Assembly of a ternary complex by the predicted minimal coiled-coil-forming domains of syntaxin, SNAP-25, and synaptobrevin. A circular dichroism study.
The assembly of target (t-SNARE) and vesicle-associated SNAP receptor (v-SNARE) proteins is a critical step for the docking of synaptic vesicles to the plasma membrane. Syntaxin-1A, SNAP-25, and synaptobrevin-2 (also known as vesicle-associated membrane protein, or VAMP-2) bind to each other with high affinity, and their binding regions are predicted to form a trimeric coiled-coil. Here, we have designed three peptides, which correspond to sequences located in the syntaxin-1A H3 domain, the C-terminal domain of SNAP-25, and a conserved central domain of synaptobrevin-2, that exhibit a high propensity to form a minimal trimeric coiled-coil. The peptides were synthesized by solid phase methods, and their interactions were studied by CD spectroscopy. In aqueous solution, the peptides were unstructured and showed no interactions with each other. In contrast, upon the addition of moderate amounts of trifluoroethanol (30%), the peptides adopted an alpha-helical structure and displayed both homomeric and heteromeric interactions. The interactions observed in ternary mixtures induce a stabilization of peptide structure that is greater than that predicted from individual binary interactions, suggesting the formation of a higher order structure compatible with the assembly of a trimeric coiled-coil.
Cánaves
J M
JM
Department of Biology, University of California San Diego, La Jolla, California 92093-0366, USA.
Montal
M
M
D000595:Amino Acid Sequence; D001923:Brain Chemistry; D002942:Circular Dichroism; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D050765:Qa-SNARE Proteins; D050683:R-SNARE Proteins; D013056:Spectrophotometry, Ultraviolet; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1
D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D050765:Qa-SNARE Proteins; D050683:R-SNARE Proteins; C494208:SNAP25 protein, human; C494212:STX1A protein, human; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1