Advances in Protein Molecular and Structural Biology Methods

Advances in Protein Molecular and Structural Biology Methods

1st Edition - January 14, 2022

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  • Editors: Timir Tripathi, Vikash Dubey
  • Paperback ISBN: 9780323902649
  • eBook ISBN: 9780323902656

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Description

Advances in Protein Molecular and Structural Biology Methods offers a complete overview of the latest tools and methods applicable to the study of proteins at the molecular and structural level. The book begins with sections exploring tools to optimize recombinant protein expression and biophysical techniques such as fluorescence spectroscopy, NMR, mass spectrometry, cryo-electron microscopy, and X-ray crystallography. It then moves towards computational approaches, considering structural bioinformatics, molecular dynamics simulations, and deep machine learning technologies. The book also covers methods applied to intrinsically disordered proteins (IDPs)followed by chapters on protein interaction networks, protein function, and protein design and engineering. It provides researchers with an extensive toolkit of methods and techniques to draw from when conducting their own experimental work, taking them from foundational concepts to practical application.

Key Features

  • Presents a thorough overview of the latest and emerging methods and technologies for protein study
  • Explores biophysical techniques, including nuclear magnetic resonance, X-ray crystallography, and cryo-electron microscopy
  • Includes computational and machine learning methods
  • Features a section dedicated to tools and techniques specific to studying intrinsically disordered proteins

Readership

Molecular biologists, biochemists, structural biologists, biophysicists, computational biologists, and researchers working with proteins at the molecular level and associated methods. Graduate, postgraduate and PhD students in the field of molecular biology, structural biology and related fields

Table of Contents

  • Cover image
  • Title page
  • Table of Contents
  • Copyright
  • Dedicated to
  • Contributors
  • About the Editors
  • Foreword
  • Preface
  • Chapter 1: Strategies to improve the expression and solubility of recombinant proteins in E. coli
  • Abstract
  • 1: Introduction
  • 2: Before starting with protein expression
  • 3: Materials required
  • 4: Standard protocol for recombinant protein expression in E. coli
  • 5: Troubleshooting strategies
  • 6: Conclusion and future perspectives
  • Chapter 2: Advances in heterologous protein expression strategies in yeast and insect systems
  • Abstract
  • Acknowledgment
  • 1: Introduction
  • 2: Heterologous protein expression strategies in yeast systems
  • 3: Heterologous protein expression strategies in insect systems
  • 4: Conclusion of baculovirus expression systems
  • Chapter 3: Methods for transient expression and purification of monoclonal antibodies in mammalian cells
  • Abstract
  • 1: Introduction
  • 2: Background experimental preparation
  • 3: Materials required for antibody purification
  • 4: Detailed step-by-step protocol for antibody purification
  • 5: Troubleshooting problems
  • 6: Conclusions
  • Chapter 4: Methods for recombinant production and purification of intrinsically disordered proteins
  • Abstract
  • 1: Before you begin
  • 2: Materials and equipment
  • 3: Step-by-step method details
  • 4: Expected outcomes
  • 5: Optimization and troubleshooting
  • Chapter 5: Methods to determine the oligomeric structure of proteins
  • Abstract
  • 1: Introduction
  • 2: Electrophoretic methods
  • 3: Size exclusion chromatography
  • 4: Dynamic light scattering
  • 5: Circular dichroism spectroscopy
  • 6: Fluorescence-based methods
  • 7: Analytical ultracentrifugation
  • 8: X-ray crystallography and NMR spectroscopy
  • 9: Mass spectroscopy
  • 10: Atomic force microscopy
  • 11: Co-immunoprecipitation
  • 12: Computational methods
  • 13: Conclusions
  • Chapter 6: Multimodal methods to study protein aggregation and fibrillation
  • Abstract
  • Acknowledgments
  • 1: Introduction
  • 2: Combination of in vitro techniques to evaluate isolated protein aggregates and fibrils
  • 3: Multimodal methods to evaluate the aggregation of proteins in cells tissues and living system
  • Chapter 7: Experimental methods to study the thermodynamics of protein–protein interactions
  • Abstract
  • 1: Introduction
  • 2: Criteria for forming a protein–protein interaction
  • 3: Characteristic features of PPI interfaces
  • 4: Thermodynamic parameters associated with PPI
  • 5: Techniques to study the thermodynamics of PPI
  • 6: Conclusions
  • Chapter 8: Experimental methods to study the kinetics of protein–protein interactions
  • Abstract
  • Acknowledgment
  • 1: Introduction
  • 2: Surface plasmon resonance
  • 3: Bio-layer interferometry
  • 4: Microscale thermophoresis
  • 5: Isothermal titration calorimetry
  • 6: Quartz crystal microbalance
  • 7: Conclusions
  • Chapter 9: Computational techniques for studying protein-protein interactions
  • Abstract
  • 1: Introduction
  • 2: Types of protein-protein complexes
  • 3: PPIs as targets for drug discovery
  • 4: Mining PPIs
  • 5: Computational techniques in PPI detection
  • 6: Comparison of available computational approaches
  • 7: PPI databases
  • 8: PPI network and visualization
  • 9: Conclusion and future perspectives
  • Chapter 10: Experimental methods to study protein–nucleic acid interactions
  • Abstract
  • 1: Introduction
  • 2: Single-molecule approaches for the identification and validation of protein–nucleic acid interactions
  • 3: Investigation of protein–nucleic acid interactions in mammalian cell lines
  • 4: Conclusions
  • Chapter 11: Advanced computational tools for quantitative analysis of protein–nucleic acid interfaces
  • Abstract
  • Acknowledgment
  • 1: Protein–RNA complexes
  • 2: Datasets for studying protein–RNA interfaces
  • 3: Tools for the analysis of protein–RNA interfaces
  • 4: Tools for protein–RNA binding site prediction
  • 5: Tools for protein–RNA binding affinity prediction
  • 6: Tools for hot spots at protein–RNA interfaces
  • 7: A brief survey of tools for studying protein–DNA interactions
  • 8: Conclusions
  • Chapter 12: Experimental techniques to study protein dynamics and conformations
  • Abstract
  • 1: Introduction
  • 2: Various methods to study protein dynamics and conformations
  • 3: Conclusion and future perspectives
  • Chapter 13: Computational techniques to study protein dynamics and conformations
  • Abstract
  • Acknowledgments
  • 1: Introduction
  • 2: “Realistic” methods: Molecular dynamics and enhanced sampling
  • 3: “Simplified” approaches: Coarse-graining and path-sampling algorithms
  • 4: A case study: The open-to-close transition of the ribose-binding protein
  • 5: Summary and conclusions
  • Chapter 14: Application of circular dichroism spectroscopy in studying protein folding, stability, and interaction
  • Abstract
  • 1: Introduction
  • 2: Theory of circular dichroism
  • 3: Application of CD spectroscopy
  • 4: Time-resolved CD spectroscopy and its uses in protein folding kinetics
  • 5: Conclusion and future perspectives
  • Conflict of interests
  • Chapter 15: Studying protein-folding dynamics using single-molecule fluorescence methods
  • Abstract
  • 1: Introduction
  • 2: Single-molecule fluorescence techniques for protein-folding dynamics
  • 3: Conclusion and future perspectives
  • Chapter 16: Advances in liquid-state NMR spectroscopy to study the structure, function, and dynamics of biomacromolecules
  • Abstract
  • 1: Introduction to liquid-state NMR spectroscopy
  • 2: Liquid-state NMR spectroscopy of biomacromolecules
  • 3: Biomolecular behavior and drug discovery
  • 4: NMR of biomacromolecules in living cells
  • 5: Summary
  • Chapter 17: In-cell NMR spectroscopy: A tool to study cellular structure biology
  • Abstract
  • 1: Introduction
  • 2: Overview of in-cell NMR
  • 3: Bioreactor systems for in-cell NMR observations
  • 4: Applications of in-cell NMR
  • 5: Conclusion and future perspectives
  • Chapter 18: Current trends in membrane protein crystallography
  • Abstract
  • Acknowledgments
  • 1: Introduction
  • 2: Expression screening of membrane proteins for crystallization
  • 3: Detergent screening and fluorescence size exclusion chromatography of membrane proteins
  • 4: Crystallization of membrane proteins
  • 5: Engineering membrane proteins to facilitate crystal formation
  • 6: X-ray sources
  • 7: Detectors
  • 8: Time-resolved crystallography
  • 9: X-ray free-electron laser
  • 10: Conclusion and future perspectives
  • Chapter 19: Advances in sample preparation and data processing for single-particle cryo-electron microscopy
  • Abstract
  • 1: Introduction
  • 2: Sample quality is the key to high-resolution structure determination
  • 3: Grid preparation for SPA
  • 4: Time-resolved cryoEM
  • 5: Advances in SPA data collection and processing
  • 6: AI/ML-based approaches in cryoEM data processing pipeline
  • 7: Conclusion and future perspectives
  • Chapter 20: Advanced mass spectrometry-based methods for protein molecular-structural biologists
  • Abstract
  • Acknowledgments
  • 1: Introduction
  • 2: Data-independent acquisitions (DIA) for accurate protein quantification
  • 3: Resolving protein structures using DIA-MS
  • 4: Conclusions and outlooks
  • Chapter 21: Developments, advancements, and contributions of mass spectrometry in omics technologies
  • Abstract
  • 1: Introduction
  • 2: Omics mass spectrometry
  • 3: Mass spectrometry in Omics technologies
  • 4: Recent developments in the mass spectrometer
  • 5: Fragmentation principles
  • 6: Conclusion and future perspectives
  • Chapter 22: Role of structural biology methods in drug discovery
  • Abstract
  • 1: Introduction
  • 2: Structural biology aided selection of drug targets
  • 3: Role of experimental and computational approaches in drug discovery
  • 4: Virtual screening
  • 5: Enhancement of ligand specificity
  • 6: Optimization of hits and drug-likeness
  • 7: Development of peptidomimetics
  • 8: Conclusion and future perspectives
  • Chapter 23: Prediction, validation, and analysis of protein structures: A beginner’s guide
  • Abstract
  • 1: Introduction
  • 2: Protein structure modeling
  • 3: Protein structure refinement and validation
  • 4: Protein structure analysis and importance of protein folding
  • 5: Recent advances in in silico protein structure determination
  • 6: Conclusion and future perspectives
  • Chapter 24: Advances in structure-based virtual screening for drug discovery
  • Abstract
  • 1: Introduction
  • 2: Drug design and the computers
  • 3: Conclusion and future perspectives
  • Chapter 25: Methods and applications of machine learning in structure-based drug discovery
  • Abstract
  • Acknowledgments
  • 1: Introduction
  • 2: Protein crystallography and AI-assisted drug discovery
  • 3: Application of ML in protein structure prediction (in silico approach)
  • 4: Virtual screening
  • 5: Conclusion and future perspectives
  • Chapter 26: Molecular dynamics simulations: Principles, methods, and applications in protein conformational dynamics
  • Abstract
  • Acknowledgments
  • 1: Introduction
  • 2: Applications of MD simulations
  • 3: Materials
  • 4: Methods
  • 5: Notes
  • 6: Utility of MD simulation: A case study on conformational dynamics of d-amino acid oxidase (DAAO)
  • 7: Conclusion and future perspectives
  • Chapter 27: Applications of molecular dynamics simulations in drug discovery
  • Abstract
  • 1: Introduction
  • 2: Identification of protein conformation ensemble and drug binding site
  • 3: Modeling protein-drug interactions
  • 4: Modeling drug-membrane interactions
  • 5: Conclusion and future perspectives
  • Chapter 28: Envisaging the conformational space of proteins by coupling machine learning and molecular dynamics
  • Abstract
  • Declaration of competing interests
  • 1: Introduction
  • 2: Conformational impact due to various environment
  • 3: Multiple conformational states of proteins
  • 4: Impact of Ramachandran plot in conformational space
  • 5: Variability in the conformation of intrinsically disordered proteins
  • 6: Conformational sampling analysis through different methods
  • 7: Role of force fields in different conformational space observation
  • 8: Conformational space assessment on the explicit and implicit solvent model
  • 9: Conformational space analysis through machine learning
  • 10: Combination of MD simulation and machine learning
  • 11: Conclusion and future perspectives
  • Chapter 29: Immunoinformatics and reverse vaccinology methods to design peptide-based vaccines
  • Abstract
  • 1: Introduction
  • 2: Peptide vaccines
  • 3: Methods and tools in reverse vaccinology
  • 4: Steps involved in reverse vaccinology
  • 5: Advantage of peptide vaccine or multi-epitope vaccines
  • 6: Conclusion and future perspectives
  • Chapter 30: Computational methods to study intrinsically disordered proteins
  • Abstract
  • Acknowledgments
  • 1: Introduction
  • 2: Bioinformatics over biophysical techniques to study IDP
  • 3: Common predictors for identification of IDPs
  • 4: Identification of molecular recognition features (MoRFs)
  • 5: Prediction of nucleic acid-binding regions
  • 6: Biological relevance of predictions
  • 7: Conclusion and future perspectives
  • Chapter 31: Experimental methods to study intrinsically disordered proteins
  • Abstract
  • 1: Introduction
  • 2: Size exclusion chromatography
  • 3: UV-vis absorption spectroscopy
  • 4: Circular dichroism spectroscopy
  • 5: Fluorescence spectroscopy
  • 6: Nuclear magnetic resonance spectroscopy
  • 7: Fourier transform infrared spectroscopy
  • 8: Electron spin resonance spectroscopy
  • 9: Raman spectroscopy
  • 10: Light scattering methods
  • 11: Microscopy-based methods
  • 12: Analytical ultracentrifugation
  • 13: Mass spectrometry
  • 14: Conclusions and future perspectives
  • Chapter 32: Analysis of structure and dynamics of intrinsically disordered regions in proteins using solution NMR methods
  • Abstract
  • 1: Introduction
  • 2: NMR chemical shift assignments of intrinsically disordered sequences
  • 3: Structural characterization of IDRPs
  • 4: Characterization of IDRP dynamics
  • 5: In-cell NMR experiments
  • 6: Conclusions and future perspectives
  • Chapter 33: Methods to study the effect of solution variables on the conformational dynamics of intrinsically disordered proteins
  • Abstract
  • 1: Introduction
  • 2: Computational tools to study the impacts of solution variables on IDPs
  • Chapter 34: Molecular simulations to study IDP-IDP interactions and their complexes
  • Abstract
  • 1: Intrinsically disordered proteins and their interactions
  • 2: Introduction of the molecular simulation techniques
  • 3: Characterizing IDP–IDP interactions and their complexes by coarse-grained models
  • 4: Challenges in coarse-grained models
  • 5: Conclusion and future perspectives
  • Chapter 35: Exploring large-scale protein function using systematic mutant analysis
  • Abstract
  • Acknowledgments
  • 1: Introduction
  • 2: Engineering systematic site saturation mutant libraries
  • 3: Screening the systematic mutant libraries for variant function
  • 4: Next-generation sequencing of the variants
  • 5: Large-scale functional mapping in proteins
  • 6: Conclusion and future perspectives
  • Chapter 36: Approaches and methods to study cell signaling: Linguistics of cellular communication
  • Abstract
  • 1: Introduction
  • 2: Molecular players in signal transduction
  • 3: Cell signaling can occur in a variety of ways
  • 4: Cell signaling orchestrates key biological processes
  • 5: Experimental techniques used to study cell signaling
  • 6: Conclusion and future perspectives
  • Chapter 37: Methods to study systems biology of signaling networks: A case study of NSCLC
  • Abstract
  • Acknowledgments
  • 1: Introduction
  • 2: Non-small cell lung carcinoma (NSCLC)
  • 3: Systems biology
  • 4: Applications of systems biology approaches in cancer studies
  • 5: System biology methods and approaches
  • 6: Analysis of the data
  • 7: Interpretations and conclusions
  • Chapter 38: Advancements in the analysis of protein post-translational modifications
  • Abstract
  • 1: Introduction
  • 2: Ubiquitination
  • 3: Types of ubiquitination
  • 4: Detection of protein ubiquitination
  • 5: Identification of the site of ubiquitination
  • 6: Ubiquitin chain architecture detection
  • 7: Conclusion and future perspectives
  • Chapter 39: Protein engineering: Methods and applications
  • Abstract
  • 1: Introduction
  • 2: Protein engineering approaches
  • 3: Directed evolution
  • 4: Semi-rational design
  • 5: De novo design
  • 6: Rational design
  • 7: Applications of protein engineering
  • 8: Conclusions
  • Chapter 40: Designer 3D-DNA nanodevices: Structures, functions, and cellular applications
  • Abstract
  • 1: Introduction
  • 2: Different approaches to realize 3D DNA polyhedral nanodevices
  • 3: Methods for characterization of DNA nanostructures
  • 4: Cellular uptake of TDN and its characterization
  • 5: Conclusions and future perspectives
  • Index

Product details

  • No. of pages: 714
  • Language: English
  • Copyright: © Academic Press 2022
  • Published: January 14, 2022
  • Imprint: Academic Press
  • Paperback ISBN: 9780323902649
  • eBook ISBN: 9780323902656

About the Editors

Timir Tripathi

Timir Tripathi is the Regional Director of Indira Gandhi National Open University (IGNOU), Regional Centre Kohima, Nagaland, India. Earlier, he served as Senior Assistant Professor and Principal Investigator at the Department of Biochemistry, North-Eastern Hill University, Shillong, India. He holds a Ph.D. from the Central Drug Research Institute, Lucknow, India. He was a visiting faculty at ICGEB, New Delhi, India (2011), and Khon Kaen University, Thailand (2015). He is known for his research in the fields of protein biophysics, biochemistry, structural biology, and drug discovery. He has over 16 years of experience in teaching and research on protein structure, function, and dynamics at the post-graduate and doctoral levels. He has developed and improved methods to investigate and analyze proteins. His research areas include protein interaction dynamics and understanding the roles of non-catalytic domains in regulating the catalytic activity of proteins. The common theme in his research is an interest in understanding biological phenomena involving proteins at the molecular, structural, and mechanistic levels. He has handled several research grants as a principal investigator from various national and international funding agencies, including DST-Russian Foundation for Basic Research, UGC-Israel Science Foundation, DBT, SERB, DHR, and ICMR. He has received several awards, including Prof. B.K. Bachhawat Memorial Young Scientist Lecture Award (2020) by the National Academy of Sciences, India, ISCB-Young Scientist Award (2019), ICMR-Shakuntala Amir Chand Prize (2018), BRSI-Malviya Memorial Award (2017), DST Fasttrack Young Scientist Award (2012), DBT Overseas Associateship Award (2012), Dr. D.M. Bose Award (2008), etc. He is an elected member of the National Academy of Sciences, India, and the Royal Society of Biology, UK. He has published more than 100 research papers, reviews, commentaries, viewpoints, and editorial articles in international journals, has edited three books, and published several book chapters. He currently serves on the editorial boards of several international journals, including International Journal of Biological Macromolecules, Acta Tropica, Scientific Reports, PLoS One, etc.

Affiliations and Expertise

Regional Director of Indira Gandhi National Open University (IGNOU), Regional Centre Kohima, Nagaland, India

Vikash Dubey

Vikash Kumar Dubey is a Professor at the School of Biochemical Engineering and Associate Dean for Academic Affairs at the Indian Institute of Technology (BHU), Varanasi, India. Prior to joining IIT (BHU), he was a postdoctoral fellow at Florida State University, USA before joining IIT Guwahati, India, where he served as Associate Professor and Professor. He has published over 130 articles in peer-reviewed journals and over 80 conference presentations/proceedings. He has seven awarded and licensed USA patents and many Indian patents. He has also written several book chapters and has guided a number of PhD students, MTech students, and postdoctoral fellows. He has received several awards and has been elected as a member of a number of scientific societies and academies; Among these, he is currently the Vice President of the Bioinformatics and Drug Discovery Society [BIDDS], India and a Fellow of the Royal Society of Biology, UK.

Affiliations and Expertise

Professor, School of Biochemical Engineering, Indian Institute of Technology (BHU), Varanasi, India

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