Single-molecule FRET–derived model of the synaptotagmin 1–SNARE fusion complex

• Published in: Nature structural & molecular biology Vol. 17; no. 3; pp. 318 - 324
• Main Authors: Choi, Ucheor B, Strop, Pavel, Vrljic, Marija, Chu, Steven, Brunger, Axel T, Weninger, Keith R
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group US 01.03.2010; Nature Publishing Group
• Subjects: 631/57/2272/2273; 631/80/313/2104; 631/80/313/2378; 631/80/86/1999; Animals; Binding sites; Biochemistry; Biological Microscopy; Biomedical and Life Sciences; Calcium - metabolism; Calcium channels; Cellular biology; Cellular signal transduction; Chromatography, Affinity; Chromatography, Gel; Chromatography, Ion Exchange; Crystal structure; Energy transfer; Fluorescence; Fluorescence Resonance Energy Transfer; Life Sciences; Membrane Biology; Membrane proteins; Membranes; Molecular biology; Neural transmission; Physiological aspects; Protein Binding; Protein Structure; Proteins; Rats; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Resonance; SNARE Proteins - chemistry; SNARE Proteins - genetics; SNARE Proteins - metabolism; Synaptotagmin I - chemistry; Synaptotagmin I - genetics; Synaptotagmin I - metabolism; United States
• ISSN: 1545-9993; 1545-9985
• DOI: 10.1038/nsmb.1763

Single-molecule FRET studies have resulted in an experimentally derived model of a synaptotagmin–SNARE complex. In this complex of SNARE with synaptotagmin 1, the arrangement of the Ca 2+ -binding loops is similar to that of the structure of SNARE-induced Ca 2+ -bound synaptotagmin 3. This suggests a common molecular mechanism by which the synaptotagmin–SNARE interaction plays a role in Ca 2+ -triggered vesicle fusion. Synchronous neurotransmission is triggered when Ca 2+ binds to synaptotagmin 1 (Syt1), a synaptic-vesicle protein that interacts with SNAREs and membranes. We used single-molecule fluorescence resonance energy transfer (FRET) between synaptotagmin's two C2 domains to determine that their conformation consists of multiple states with occasional transitions, consistent with domains in random relative motion. SNARE binding results in narrower intrasynaptotagmin FRET distributions and less frequent transitions between states. We obtained an experimentally determined model of the elusive Syt1–SNARE complex using a multibody docking approach with 34 FRET-derived distances as restraints. The Ca 2+ -binding loops point away from the SNARE complex, so they may interact with the same membrane. The loop arrangement is similar to that of the crystal structure of SNARE-induced Ca 2+ -bound Syt3, suggesting a common mechanism by which the interaction between synaptotagmins and SNAREs aids in Ca 2+ -triggered fusion.