An Extended Helical Conformation in Domain 3a of Munc18-1 Provides a Template for SNARE (Soluble N-Ethylmaleimide-sensitive Factor Attachment Protein Receptor) Complex Assembly*
| Autor: | Thomas H. Söllner, Joerg Malsam, Maximilian Pfau, Daniel Parisotto, Matthias P. Mayer, Andrea Scheutzow, Klemens Wild, Irmgard Sinning |
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| Jazyk: | angličtina |
| Rok vydání: | 2014 |
| Předmět: |
Vesicle fusion
VAMP2 Vesicle-Associated Membrane Protein 2 Vesicle docking Mutation Missense SNAP25 Syntaxin 1 Munc-18 Cell Biology Biology Biochemistry Protein Structure Secondary Protein Structure Tertiary Rats Structure-Activity Relationship Munc18 Proteins Amino Acid Substitution Biophysics Animals Molecular Biology SNARE complex assembly |
| Popis: | Munc18-1, a SEC1/Munc18 protein and key regulatory protein in synaptic transmission, can either promote or inhibit SNARE complex assembly. Although the binary inhibitory interaction between Munc18-1 and closed syntaxin 1 is well described, the mechanism of how Munc18-1 stimulates membrane fusion remains elusive. Using a reconstituted assay that resolves vesicle docking, priming, clamping, and fusion during synaptic exocytosis, we show that helix 12 in domain 3a of Munc18-1 stimulates SNAREpin assembly and membrane fusion. A single point mutation (L348R) within helix 12 selectively abolishes VAMP2 binding and the stimulatory function of Munc18-1 in membrane fusion. In contrast, targeting a natural switch site (P335A) at the start of helix 12, which can result in an extended α-helical conformation, further accelerates lipid-mixing. Together with structural modeling, the data suggest that helix 12 provides a folding template for VAMP2, accelerating SNAREpin assembly and membrane fusion. Analogous SEC1/Munc18-SNARE interactions at other transport steps may provide a general mechanism to drive lipid bilayer merger. At the neuronal synapse, Munc18-1 may convert docked synaptic vesicles into a readily releasable pool. |
| Databáze: | OpenAIRE |
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