Advanced Spectroscopic Methods to Study Biomolecular Structure and Dynamics

Front Cover
Advanced Spectroscopic Methods to Study Biomolecular Structure and Dynamics
Copyright
Dedication
Contents
Contributors
Editors biography
Foreword
Preface
Chapter 1: Fundamentals of spectroscopy for biomolecular structure and dynamics
1. Introduction to spectroscopy
1.1. Electromagnetic spectrum
1.2. Electron energy level and Jablonski diagram
1.3. Basic properties of light
1.3.1. Reflection
1.3.2. Refraction
1.3.3. Diffraction
1.3.4. Scattering
1.3.5. Polarization
1.4. Spectroscopy: Types and parameters
1.5. Biomolecular structure and dynamics
2. General design of a spectrometer instrumentation
3. Various spectroscopic methods
3.1. Fluorescence spectroscopy-based methods
3.2. Circular dichroism spectroscopy
3.3. Nuclear magnetic resonance spectroscopy
3.4. Infrared spectroscopy
3.5. Raman spectroscopy
3.6. Light scattering methods
3.7. Mass spectrometry
3.8. X-ray spectroscopy
4. Conclusions
References
Chapter 2: Fluorescence-based techniques to assess biomolecular structure and dynamics
1. Introduction
1.1. Types of fluorescence measurements
1.2. Fluorophores
2. Environmental effects used in fluorescence structural studies
2.1. Environment-dependent shifts of fluorescence parameters
2.2. Time-resolved emission spectra
2.3. Ultrafast fluorescence upconversion spectroscopy
2.4. Red-edge excitation shifts
2.5. Voltage clamp fluorometry
3. Förster resonance energy transfer
3.1. Transition-metal ion FRET
3.2. Lanthanide-based resonance energy transfer
3.3. Determination of quaternary protein structure by FRET
3.4. Bioluminescence resonance energy transfer
4. Fluorescence quenching
4.1. Photoinduced electron transfer as a molecular ruler
4.2. Depth-dependent quenching in lipid bilayers
4.3. Thermal shift assay
5. Fluorescence anisotropy
5.1. Time-resolved anisotropy measurements
5.2. Determination of homoFRET by anisotropy measurements
6. Diffusion-based estimation of biomolecular size and shape
6.1. Analytical ultracentrifugation
6.2. Microscale thermophoresis
6.3. Fluorescence correlation spectroscopy
7. Biomolecules in situ
7.1. CLSM and its auxiliary techniques
7.2. FLIM complements localization data
7.3. Superresolution and single-molecule studies
8. Conclusions
References
Chapter 3: Structural analysis of biomacromolecules using circular dichroism spectroscopy
1. Introduction
2. Applications of CD in protein studies
2.1. CD in the analysis of protein structure
2.2. CD spectra of proteins
2.3. Calculation of secondary structural by CD spectrogram
2.3.1. Method I-Approximate estimation
2.3.2. Method II-Deconvolution method based on reference databases
2.4. Recent advances in predicting secondary structure using CD
2.5. Changes in CD spectra by environmental factors
2.5.1. pH
2.5.2. Temperature
2.5.3. Chemical modifications
2.6. Changes in CD spectra by protein-related interactions
2.6.1. Protein-protein interaction
2.6.2. Protein-ligand interaction
3. Applications of CD in studying polysaccharides
3.1. Basic knowledge of CD spectrum of polysaccharides
3.2. Polysaccharides in the vacuum UV region
3.3. Polysaccharides in the induced CD region
3.4. Polysaccharides CD spectra affected by environmental parameters
4. Applications of CD in nucleic acid measurements
4.1. Basic understanding of CD spectrum of nucleic acids
4.2. CD spectra of nucleic acids: From simple sequence to native DNA
4.3. Binding of drugs with DNA
5. Conclusions and outlook
References
Chapter 4: Nuclear magnetic resonance spectroscopy for protein structure, folding, and dynamics
1. Introduction
2. NMR spectroscopy approaches to study protein folding and dynamics
2.1. Real-time NMR spectroscopy
2.2. Multidimensional real-time NMR spectroscopy
2.3. Polarization-enhanced fast-pulsing techniques
2.4. 2D real-time NMR spectroscopy
2.5. Chemical shifts
2.6. Spin-lattice (T1) and spin-spin (T2) and nuclear Overhauser effect
2.7. Relaxation dispersion (RD) techniques
2.8. Rotating frame relaxation
2.9. Paramagnetic relaxation enhancement (PRE)
2.10. ZZ-exchange
2.11. Chemical exchange saturation transfer (CEST)
2.12. Hydrogen-deuterium exchange (H-D or H/D exchange)
3. Conclusion and future prospects
References
Chapter 5: Advanced NMR spectroscopy methods to study protein structure and dynamics
1. Introduction
2. Traditional NMR spectroscopy approaches for small-medium-sized proteins
3. Protein backbone dynamics
3.1. The heteronuclear NOE
3.2. T1 relaxation
3.3. T2 relaxation
3.4. Model-free analysis
4. NMR spectroscopy for large proteins
4.1. Deuteration
4.2. Selective methyl labeling
4.3. TROSY
5. Methods for probing protein dynamics of large proteins
5.1. Fast (ps-ns) time scale motions
5.1.1. Deuterium relaxation
5.1.2. ``Forbidden´´ multiple quantum transitions
5.2. Slow (μs-ms) time scale motions
5.2.1. Zero quantum (ZQ) and double quantum (DQ) relaxation
5.2.2. Chemical exchange saturation transfer (CEST) and dark state excitation saturation transfer (DEST)
5.2.3. ZZ-exchange spectroscopy
5.2.4. CPMG relaxation dispersion experiments
6. Methods for simultaneous study of the structure and dynamics of proteins
6.1. Paramagnetic relaxation enhancement
6.2. Residual dipolar couplings
7. Conclusions
Acknowledgments
References
Chapter 6: Applications of infrared spectroscopy to study proteins
1. Introduction
2. Infrared spectrum
3. Infrared spectrum for the structural characterization of proteins
3.1. Alteration in the chemical structure of proteins
3.2. Understanding the redox state and bonding in proteins
3.3. Conformational aspects and hydrogen bonding
3.4. Conformational freedom and electric fields
4. Infrared spectrophotometers
4.1. Fourier-transform IR (FT-IR) spectrophotometer
4.2. Dispersive IR spectrophotometers
5. Types of IR measurements
5.1. Transmission measurements
5.2. Attenuated total reflectance (ATR) measurements
6. IR absorption and detection of amino acid side chains
7. IR absorption and detection of the protein backbone
7.1. Vibrations of NH stretching (3300cm-1 for amide A and 3070cm-1 for amide B)
7.2. Vibrations of amide I (1650cm-1)
7.3. Vibrations of amide II (1550cm-1)
7.4. Vibrations of amide III (1400-1200cm-1)
8. IR spectroscopy for studying proteins
8.1. Understanding the protein secondary structure
8.2. Flexibility of proteins
8.3. Function of proteins
8.4. Measuring enzyme activity
8.5. Water and hydrated proton in proteins
9. Studying proteins with IR spectroscopy: Case studies
10. Conclusion and future perspectives
References
Chapter 7: Raman spectroscopy to study biomolecules, their structure, and dynamics
1. Introduction
2. Applications of Raman spectroscopy
2.1. Applications in microbiology
2.1.1. In studying viruses
2.1.2. In studying excretion
2.1.3. In studying bacteria
2.1.4. In studying fungi
2.2. Applications in plants
2.2.1. In studying pollens
2.2.2. In studying leaves, fruits, and seeds
2.3. Applications in animal science
2.3.1. In studying bones
2.3.2. In studying animal cells and tissues
3. Conclusions
Acknowledgments
References
Chapter 8: Spectroscopic investigation of biomolecular dynamics using light scattering methods
1. Introduction
2. Basics of light scattering
3. Applications of light scattering methods
3.1. Protein folding and unfolding
3.2. Conformational fluctuations, disorder, and transitions
4. Conclusion and future perspectives
References
Chapter 9: Protein footprinting by mass spectrometry: H/D exchange, specific amino acid labeling, and fast photochemical ...
1. Introduction
2. Hydrogen-deuterium exchange mass spectrometry
2.1. Introduction to HDX-MS
2.2. HDX-MS workflow and mechanism
2.3. Recent applications
2.3.1. Epitope mapping, DBP, and inhibitory antibodies
2.3.2. Small-molecule binding study, ApoE3 and EZ482
2.3.3. Metal-binding study: Troponin C and Ca2+ binding
2.3.4. Metal-binding study: Calprotectin and Ca2+ binding
3. Specific amino acid labeling
3.1. Introduction
3.2. Recent applications
3.2.1. FMO orientation between the membrane and chlorosome, elucidated by GEE footprinting
3.2.2. Siderocalin: Footprint Arg and Lys
3.2.3. Protocol for the development of amino acid specific footprinting
4. FPOP for protein structural studies
4.1. Introduction
4.2. Recent applications
4.2.1. Epitope mapping using FPOP
4.2.2. FPOP for studying the protein aggregation
4.2.3. Submillisecond folding probed by FPOP
5. Conclusions
Acknowledgments
References
Chapter 10: Small-angle scattering techniques for biomolecular structure and dynamics
1. Introduction to small-angle scattering experiments
1.1. Instrumental layouts
1.1.1. Small-angle X-ray scattering
1.1.2. Small-angle neutron scattering
1.1.3. Small-angle light scattering
1.2. Beam sources and their interaction with the sample
1.2.1. SAXS
1.2.2. SANS
1.2.3. SALS
1.3. Large facilities and bench-top instruments
2. Structural studies
2.1. Single-protein analysis
2.2. Aggregates and protein material
3. Dynamics analysis
3.1. Sample environment
3.2. Changes in temperature
3.3. Changes in pressure
3.4. Shear and rheology
3.5. Magnetic fields
3.6. Electric fields
4. Models
4.1. General models for the SAS pattern
4.1.1. Guinier region
4.1.2. Fractal regime
4.1.3. Porod region
4.1.4. Unified model
4.2. Biomolecular models for the SAS pattern
4.2.1. Protein solutions
4.2.2. Membranes
4.3. Deep learning methods for the reconstruction of 3D models
5. Conclusions
Acknowledgments
References
Chapter 11: Advances in X-ray crystallography methods to study structural dynamics of macromolecules
1. Introduction
2. Protein extraction and purification
2.1. Detergents and surfactants
2.1.1. Maltose-neopentyl glycol (MNG) compounds
2.1.2. Glucose-neopentyl glycerol (GNG) compounds
2.1.3. Nonionic amphipols (NAPoI)
2.1.4. Calixarene
2.1.5. Fluorinated surfactants
2.1.6. Commercial detergent screen kits
2.2. Membrane mimetics
2.2.1. Nanodiscs
2.2.2. Styrene maleic acid copolymer lipid particles (SMALPs)
2.2.3. Saposin-lipoprotein nanoparticle system (Salipro)
3. Increasing the solubility and stability of proteins
3.1. Crystallization chaperones
3.2. Thermostabilizing mutations
4. Assessing the homogeneity and purity of protein samples
4.1. UV-vis and fluorescence spectroscopy
4.2. Size exclusion chromatography
4.3. Dynamic light scattering
4.4. Size exclusion chromatography with multiangle light scattering (SEC-MALS)
5. New crystallization methods
5.1. Automation of crystallization
5.2. On-chip crystal growth
6. New crystallization additives
6.1. Porous nucleants
6.2. Molecularly imprinted polymers (MIPs)
6.3. Crystallophore (Tb-Xo4)
6.4. Other nucleants
7. Advances in instrument and data-processing software
7.1. Automations in screening crystallization conditions
7.2. Detecting protein crystals using an automated plate imager
7.3. Advances in synchrotron radiation instrumentation
7.4. In situ X-ray screening and data collection
7.5. In situ data collection using X-ray free-electron laser (XFEL)
7.6. The computational tools available in protein crystallography
8. Conclusions and future perspectives
References
Chapter 12: Spectroscopic methods to study protein folding kinetics: Methodology, data analysis, and interpretation of the&
1. Introduction
2. Kinetics of protein folding
2.1. Principle
2.2. Methodology
2.3. Data analysis and interpretation
2.3.1. Obtaining the experimental rate constant
2.3.2. Transition state
2.3.3. Modeling of the folding/unfolding pathway
3. Applications in protein engineering
4. Conclusions
References
Chapter 13: Spectroscopic methods to study the thermodynamics of biomolecular interactions
1. Introduction
2. Overview of biomolecular forces
2.1. Hydrogen bonding
2.2. Hydrophobic interactions
2.3. Van der Waals interactions
2.4. Electrostatic interactions
2.5. Configurational entropy
2.6. Bonded interactions
3. Thermodynamics overview
3.1. Thermodynamics of protein folding, binding reactions, and interactions
3.2. Enthalpy contributions
3.3. Cooperativity
4. Methods for binding constant and thermodynamics study
4.1. Differential scanning calorimetry and isothermal titration calorimetry
4.2. Spectroscopic techniques for thermodynamics studies
4.2.1. Ultraviolet-visible (UV-vis) spectroscopy
4.2.2. Fluorescence spectroscopy
4.2.3. Circular dichroism (CD) spectroscopy
4.2.4. Nuclear magnetic resonance (NMR) spectroscopy
4.2.5. Atomic force spectroscopy
4.2.6. Mass spectrometry
5. Conclusions
Acknowledgments
References
Chapter 14: Spectroscopic methods to detect and analyze protein oligomerization, aggregation, and fibrillation
1. Introduction
2. UV-visible spectroscopy
2.1. Principle
2.2. Indirect detection of protein conformation changes
2.3. Direct detection of protein conformational changes
2.4. Estimating the concentration of proteins
2.5. Advantages and limitations
3. Circular dichroism spectroscopy
3.1. Principle
3.2. Far-UV CD
3.3. Near-UV CD
3.4. Advantages and limitations
4. Fluorescence spectroscopy
4.1. Principle
4.2. Intrinsic and extrinsic fluorophores
4.3. Sample properties
4.4. Steady-state fluorescence
4.5. Fluorescence anisotropy
4.6. Time-resolved fluorescence spectroscopy
4.7. Fluorescence correlation spectroscopy
4.8. Advantages and limitations
5. Infrared spectroscopy
5.1. Principle
5.2. FTIR spectroscopy to analyze amyloid aggregates
5.3. FTIR spectroscopy to analyze nonamyloid aggregates
5.4. Combinatorial studies with FTIR
5.5. Advantages and limitations
6. Dynamic light scattering spectroscopy
6.1. Principle
6.2. Protein aggregation analysis by DLS
6.3. Advantages and limitations
7. Raman spectroscopy
7.1. Principle
7.2. Protein analysis by deep UV Raman spectroscopy (DUVRR)
7.3. Protein analysis by surface-enhanced Raman spectroscopy (SERS)
7.4. Advantages and limitations
8. NMR spectroscopy
8.1. Principle
8.2. Solution-state NMR
8.3. Solid-state NMR
8.4. Advantages and disadvantages
9. Conclusion
Acknowledgments
References
Chapter 15: Multimodal spectroscopic methods for the analysis of carbohydrates
1. Introduction
2. Sample preparation for the spectroscopic analysis of carbohydrates
2.1. Extraction and purification
2.1.1. Pressurized liquid extraction
2.1.2. Field flow fractionation
2.1.3. Chromatographic procedures
2.1.4. Other important fractionation techniques
2.2. Purity estimation
2.3. Chemical modifications for analysis
3. Advanced analysis of carbohydrates
3.1. High-performance liquid chromatography
3.1.1. HPLC detectors
3.2. Mass spectrometry
3.3. Infrared spectroscopy
3.4. Raman spectroscopy
3.5. Nuclear magnetic resonance spectroscopy
3.6. X-ray diffraction analysis
3.7. Multidimensional techniques for carbohydrate analysis
4. Conclusions
References
Chapter 16: Integration of spectroscopic and computational data to analyze protein structure, function, folding, and d
1. Protein structures: A race with time
2. Spectroscopic tools to study protein structure and dynamics
2.1. CD spectroscopy
2.2. Fluorescence spectroscopy
2.3. NMR spectroscopy
2.4. FTIR spectroscopy
2.5. Raman spectroscopy
2.6. X-ray crystallography
2.7. Small-angle X-ray scattering
3. Computational tools to study protein structure and dynamics
3.1. Modeling the protein structure
3.1.1. Template-based modeling
3.1.2. Template-free modeling
3.2. Molecular dynamics simulation of protein structure
3.3. Secondary structure prediction using the DSSP tool
4. Integrating spectroscopic data with computational data
5. Case studies
5.1. Integrating data from CD spectroscopy and computational analysis
5.2. Urea denaturation analysis: Integrating fluorescence and MD simulation data
5.3. Integrating CD, fluorescence, and MD simulation data to understand the roles of specific mutations in protein struct ...
6. Conclusion and future perspectives
References
Chapter 17: Advance data handling tools for easy, fast, and accurate interpretation of spectroscopic data
1. Introduction
2. Spectroscopic data handling tools
2.1. Origin
2.1.1. To install origin
2.1.2. To use origin
2.2. SigmaPlot
2.2.1. To install SigmaPlot
2.2.2. To use SigmaPlot
2.3. JCAMP-DX
2.3.1. To install JCAMP-DX
2.3.2. To use JCAMP-DX
2.3.3. JDXview
To install JDXview
2.4. jsNMR
2.4.1. Ways to load the spectrum
2.4.2. Zooming and panning
2.4.3. Analysis and processing
2.4.4. Spectral management
2.4.5. File types that are commonly used
2.5. Unscrambler
2.5.1. To install Unscrambler
2.5.2. To run Unscrambler
2.5.3. To use Unscrambler
2.6. MBROLE 2.0
2.6.1. To use MBROLE 2.0
2.7. MASCOT
2.7.1. To use MASCOT
2.8. MaxQuant
2.8.1. To use MaxQuant
2.9. RAMANMETRIX
2.9.1. To use RAMANMETRIX
2.10. INSPECTOR
2.10.1. To use INSPECTOR
3. Conclusions
References
Index
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