Synaptobrevin transmembrane domain dimerization-revisited.
Synaptobrevin is a membrane-spanning soluble N-ethyl maleimid-sensitive factor (NSF) attachment protein receptor (SNARE) protein of synaptic vesicles that is essential for neurotransmitter release. Various lines of evidence indicate that it exists alternatively as a monomer, as a homodimer, as a heterodimer with synaptophysin, or as a ternary complex with other SNAREs at the various stages of the synaptic vesicle cycle. Homodimerization of synaptobrevin was previously shown by different authors to depend on its single transmembrane segment, and the crucial residues forming the helix-helix interface have been mapped. Since another recent study challenged these results, we reinvestigated this issue. Here, we show that native synaptobrevin can be cross-linked in synaptic vesicle membranes to a homodimer by disulfide bond formation between cysteine residues of the transmembrane segment. Further, we demonstrate that determination of synaptobrevin transmembrane segment interactions in membranes or in detergent solution requires careful control of experimental conditions. Thus, our present results corroborate that homodimerization of synaptobrevin is mediated by its transmembrane segment.
Roy
Rana
R
Lehrstuhl Chemie der Biopolymere, Technische Universität München, Weihenstephaner Berg 3, 85354 Freising, Germany.
Laage
Rico
R
Langosch
Dieter
D
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002942:Circular Dichroism; D017124:Conserved Sequence; D003432:Cross-Linking Reagents / Q000737:chemistry; D003545:Cysteine / Q000737:chemistry; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D008297:Male; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D050683:R-SNARE Proteins; D051381:Rats; D017208:Rats, Wistar; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D050600:SNARE Proteins; D017386:Sequence Homology, Amino Acid; D013570:Synaptic Membranes / Q000737:chemistry / Q000378:metabolism*; D033921:Vesicular Transport Proteins
D003432:Cross-Linking Reagents; D004220:Disulfides; D008565:Membrane Proteins; D050683:R-SNARE Proteins; D011993:Recombinant Fusion Proteins; D050600:SNARE Proteins; D033921:Vesicular Transport Proteins; D003545:Cysteine