Electron Crystallography of Soluble and Membrane Proteins
Beschreibung:
The basic principle of electron crystallography is to calculate a 3D density map by combining the amplitudes obtained from electron diffraction patterns with the experimental phases calculated from images of two-dimensional crystals of membrane or soluble proteins. This technology is very well developed and has produced a number of atomic models of membrane proteins in a lipid environment. Focused on comprehensive experimental protocols, Electron Crystallography of Soluble and Membrane Proteins: Methods and Protocols covers the entire range of techniques used in electron crystallography, including protein sample preparation, 2D crystallization, and screening in negative stain over electron cryo-microscopy (cryo-EM) and data processing, as well as modeling of conformational changes. Additional chapters provide perspective on past, present, and future challenges as well as complementary methods. Written for the popular Methods in Molecular Biology(TM) series, the work contains the kind of detailed descriptions and implementation advice necessary to ensure successful results.
Comprehensive and cutting-edge, Electron Crystallography of Soluble and Membrane Proteins: Methods and Protocols serves laboratories new to the methods as well as state-of-the-art facilities pursuing this exciting area of protein science.
Introduction to Electron Crystallography.- Practical Aspects in Expression and Purification of Membrane Proteins for Structural Analysis.- Two-Dimensional Crystallization of Membrane Proteins by Reconstitution through Dialysis.- Monolayer Two-Dimensional Crystallization.- Screening for Two-Dimensional Crystals by Transmission Electron Microscopy of Negatively Stained Samples.- Low Dose Techniques and Cryo-Electron Microscopy.- Grid Preparation for Cryo-Electron Microscopy.- Recording High-Resolution Images of Two-Dimensional Crystals of Membrane Proteins.- The Collection of High-Resolution Electron Diffraction Data.- Image Processing of 2D Crystal Images.- Merging of Image Data in Electron Crystallography.- Evaluation of Electron Crystallographic Data From Images of Two-Dimensional Crystals.- Modeling, Docking, and Fitting of Atomic Structures to 3D Maps from Cryo-Electron Microscopy.- Phasing Electron Diffraction Data by Molecular Replacement: Strategy for Structure Determination and Refinement.- High-Throughput Methods for Electron Crystallography.- Automated Grid Handling and Image Acquisition for Two-Dimensional Crystal Screening.- Automation of Data Acquisition in Electron Crystallography.- Automation of Image Processing in Electron Crystallography.- Choice and Maintenance of Equipment for Electron Crystallography.- Future Developments in Instrumentation for Electron Crystallography.- Tubular Crystals and Helical Arrays: Structural Determination of HIV-1 Capsid Assemblies Using Iterative Helical Real-Space Reconstruction.- Single Particle Electron Microscopy.- Electron Tomography of Paracrystalline 2D Arrays.- High-Resolution Imaging of 2D Outer Membrane Protein F (OmpF) Crystals by Atomic Force Microscopy.- Determination of Soluble and Membrane Protein Structures by X-Ray Crystallography.- Solution Nuclear Magnetic Resonance (NMR) Spectroscopy.- Structure-Function Insights of Membrane and Soluble Proteins Revealed by Electron Crystallography.- Lipid Monolayer and Sparse Matrix Screening for Growing Two-Dimensional Crystals for Electron Crystallography: Methods and Examples.- Processing of Electron Diffraction Patterns with the XDP Program.- Future Directions of Electron Crystallography.