title: Fast Photochemical Oxidation of Proteins Maps the Topology of Intrinsic Membrane Proteins: Light-Harvesting Complex 2 in a Nanodisc.
abstract: Although membrane proteins are crucial participants in photosynthesis and other biological processes, many lack high-resolution structures. Prior to achieving a high-resolution structure, we are investigating whether MS-based footprinting can provide coarse-grained protein structure by following structural changes that occur upon ligand binding, pH change, and membrane binding. Our platform probes topology and conformation of membrane proteins by combining MS-based footprinting, specifically fast photochemical oxidation of proteins (FPOP), and lipid Nanodiscs, which are more similar to the native membrane environment than are the widely used detergent micelles. We describe here results that show a protein's outer membrane regions are more heavily footprinted by OH radicals whereas the regions spanning the lipid bilayer remain inert to the labeling. Nanodiscs generally exhibit more protection of membrane proteins compared to detergent micelles and less shielding to those protein residues that exist outside the membrane. The combination of immobilizing the protein in Nanodiscs and footprinting with FPOP is a feasible approach to map extra-membrane protein surfaces, even at the amino-acid level, and to illuminate intrinsic membrane protein topology.
authors: Department of Chemistry, Washington University in St. Louis , St. Louis, Missouri 63130, United States.|Photosynthetic Antenna Research Center, Washington University in St. Louis , St. Louis, Missouri 63130, United States.
authors: Yue
authors: Y
authors: Lu
authors: Department of Chemistry, Washington University in St. Louis , St. Louis, Missouri 63130, United States.|Photosynthetic Antenna Research Center, Washington University in St. Louis , St. Louis, Missouri 63130, United States.
authors: Hao
authors: H
authors: Zhang
authors: Photosynthetic Antenna Research Center, Washington University in St. Louis , St. Louis, Missouri 63130, United States.
authors: Dariusz M
authors: DM
authors: Niedzwiedzki
authors: Photosynthetic Antenna Research Center, Washington University in St. Louis , St. Louis, Missouri 63130, United States.|Department of Energy, Environmental and Chemical Engineering, Washington University in St. Louis , St. Louis, Missouri 63130, United States.
authors: Jing
authors: J
authors: Jiang
authors: Department of Chemistry, Washington University in St. Louis , St. Louis, Missouri 63130, United States.|Photosynthetic Antenna Research Center, Washington University in St. Louis , St. Louis, Missouri 63130, United States.|Department of Biology, Washington University in St. Louis , St. Louis, Missouri 63130, United States.
authors: Robert E
authors: RE
authors: Blankenship
authors: Department of Chemistry, Washington University in St. Louis , St. Louis, Missouri 63130, United States.|Photosynthetic Antenna Research Center, Washington University in St. Louis , St. Louis, Missouri 63130, United States.
authors: Michael L
authors: ML
authors: Gross
mesh_terms: D017665:Hydroxyl Radical / Q000737:chemistry / Q000378:metabolism; D045342:Light-Harvesting Protein Complexes / Q000737:chemistry / Q000378:metabolism*; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008823:Micelles; D049329:Nanostructures / Q000737:chemistry*; D010084:Oxidation-Reduction; D055668:Photochemical Processes; D025941:Protein Interaction Mapping
chemical_list: D045342:Light-Harvesting Protein Complexes; D008051:Lipid Bilayers; D008565:Membrane Proteins; D008823:Micelles; D017665:Hydroxyl Radical