title: A conformational switch in complexin is required for synaptotagmin to trigger synaptic fusion.
abstract: The crystal structure of complexin bound to a prefusion SNAREpin mimetic shows that the accessory helix extends away from the SNAREpin in an 'open' conformation, binding another SNAREpin and inhibiting its assembly, to clamp fusion. In contrast, the accessory helix in the postfusion complex parallels the SNARE complex in a 'closed' conformation. Here we use targeted mutations, FRET spectroscopy and a functional assay that reconstitutes Ca(2+)-triggered exocytosis to show that the conformational switch from open to closed in complexin is needed for synaptotagmin-Ca(2+) to trigger fusion. Triggering fusion requires the zippering of three crucial aspartate residues in the switch region (residues 64-68) of v-SNARE. Conformational switching in complexin is integral to clamp release and is probably triggered when its accessory helix is released from its trans-binding to the neighboring SNAREpin, allowing the v-SNARE to complete zippering and open a fusion pore.
authors.affiliation: Department of Cell Biology, Yale University School of Medicine, New Haven, Connecticut, USA.
authors.forename: Shyam S
authors.initials: SS
authors.lastname: Krishnakumar
authors.forename: Daniel T
authors.initials: DT
authors.lastname: Radoff
authors.forename: Daniel
authors.initials: D
authors.lastname: Kümmel
authors.forename: Claudio G
authors.initials: CG
authors.lastname: Giraudo
authors.forename: Feng
authors.initials: F
authors.lastname: Li
authors.forename: Lavan
authors.initials: L
authors.lastname: Khandan
authors.forename: Stephanie Wood
authors.initials: SW
authors.lastname: Baguley
authors.forename: Jeff
authors.initials: J
authors.lastname: Coleman
authors.forename: Karin M
authors.initials: KM
authors.lastname: Reinisch
authors.forename: Frederic
authors.initials: F
authors.lastname: Pincet
authors.forename: James E
authors.initials: JE
authors.lastname: Rothman
mesh_terms: D033942:Adaptor Proteins, Vesicular Transport / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D018360:Crystallography, X-Ray; D006801:Humans; D008561:Membrane Fusion / Q000502:physiology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D017434:Protein Structure, Tertiary; D051381:Rats; D050825:Synaptosomal-Associated Protein 25 / Q000737:chemistry / Q000378:metabolism; D050857:Synaptotagmins / Q000378:metabolism / Q000502:physiology*; D050827:Syntaxin 1 / Q000737:chemistry / Q000378:metabolism; D050988:Vesicle-Associated Membrane Protein 2 / Q000737:chemistry / Q000378:metabolism / Q000502:physiology
chemical_list: D033942:Adaptor Proteins, Vesicular Transport; D009419:Nerve Tissue Proteins; C494208:SNAP25 protein, human; C494213:Stx1a protein, rat; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; C494807:VAMP2 protein, human; D050988:Vesicle-Associated Membrane Protein 2; C095992:complexin I; D050857:Synaptotagmins