title: Solution and membrane-bound conformations of the tandem C2A and C2B domains of synaptotagmin 1: Evidence for bilayer bridging.
abstract: Synaptotagmin 1 (syt1) is a synaptic vesicle membrane protein that functions as the Ca(2)(+) sensor in neuronal exocytosis. Here, site-directed spin labeling was used to generate models for the solution and membrane-bound structures of a soluble fragment of syt1 containing its two C2 domains, C2A and C2B. In solution, distance restraints between the two C2 domains of syt1 were measured using double electron-electron resonance and used in a simulated annealing routine to generate models for the structure of the tandem C2A-C2B fragment. The data indicate that the two C2 domains are flexibly linked and do not interact with each other in solution, with or without Ca(2+). However, the favored orientation is one where the Ca(2+)-binding loops are oriented in opposite directions. A similar approach was taken for membrane-associated C2A-C2B, combining both distances and bilayer depth restraints with simulated annealing. The restraints can only be satisfied if the Ca(2+) and membrane-binding surfaces of the domains are oriented in opposite directions so that C2A and C2B are docked to opposing bilayers. The result suggests that syt1 functions to bridge across the vesicle and plasma membrane surfaces in a Ca(2+)-dependent manner.
authors.affiliation: Department of Chemistry and Biophysics, University of Virginia, Charlottesville, 22904-4319, USA.
authors.forename: Dawn Z
authors.initials: DZ
authors.lastname: Herrick
authors.forename: Weiwei
authors.initials: W
authors.lastname: Kuo
authors.forename: Hao
authors.initials: H
authors.lastname: Huang
authors.forename: Charles D
authors.initials: CD
authors.lastname: Schwieters
authors.forename: Jeffrey F
authors.initials: JF
authors.lastname: Ellena
authors.forename: David S
authors.initials: DS
authors.lastname: Cafiso
mesh_terms: D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D008051:Lipid Bilayers / Q000378:metabolism*; D008958:Models, Molecular; D018583:Pliability / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D051381:Rats; D012996:Solutions; D050863:Synaptotagmin I / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D008051:Lipid Bilayers; D012996:Solutions; D050863:Synaptotagmin I; D002118:Calcium