biostudies-literatureKhan YANIMH NIH HHSNCI NIH HHSNIGMS NIH HHSNIH HHS1708-1720https://www.ebi.ac.uk/biostudies/studies/S-EPMC12440825biostudies-literatureUnknown32(9)SNARE (soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein receptor) proteins drive membrane fusion at different cell compartments as their core domains zipper into a parallel four-helix bundle. After fusion, these bundles are disassembled by the AAA+ (ATPase associated with diverse cellular activities) protein Sec18/NSF and its adaptor Sec17/α-SNAP to make them available for subsequent rounds of membrane fusion. SNARE domains are often flanked by C-terminal transmembrane or N-terminal domains. Previous structures of the NSF-α-SNAP-SNARE complex revealed binding to the D1 ATPase pore, posing a topological constraint as SNARE transmembrane domains would prevent complete substrate threading as suggested for other AAA+ systems. Using mass spectrometry in yeast cells, we show N-Nature structural & molecular biologySNARE disassembly requires Sec18/NSF side loading.PMC12440825S10 OD030473P30 CA124435F31 MH134477R35 GM118037R01 MH063105Mckenzie GSingal BMontabana EEsquivies LKhan YAMclaughlin TChoi UBPfuetzner RADeLong KBrunger ATWickner WTWhite KILiu FfalseSNARE disassembly requires Sec18/NSF side loading.SNARE (soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein receptor) proteins drive membrane fusion at different cell compartments as their core domains zipper into a parallel four-helix bundle. After fusion, these bundles are disassembled by the AAA+ (ATPase associated with diverse cellular activities) protein Sec18/NSF and its adaptor Sec17/α-SNAP to make them available for subsequent rounds of membrane fusion. SNARE domains are often flanked by C-terminal transmembrane or N-terminal domains. Previous structures of the NSF-α-SNAP-SNARE complex revealed binding to the D1 ATPase pore, posing a topological constraint as SNARE transmembrane domains would prevent complete substrate threading as suggested for other AAA+ systems. Using mass spectrometry in yeast cells, we show N-2025-01-01T00:00:00Z2025 Sep2026-06-03T08:00:58.524Z2026-04-26T03:12:05.632ZS-EPMC124408254060431010.1038/s41594-025-01590-w