title: Mixed and non-cognate SNARE complexes. Characterization of assembly and biophysical properties.
abstract: Assembly of soluble N-ethylmaleimide-sensitive fusion attachment protein receptor (SNARE) proteins between two opposing membranes is thought to be the key event that initiates membrane fusion. Many new SNARE proteins have recently been localized to distinct intracellular compartments, supporting the view that sets of specific SNAREs are specialized for distinct trafficking steps. We have now investigated whether other SNAREs can form complexes with components of the synaptic SNARE complex including synaptobrevin/VAMP 2, SNAP-25, and syntaxin 1. When the Q-SNAREs syntaxin 2, 3, and 4, and the R-SNARE endobrevin/VAMP 8 were used in various combinations, heat-resistant complexes were formed. Limited proteolysis revealed that these complexes contained a protease-resistant core similar to that of the synaptic complex. All complexes were disassembled by the ATPase N-ethylmaleimide-sensitive fusion protein and its cofactor alpha-SNAP. Circular dichroism spectroscopy showed that major conformational changes occur during assembly, which are associated with induction of structure from unstructured monomers. Furthermore, no preference for synaptobrevin was observed during the assembly of the synaptic complex when endobrevin/VAMP 8 was present in equal concentrations. We conclude that cognate and non-cognate SNARE complexes are very similar with respect to biophysical properties, assembly, and disassembly, suggesting that specificity of membrane fusion in intracellular membrane traffic is not due to intrinsic specificity of SNARE pairing.
authors.affiliation: Department of Neurobiology, Max Planck Institute for Biophysical Chemistry, D-37077 Göttingen, Germany.
authors.forename: D
authors.initials: D
authors.lastname: Fasshauer
authors.forename: W
authors.initials: W
authors.lastname: Antonin
authors.forename: M
authors.initials: M
authors.lastname: Margittai
authors.forename: S
authors.initials: S
authors.lastname: Pabst
authors.forename: R
authors.initials: R
authors.lastname: Jahn
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism; D003001:Cloning, Molecular; D003902:Detergents; D019286:Endopeptidase K / Q000378:metabolism; D008561:Membrane Fusion; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000378:metabolism*; D017433:Protein Structure, Secondary; D050765:Qa-SNARE Proteins; D050683:R-SNARE Proteins; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D050600:SNARE Proteins; D016415:Sequence Alignment; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; D033921:Vesicular Transport Proteins
chemical_list: D002352:Carrier Proteins; D003902:Detergents; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D050765:Qa-SNARE Proteins; D050683:R-SNARE Proteins; D011994:Recombinant Proteins; D050600:SNARE Proteins; C494210:Snap25 protein, rat; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; C494213:Stx1a protein, rat; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; C491830:Vamp8 protein, rat; D033921:Vesicular Transport Proteins; D019286:Endopeptidase K