STRUCTURE AND CONFORMATIONAL-CHANGES IN NSF AND ITS MEMBRANE-RECEPTORCOMPLEXES VISUALIZED BY QUICK-FREEZE DEEP-ETCH ELECTRON-MICROSCOPY/

Using quick-freeze/deep-etch electron microscopy of recombinant protei ns adsorbed to mica, we show that NSF, the oligomeric ATPase involved in membrane fusion, is a hollow 10 x 16 nm cylinder whose conformation depends upon nucleotide binding. Depleted of nucleotide, NSF converts to a ''splayed'' protease-sensitive conformation that reveals its sub unit composition. NSF's synaptic membrane substrate, the ternary SNARE complex containing syntaxin, SNAP-25, and synaptobrevin, is a 4 x 14 nm rod with a ''tail'' at one end, corresponding to the N-terminus of syntaxin. Using epitope tags, antibodies, and maltose-binding protein markers, we find that syntaxin and synaptobrevin are aligned in parall el in the complex, with their membrane anchors located at the same end of the rod. This SNARE rod binds with alpha-SNAP to one end of the NS F cylinder to form an asymmetric ''20S'' complex. Together, these imag es suggest how NSF could dissociate the SNARE complex and how associat ion and dissociation of the complex could be related to membrane fusio n.