1. The G Protein-Coupled Receptor Rhodopsin: A Historical Perspective Lukas Hofmann and Krzysztof Palczewski
Part II: Rhodopsin Expression, Regeneration and Purification for Structural Studies
2. Rhodopsin Purification from Dark Adapted Bovine Retina Elise Blankenship and David T. Lodowski
3. Mammalian Expression, Purification, and Crystallization of Rhodopsin Variants Daniel Mattle, Ankita Singhal, Georg Schmid, Roger Dawson, and Jörg Standfuss
4. Imaging of Rhodopsin Crystals with Two-Photon Microscopy Grazyna Palczewska and David Salom
Part III: Structure-function characterization of rhodopsin
5. Functional Stability of Rhodopsin in a Bicelle System: Evaluating G-Protein Activation by Rhodopsin in Bicelles Ali I. Kaya, T. M. Iverson, and Heidi E. Hamm
6. The Rhodopsin-Arrestin-1 Interaction in Bicelles Qiuyan Chen, Sergey A. Vishnivetskiy, Tiandi Zhuang, Min-Kyu Cho,Tarjani M. Thaker, Charles R. Sanders, Vsevolod V. Gurevich, and T. M. Iverson
7. Detection of Structural Waters and Their Role in Structural Dynamics of Rhodopsin Activation Liwen Wang and Mark Chance
8. Probing Conformational Changes in Rhodopsin Using Hydrogen-Deuterium Exchange Coupled to Mass Spectrometry Tivadar Orban and Yaroslav Tsybovsky
9. Analysis of Conformational Changes in Rhodopsin by Histidine Hydrogen-Deuterium Exchange David T. Lodowski and Masaru Miyagi
10. Investigation of Rhodopsin Dynamics in its Signaling State by Solid-State Deuterium NMR Spectroscopy Andrey V. Struts, Udeep Chawla, Suchithranga M.D.C. Perera, and Michael F. Brown
11. Sequential Structural Changes in Rhodopsin Occurring upon Photoactivation Naoki Kimata, Andreyah Pope, Dawood Rashid, Philip J. Reeves, and Steven O. Smith
12. Dynamic Single-Molecule Force Spectroscopy of Rhodopsin in Native Membranes Paul S.-H. Park and Daniel J. Müller
Part IV: Rhodopsin Supramolecular Organization and its Complexes with Partner Proteins
13. High-Resolution Atomic Force Microscopy Imaging of Rhodopsin in Rod Outer Segment Disk Membranes Patrick D. Bosshart, Andreas Engel, and Dimitrios Fotiadis
14. Detection of Rhodopsin Dimerization in situ by PIE-FCCS, a Time-Resolved Fluorescence Spectroscopy Adam W. Smith
15. Oligomeric State of Rhodopsin within Rhodopsin-Transducin Complex Probed with Succinylated Concanavalin A Beata Jastrzebska
16. Quantification of Arrestin-Rhodopsin Binding Stoichiometry Ciara C.M. Lally and Martha E. Sommer
17. Rhodopsin Transient Complexes Investigated by Systems Biology Approaches Daniele Dell’Orco
Part V: Rhodopsin and Photoreceptors
18. Three Dimensional Architecture of Murine Rod Cilium Revealed by Cryo-EM Theodore G. Wensel and Jared C. Gilliam
19. Monitoring of Rhodopsin Trafficking and Mistrafficking in Live Photoreceptors Kerrie H. Lodowski and Yoshikazu Imanishi
20. Measurements of Rhodopsin Diffusion within Signaling Membrane Microcompartments in Live Photoreceptors Mehdi Najafi and Peter D. Calvert
Part VI: Treatment Strategies of Retinal Degenerative Disease
21. Kinetics of Rhodopsin’s Chromophore Monitored in a Single Photoreceptor Leopold Adler IV, Nicholas P. Boyer, Chunhe Chen, and Yiannis Koutalos
22. Supplementation with Vitamin A Derivatives to Rescue Vision in Animal Models of Degenerative Retinal Diseases Lindsay Perusek, Akiko Maeda, and Tadao Maeda
23. Sustained Delivery of Retinoids to Prevent Photoreceptor Death Peter H. Tang and Rosalie K. Crouch
24. High-Throughput Screening Assays to Identify Small-Molecules Preventing Photoreceptor Degeneration Caused by the Rhodopsin P23H Mutation Yuanyuan Chen and Hong Tang
25. Gene Therapy to Rescue Retinal Degeneration Caused by Mutations in Rhodopsin Brian P. Rossmiller, Renee C. Ryals, and Alfred S. Lewin
Beata Jastrzebska, Ph.D. Department of Pharmacology, W317 Case Western Reserve University 10900 Euclid Avenue Cleveland, OH 44106-4965
This volume explores the technological improvements in protein engineering, expression, purification, and crystallization of several rhodopsin photoactive intermediates, thus increasing our understanding of rhodopsin activation. The first chapters of the book focus on methods developed to study fundamentals of rhodopsin structure and function, starting with improved purification protocols of native and mutated rhodopsin, followed by methods used for rhodopsin reconstitution into lipid bilayers stabilizing rhodopsin function properties, and finally describing recently developed methods to study structural dynamics of rhodopsin activation and its mechanistic properties. Subsequently, chapters underline various techniques that have been developed to visualize the rhodopsin dimer and to study its functional significance. The next few chapters highlight cutting-edge imaging techniques of photoreceptors, rhodopsin trafficking, and its diffusion within signaling membranes. Finally the book concludes with recent developments that could be potentially beneficial in patient treatments, and treatment strategies for retinal degenerative diseases. Written in the highly successful Methods in Molecular Biology series format, the chapters include the kind of detailed description and implementation advice that is crucial for getting optimal results in the laboratory.
Timely and practical, Rhodopsin: Methods and Protocols reaches out to researchers and health practitioners, and provides timely protocol useful for studying structural and functional properties of rhodopsin.