithm is freely available at http://3.17.12.213/pent-un-fold and http://chemres.bsmu.by/PentUnFOLD.htm.
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Affiliation(s)
| | | | - Tatyana Aleksandrovna Khrustaleva
- Biochemical Group of the Multidisciplinary Diagnostic Laboratory, Institute of Physiology of the National Academy of Sciences of Belarus, Minsk, Belarus
| | - Tihon Evgenyevich Kasko
- Department of General Chemistry, Belarusian State Medical University, Dzerzinskogo 83, Minsk, Belarus
| | - Vadim Dmitrievich Popkov
- Department of General Chemistry, Belarusian State Medical University, Dzerzinskogo 83, Minsk, Belarus
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Chae MK, Lee NK, Jung Y, Johner A, Joanny JF. Partially Globular Conformations from Random Charge Sequences. ACS Macro Lett 2022; 11:382-386. [PMID: 35575372 DOI: 10.1021/acsmacrolett.1c00655] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
Overall charged polymers with quenched charge sequences often adopt partially globular structures which result from the interplay between the disorder in charge sequences and thermal fluctuations. Simple energetic considerations show that structures consisting of alike (equal-size-equal-charge) globules are not favorable: the structures are intrinsically heterogeneous. We predict the globule distributions with the lowest energies in the size-charge space. The favorable structures comprise large (undercharged) and a majority of small (overcharged) globules. These distributions build a well characterized compact subset, which suggests some order. We also perform large scale molecular dynamics simulations on random quenched +/- sequences. Simulation results show that, despite disorder, the random charge sequences preferentially visit the predicted low energy structures and the predicted order emerges in the pearl-size distribution. This good agreement validates a posteriori the simple expression used for the energy. Implications for polyampholytes, polyelectrolytes, and intrinsically disordered proteins are discussed.
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Affiliation(s)
- Min-Kyung Chae
- Department of Physics and Astronomy, Sejong University, Seoul 05006, Korea
| | - Nam-Kyung Lee
- Department of Physics and Astronomy, Sejong University, Seoul 05006, Korea
| | - Youngkyun Jung
- Supercomputing Center, Korea Institute of Science and Technology Information, Daejeon 34141, Korea
| | - Albert Johner
- Université de Strasbourg, CNRS, Institut Charles Sadron (ICS), UPR 22, F-67000 Strasbourg, France
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40
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Naullage PM, Haghighatlari M, Namini A, Teixeira JMC, Li J, Zhang O, Gradinaru CC, Forman-Kay JD, Head-Gordon T. Protein Dynamics to Define and Refine Disordered Protein Ensembles. J Phys Chem B 2022; 126:1885-1894. [PMID: 35213160 PMCID: PMC10122607 DOI: 10.1021/acs.jpcb.1c10925] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
Intrinsically disordered proteins and unfolded proteins have fluctuating conformational ensembles that are fundamental to their biological function and impact protein folding, stability, and misfolding. Despite the importance of protein dynamics and conformational sampling, time-dependent data types are not fully exploited when defining and refining disordered protein ensembles. Here we introduce a computational framework using an elastic network model and normal-mode displacements to generate a dynamic disordered ensemble consistent with NMR-derived dynamics parameters, including transverse R2 relaxation rates and Lipari-Szabo order parameters (S2 values). We illustrate our approach using the unfolded state of the drkN SH3 domain to show that the dynamical ensembles give better agreement than a static ensemble for a wide range of experimental validation data including NMR chemical shifts, J-couplings, nuclear Overhauser effects, paramagnetic relaxation enhancements, residual dipolar couplings, hydrodynamic radii, single-molecule fluorescence Förster resonance energy transfer, and small-angle X-ray scattering.
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Affiliation(s)
- Pavithra M Naullage
- Pitzer Center for Theoretical Chemistry, University of California, Berkeley, California 94720, United States
- Department of Chemistry, University of California, Berkeley, California 94720, United States
| | - Mojtaba Haghighatlari
- Pitzer Center for Theoretical Chemistry, University of California, Berkeley, California 94720, United States
- Department of Chemistry, University of California, Berkeley, California 94720, United States
| | - Ashley Namini
- Molecular Medicine Program, Hospital for Sick Children, Toronto, Ontario M5G 0A4, Canada
| | - João M C Teixeira
- Molecular Medicine Program, Hospital for Sick Children, Toronto, Ontario M5G 0A4, Canada
| | - Jie Li
- Pitzer Center for Theoretical Chemistry, University of California, Berkeley, California 94720, United States
- Department of Chemistry, University of California, Berkeley, California 94720, United States
| | - Oufan Zhang
- Pitzer Center for Theoretical Chemistry, University of California, Berkeley, California 94720, United States
- Department of Chemistry, University of California, Berkeley, California 94720, United States
| | - Claudiu C Gradinaru
- Department of Chemical and Physical Sciences, University of Toronto Mississauga, Mississauga, Ontario L5L 1C6, Canada
| | - Julie D Forman-Kay
- Molecular Medicine Program, Hospital for Sick Children, Toronto, Ontario M5G 0A4, Canada
- Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada
| | - Teresa Head-Gordon
- Pitzer Center for Theoretical Chemistry, University of California, Berkeley, California 94720, United States
- Department of Chemistry, University of California, Berkeley, California 94720, United States
- Department of Chemical and Biomolecular Engineering, University of California, Berkeley, California 94720, United States
- Department of Bioengineering, University of California, Berkeley, California 94720, United States
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41
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Madeira PP, Rocha IL, Rosa ME, Freire MG, Coutinho JA. On the aggregation of bovine serum albumin. J Mol Liq 2022. [DOI: 10.1016/j.molliq.2021.118183] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/19/2022]
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42
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Wang D, Wu S, Wang D, Song X, Yang M, Zhang W, Huang S, Weng J, Liu Z, Wang W. The importance of the compact disordered state in the fuzzy interactions between intrinsically disordered proteins. Chem Sci 2022; 13:2363-2377. [PMID: 35310482 PMCID: PMC8864705 DOI: 10.1039/d1sc06825c] [Citation(s) in RCA: 17] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/07/2021] [Accepted: 01/21/2022] [Indexed: 11/23/2022] Open
Abstract
The intrinsically disordered C-terminal domain (CTD) of protein 4.1G is able to specifically bind a 26-residue intrinsically disordered region of NuMA, forming a dynamic fuzzy complex. As one of a few cases of extremely fuzzy interactions between two intrinsically disordered proteins/regions (IDPs/IDRs) without induced folding, the principle of the binding is unknown. Here, we combined experimental and computational methods to explore the detailed mechanism of the interaction between 4.1G-CTD and NuMA. MD simulations suggest that the kinetic hub states in the structure ensemble of 4.1G-CTD are favorable in the fuzzy complex. The feature of these hub states is that the binding 'hot spot' motifs βA and βB exhibit β strand propensities and are well packed to each other. The binding between 4.1G-CTD and NuMA is disrupted at low pH, which changes the intramolecular packing of 4.1G-CTD and weakens the packing between βA and βB motifs. Low pH conditions also lead to increased hydrodynamic radius and acceleration of backbone dynamics of 4.1G-CTD. All these results underscore the importance of tertiary structural arrangements and overall compactness of 4.1G-CTD in its binding to NuMA, i.e. the compact disordered state of 4.1G-CTD is crucial for binding. Different from the short linear motifs (SLiMs) that are often found to mediate IDP interactions, 4.1G-CTD functions as an intrinsically disordered domain (IDD), which is a functional and structural unit similar to conventional protein domains. This work sheds light on the molecular recognition mechanism of IDPs/IDRs and expands the conventional structure-function paradigm in protein biochemistry.
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Affiliation(s)
- Dan Wang
- Department of Chemistry, Multiscale Research Institute of Complex Systems and Institute of Biomedical Sciences, Fudan University Shanghai 200438 China
| | - Shaowen Wu
- Guangdong Key Laboratory for Crop Germplasm Resources Preservation and Utilization, Agro-biological Gene Research Center, Guangdong Academy of Agricultural Sciences Guangzhou 510640 Guangdong China
| | | | - Xingyu Song
- Department of Chemistry, Multiscale Research Institute of Complex Systems and Institute of Biomedical Sciences, Fudan University Shanghai 200438 China
| | - Maohua Yang
- Department of Chemistry, Multiscale Research Institute of Complex Systems and Institute of Biomedical Sciences, Fudan University Shanghai 200438 China
| | - Wolun Zhang
- LightEdge Technologies Limited Zhongshan 528403 China
| | - Shaohui Huang
- Institute of Biophysics, Chinese Academy of Sciences Beijing 100101 China
- University of Chinese Academy of Science Beijing 101408 China
| | - Jingwei Weng
- Department of Chemistry, Multiscale Research Institute of Complex Systems and Institute of Biomedical Sciences, Fudan University Shanghai 200438 China
| | - Zhijun Liu
- National Facility for Protein Science in Shanghai, Zhangjiang Lab, Shanghai Advanced Research Institute, Chinese Academy of Sciences Shanghai 201210 China
| | - Wenning Wang
- Department of Chemistry, Multiscale Research Institute of Complex Systems and Institute of Biomedical Sciences, Fudan University Shanghai 200438 China
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Translocation, Rejection and Trapping of Polyampholytes. Polymers (Basel) 2022; 14:polym14040797. [PMID: 35215709 PMCID: PMC8877523 DOI: 10.3390/polym14040797] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/21/2022] [Revised: 02/11/2022] [Accepted: 02/13/2022] [Indexed: 12/04/2022] Open
Abstract
Polyampholytes (PA) are a special class of polymers comprising both positive and negative monomers along their sequence. Most proteins have positive and negative residues and are PAs. Proteins have a well-defined sequence while synthetic PAs have a random charge sequence. We investigated the translocation behavior of random polyampholyte chains through a pore under the action of an electric field by means of Monte Carlo simulations. The simulations incorporated a realistic translocation potential profile along an extended asymmetric pore and translocation was studied for both directions of engagement. The study was conducted from the perspective of statistics for disordered systems. The translocation behavior (translocation vs. rejection) was recorded for all 220 sequences comprised of N = 20 charged monomers. The results were compared with those for 107 random sequences of N = 40 to better demonstrate asymptotic laws. At early times, rejection was mainly controlled by the charge sequence of the head part, but late translocation/rejection was governed by the escape from a trapped state over an antagonistic barrier built up along the sequence. The probability distribution of translocation times from all successful attempts revealed a power-law tail. At finite times, there was a population of trapped sequences that relaxed very slowly (logarithmically) with time. If a subensemble of sequences with prescribed net charge was considered the power-law decay was steeper for a more favorable net charge. Our findings were rationalized by theoretical arguments developed for long chains. We also provided operational criteria for the translocation behavior of a sequence, explaining the selection by the translocation process. From the perspective of protein translocation, our findings can help rationalize the behavior of intrinsically disordered proteins (IDPs), which can be modeled as polyampholytes. Most IDP sequences have a strong net charge favoring translocation. Even for sequences with those large net charges, the translocation times remained very dispersed and the translocation was highly sequence-selective.
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44
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Reid KM, Singh AK, Bikash CR, Wei J, Tal-Gan Y, Vinh NQ, Leitner DM. The origin and impact of bound water around intrinsically disordered proteins. Biophys J 2022; 121:540-551. [PMID: 35074392 PMCID: PMC8874019 DOI: 10.1016/j.bpj.2022.01.011] [Citation(s) in RCA: 19] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2021] [Revised: 12/14/2021] [Accepted: 01/13/2022] [Indexed: 12/29/2022] Open
Abstract
Proteins and water couple dynamically over a wide range of time scales. Motivated by their central role in protein function, protein-water dynamics and thermodynamics have been extensively studied for structured proteins, where correspondence to structural features has been made. However, properties controlling intrinsically disordered protein (IDP)-water dynamics are not yet known. We report results of megahertz-to-terahertz dielectric spectroscopy and molecular dynamics simulations of a group of IDPs with varying charge content along with structured proteins of similar size. Hydration water around IDPs is found to exhibit more heterogeneous rotational and translational dynamics compared with water around structured proteins of similar size, yielding on average more restricted dynamics around individual residues of IDPs, charged or neutral, compared with structured proteins. The on-average slower water dynamics is found to arise from excess tightly bound water in the first hydration layer, which is related to greater exposure to charged groups. The more tightly bound water to IDPs correlates with the smaller hydration shell found experimentally, and affects entropy associated with protein-water interactions, the contribution of which we estimate based on the dielectric measurements and simulations. Water-IDP dynamic coupling at terahertz frequencies is characterized by the dielectric measurements and simulations.
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Affiliation(s)
- Korey M. Reid
- Department of Chemistry, University of Nevada, Reno, Nevada
| | - Abhishek K. Singh
- Department of Physics and Center for Soft Matter and Biological Physics, Virginia Tech, Blacksburg, Virginia
| | | | - Jessica Wei
- Department of Chemistry, University of Nevada, Reno, Nevada
| | - Yftah Tal-Gan
- Department of Chemistry, University of Nevada, Reno, Nevada
| | - Nguyen Q. Vinh
- Department of Physics and Center for Soft Matter and Biological Physics, Virginia Tech, Blacksburg, Virginia,Corresponding author
| | - David M. Leitner
- Department of Chemistry, University of Nevada, Reno, Nevada,Corresponding author
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45
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Naung MT, Martin E, Munro J, Mehra S, Guy AJ, Laman M, Harrison GLA, Tavul L, Hetzel M, Kwiatkowski D, Mueller I, Bahlo M, Barry AE. Global diversity and balancing selection of 23 leading Plasmodium falciparum candidate vaccine antigens. PLoS Comput Biol 2022; 18:e1009801. [PMID: 35108259 PMCID: PMC8843232 DOI: 10.1371/journal.pcbi.1009801] [Citation(s) in RCA: 24] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/04/2021] [Revised: 02/14/2022] [Accepted: 01/03/2022] [Indexed: 12/30/2022] Open
Abstract
Investigation of the diversity of malaria parasite antigens can help prioritize and validate them as vaccine candidates and identify the most common variants for inclusion in vaccine formulations. Studies of vaccine candidates of the most virulent human malaria parasite, Plasmodium falciparum, have focused on a handful of well-known antigens, while several others have never been studied. Here we examine the global diversity and population structure of leading vaccine candidate antigens of P. falciparum using the MalariaGEN Pf3K (version 5.1) resource, comprising more than 2600 genomes from 15 malaria endemic countries. A stringent variant calling pipeline was used to extract high quality antigen gene 'haplotypes' from the global dataset and a new R-package named VaxPack was used to streamline population genetic analyses. In addition, a newly developed algorithm that enables spatial averaging of selection pressure on 3D protein structures was applied to the dataset. We analysed the genes encoding 23 leading and novel candidate malaria vaccine antigens including csp, trap, eba175, ama1, rh5, and CelTOS. Our analysis shows that current malaria vaccine formulations are based on rare haplotypes and thus may have limited efficacy against natural parasite populations. High levels of diversity with evidence of balancing selection was detected for most of the erythrocytic and pre-erythrocytic antigens. Measures of natural selection were then mapped to 3D protein structures to predict targets of functional antibodies. For some antigens, geographical variation in the intensity and distribution of these signals on the 3D structure suggests adaptation to different human host or mosquito vector populations. This study provides an essential framework for the diversity of P. falciparum antigens to be considered in the design of the next generation of malaria vaccines.
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Affiliation(s)
- Myo T. Naung
- Population Health and Immunity Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia
- Department of Medical Biology, University of Melbourne, Carlton, Victoria, Australia
- Institute of Mental and Physical Health and Clinical Translation (IMPACT), School of Medicine, Deakin University, Geelong, Victoria, Australia
| | - Elijah Martin
- Population Health and Immunity Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia
- Life Sciences Discipline, Burnet Institute, Melbourne, Victoria, Australia
| | - Jacob Munro
- Population Health and Immunity Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia
| | - Somya Mehra
- Population Health and Immunity Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia
- Life Sciences Discipline, Burnet Institute, Melbourne, Victoria, Australia
| | - Andrew J. Guy
- School of Science, RMIT University, Melbourne, Victoria, Australia
| | - Moses Laman
- Vector Borne Diseases Unit, Papua New Guinea Institute of Medical Research, Madang, Papua New Guinea
| | - G. L. Abby Harrison
- Population Health and Immunity Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia
- Department of Medical Biology, University of Melbourne, Carlton, Victoria, Australia
| | - Livingstone Tavul
- Vector Borne Diseases Unit, Papua New Guinea Institute of Medical Research, Madang, Papua New Guinea
| | - Manuel Hetzel
- Swiss Tropical Public Health Institute, Basel, Switzerland
- University of Basel, Basel, Switzerland
| | - Dominic Kwiatkowski
- Sanger Institute, Hinxton, United Kingdom
- Big Data Institute, University of Oxford, Oxford, United Kingdom
| | - Ivo Mueller
- Population Health and Immunity Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia
- Department of Medical Biology, University of Melbourne, Carlton, Victoria, Australia
- Division of Parasites and Insect Vectors, Pasteur Institute, Paris, France
| | - Melanie Bahlo
- Population Health and Immunity Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia
- Department of Medical Biology, University of Melbourne, Carlton, Victoria, Australia
| | - Alyssa E. Barry
- Population Health and Immunity Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia
- Department of Medical Biology, University of Melbourne, Carlton, Victoria, Australia
- Institute of Mental and Physical Health and Clinical Translation (IMPACT), School of Medicine, Deakin University, Geelong, Victoria, Australia
- Life Sciences Discipline, Burnet Institute, Melbourne, Victoria, Australia
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46
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Ledoux J, Trouvé A, Tchertanov L. The Inherent Coupling of Intrinsically Disordered Regions in the Multidomain Receptor Tyrosine Kinase KIT. Int J Mol Sci 2022; 23:ijms23031589. [PMID: 35163518 PMCID: PMC8835827 DOI: 10.3390/ijms23031589] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/05/2022] [Revised: 01/24/2022] [Accepted: 01/26/2022] [Indexed: 02/04/2023] Open
Abstract
RTK KIT regulates a variety of crucial cellular processes via its cytoplasmic domain (CD), which is composed of the tyrosine kinase domain, crowned by the highly flexible domains—the juxtamembrane region, kinase insertion domain, and C-tail, which are key recruitment regions for downstream signalling proteins. To prepare a structural basis for the characterization of the interactions of KIT with its signalling proteins (KIT INTERACTOME), we generated the 3D model of the full-length CD attached to the transmembrane helix. This generic model of KIT in inactive state was studied by molecular dynamics simulation under conditions mimicking the natural environment of KIT. With the accurate atomistic description of the multidomain KIT dynamics, we explained its intrinsic (intra-domain) and extrinsic (inter-domain) disorder and represented the conformational assemble of KIT through free energy landscapes. Strongly coupled movements within each domain and between distant domains of KIT prove the functional interdependence of these regions, described as allosteric regulation, a phenomenon widely observed in many proteins. We suggested that KIT, in its inactive state, encodes all properties of the active protein and its post-transduction events.
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47
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Electrostatically induced pKa shifts in oligopeptides: the upshot of neighboring side chains. Amino Acids 2022; 54:277-287. [DOI: 10.1007/s00726-021-03116-2] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2021] [Accepted: 11/29/2021] [Indexed: 11/01/2022]
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48
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Schultz CJ, Wu Y, Baumann U. A targeted bioinformatics approach identifies highly variable cell surface proteins that are unique to Glomeromycotina. MYCORRHIZA 2022; 32:45-66. [PMID: 35031894 PMCID: PMC8786786 DOI: 10.1007/s00572-021-01066-x] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 09/07/2021] [Accepted: 12/24/2021] [Indexed: 06/14/2023]
Abstract
Diversity in arbuscular mycorrhizal fungi (AMF) contributes to biodiversity and resilience in natural environments and healthy agricultural systems. Functional complementarity exists among species of AMF in symbiosis with their plant hosts, but the molecular basis of this is not known. We hypothesise this is in part due to the difficulties that current sequence assembly methodologies have assembling sequences for intrinsically disordered proteins (IDPs) due to their low sequence complexity. IDPs are potential candidates for functional complementarity because they often exist as extended (non-globular) proteins providing additional amino acids for molecular interactions. Rhizophagus irregularis arabinogalactan-protein-like proteins (AGLs) are small secreted IDPs with no known orthologues in AMF or other fungi. We developed a targeted bioinformatics approach to identify highly variable AGLs/IDPs in RNA-sequence datasets. The approach includes a modified multiple k-mer assembly approach (Oases) to identify candidate sequences, followed by targeted sequence capture and assembly (mirabait-mira). All AMF species analysed, including the ancestral family Paraglomeraceae, have small families of proteins rich in disorder promoting amino acids such as proline and glycine, or glycine and asparagine. Glycine- and asparagine-rich proteins also were found in Geosiphon pyriformis (an obligate symbiont of a cyanobacterium), from the same subphylum (Glomeromycotina) as AMF. The sequence diversity of AGLs likely translates to functional diversity, based on predicted physical properties of tandem repeats (elastic, amyloid, or interchangeable) and their broad pI ranges. We envisage that AGLs/IDPs could contribute to functional complementarity in AMF through processes such as self-recognition, retention of nutrients, soil stability, and water movement.
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Affiliation(s)
- Carolyn J Schultz
- School of Agriculture, Food, and Wine, Waite Research Institute, University of Adelaide, Adelaide, SA, Australia.
| | - Yue Wu
- School of Agriculture, Food, and Wine, Waite Research Institute, University of Adelaide, Adelaide, SA, Australia
| | - Ute Baumann
- School of Agriculture, Food, and Wine, Waite Research Institute, University of Adelaide, Adelaide, SA, Australia
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49
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Abstract
Recombinant protein expression in E. coli often induces the expressed protein to accumulate in insoluble aggregates, named inclusion bodies (IBs), that represent easy to isolate, highly pure protein reservoirs. IBs can be solubilized by denaturing agents but this procedure requires, for complex globular proteins, a refolding step that can be challenging. However, the lack of cooperatively folded tertiary structure in intrinsically disordered proteins (IDP) makes them ideal candidates for this purification strategy. Given the wide abundance of IDPs, their relevance in many disease areas and the numerous IDP-associated biological functions, the interest in this class of proteins has increased substantially over the last decade. Here we present a broad and versatile method for the production and isolation of IDPs from inclusion bodies under denaturant conditions that overcomes the challenges associated with the propensity of these sequences to precipitate from solution and becoming proteolytically degraded.
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Gomes G, do Amaral MJ, Bagri KM, Vasconcellos LM, Almeida MDS, Alvares LE, Mermelstein C. New Findings on LMO7 Transcripts, Proteins and Regulatory Regions in Human and Vertebrate Model Organisms and the Intracellular Distribution in Skeletal Muscle Cells. Int J Mol Sci 2021; 22:ijms222312885. [PMID: 34884689 PMCID: PMC8657913 DOI: 10.3390/ijms222312885] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2021] [Revised: 11/25/2021] [Accepted: 11/26/2021] [Indexed: 12/04/2022] Open
Abstract
LMO7 is a multifunctional PDZ–LIM protein that can interact with different molecular partners and is found in several intracellular locations. The aim of this work was to shed light on LMO7 evolution, alternative transcripts, protein structure and gene regulation through multiple in silico analyses. We also explored the intracellular distribution of the LMO7 protein in chicken and zebrafish embryonic skeletal muscle cells by means of confocal fluorescence microscopy. Our results revealed a single LMO7 gene in mammals, sauropsids, Xenopus and in the holostean fish spotted gar while two lmo7 genes (lmo7a and lmo7b) were identified in teleost fishes. In addition, several different transcripts were predicted for LMO7 in human and in major vertebrate model organisms (mouse, chicken, Xenopus and zebrafish). Bioinformatics tools revealed several structural features of the LMO7 protein including intrinsically disordered regions. We found the LMO7 protein in multiple intracellular compartments in chicken and zebrafish skeletal muscle cells, such as membrane adhesion sites and the perinuclear region. Curiously, the LMO7 protein was detected within the nuclei of muscle cells in chicken but not in zebrafish. Our data showed that a conserved regulatory element may be related to muscle-specific LMO7 expression. Our findings uncover new and important information about LMO7 and open new challenges to understanding how the diverse regulation, structure and distribution of this protein are integrated into highly complex vertebrate cellular milieux, such as skeletal muscle cells.
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Affiliation(s)
- Geyse Gomes
- Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-901, Brazil; (G.G.); (K.M.B.); (L.M.V.)
| | | | - Kayo Moreira Bagri
- Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-901, Brazil; (G.G.); (K.M.B.); (L.M.V.)
| | - Larissa Melo Vasconcellos
- Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-901, Brazil; (G.G.); (K.M.B.); (L.M.V.)
| | - Marcius da Silva Almeida
- Instituto de Bioquímica Médica Leopoldo de Meis, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-901, Brazil;
| | - Lúcia Elvira Alvares
- Departamento de Bioquímica e Biologia Tecidual, Universidade de Campinas (UNICAMP), Campinas, São Paulo 13083-872, Brazil;
| | - Claudia Mermelstein
- Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-901, Brazil; (G.G.); (K.M.B.); (L.M.V.)
- Correspondence:
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