Homo- and heterooligomeric SNARE complexes studied by site-directed spin labeling

SNARE (soluble NSF acceptor protein receptor) proteins are thought to media te membrane fusion by assem bling into heterooligomeric complexes that conn ect the fusing membranes and initiate the fusion reaction. Here we used sit e-directed spin labeling to map conformational changes that occur upon homo - and heterooligomeric complex formation of neuronal SNARE proteins. We fou nd that the soluble domains of synaptobrevin, SNAP-25, and syntaxin 1 are u nstructured. At higher concentrations, the SNARE motif of syntaxin 1 forms homooligomeric helical bundles with at least some of the alpha -helices ali gned in parallel. In the assembled SNARE complex, mapping of thirty side ch ain positions yielded spectra which are in good agreement with the recently published crystal structure. The loop region of SNAP-25 that connects the two SNARE motifs is largely unstructured. C-terminal truncation of synaptob revin resulted in complexes that are completely folded N-terminal of the tr uncation but become unstructured at the C-terminal end. The binary complex of syntaxin and SNAP-25 consists of a parallel four helix-bundle with prope rties resembling that of the ternary complex.