title: Structural Characterization of Membrane-Curving Proteins: Site-Directed Spin Labeling, EPR, and Computational Refinement.
abstract: Endocytosis and other membrane remodeling processes require the coordinated generation of different membrane shapes. Proteins capable of manipulating lipid bilayers mediate these events using mechanisms that are not fully understood. Progress is limited by the small number of structures solved for proteins bound to different membrane shapes and tools capable of resolving such information. However, recent studies have shown site-directed spin labeling (SDSL) in combination with electron paramagnetic resonance (EPR) to be capable of obtaining high-resolution structural information for proteins bound to different membrane shapes. This technique can be applied to proteins with no known structure or proteins with structures known in solution. By refining the data obtained by EPR with computational modeling, 3D structures or structural models of membrane-bound proteins can be generated. In this chapter, we highlight the basic considerations and steps required to investigate the structures of membrane-bound proteins using SDSL, EPR, and computational refinement.
authors.affiliation: Department of Biochemistry and Molecular Biology, Zilkha Neurogenetic Institute, University of Southern California, Los Angeles, California, USA.
authors.forename: Mark R
authors.initials: MR
authors.lastname: Ambroso
authors.affiliation: Department of Pharmacology and Pharmaceutical Sciences, University of Southern California, Los Angeles, California, USA.
authors.forename: Ian S
authors.initials: IS
authors.lastname: Haworth
authors.affiliation: Department of Biochemistry and Molecular Biology, Zilkha Neurogenetic Institute, University of Southern California, Los Angeles, California, USA. Electronic address: langen@usc.edu.
authors.forename: Ralf
authors.initials: R
authors.lastname: Langen
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D006801:Humans; D008051:Lipid Bilayers / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry*; D008958:Models, Molecular; D011487:Protein Conformation; D013113:Spin Labels
chemical_list: D008051:Lipid Bilayers; D008565:Membrane Proteins; D013113:Spin Labels
keywords: Continuous wave EPR; Double electron–electron resonance; Lipid building; Power saturation; Simulated annealing