Molecular Dynamics Simulations of SNARE Complex Unzipping

Abstract

The SNARE proteins facilitate biological membrane fusion. The neuronal SNARE proteins, namely VAMP-2, also called synaptobrevin 2, SNAP-25, and syntaxin-1A together form a coiled-coil complex. The formation of this SNARE complex is vital for vesicle-plasma membrane fusion resulting in neurotransmitter exocytosis. VAMP-2, located on vesicular membrane, and syntaxin-1A, located on the plasma membrane, are transmembrane proteins, each with a single transmembrane domain. SNAP-25 is associated with the plasma membrane through lipid anchors and forms a binary t-SNARE complex with syntaxin-1A. For fusion to occur, the VAMP-2 zips up with the binary t-SNARE complex to form a trans SNARE complex. To better understand the mechanism and energetics of formation of the ternary SNARE complex, we carried out all-atom molecular dynamics simulations using the OPLS-AA force field to investigate the unzipping of the SNARE complex. The initial structure of the soluble part of the SNARE complex was based on the 1SFC pdb (Sutton et al., 1998 Nature 95:347-53). The missing residues R262-K265 of syntaxin-1A were modeled based on 3IPD pdb (Stein et al., 2009 Nature 460:525-28) and the two C-terminal residues S205-G206 of SNAP25 added as helical extension using Modeller. For unzipping, forces were applied between the main chain atoms of syntaxin K265 and those of VAMP-2 N92, as in unzipping experiments using optical tweezers (Gao et al., 2012 Science 337:1340-43). Application of harmonic forces between these residues resulted in unzipping of layers 8 to 6. Unexpectedly, this unzipping occurred through separation of syntaxin while the SNAP-25 SNARE domains SN1 and SN2 remained associated with VAMP-2. These results suggest that mechanical unzipping may not necessarily lead to separation of VAMP-2 from the binary t-SNARE complex and that the C-terminal association of SNAP-25 with syntaxin may be weaker than that with VAMP-2. Supported by ERC Advanced grant No.322699.