The N-terminal peptide of the syntaxin Tlg2p modulates binding of its closed conformation to Vps45p
Department of Biochemistry and Molecular Pharmacology
Binding, Competitive; Circular Dichroism; Electrophoretic Mobility Shift Assay; Immunoblotting; Kinetics; Models, Molecular; Mutation; Protein Binding; Protein Conformation; Protein Structure, Tertiary; Qa-SNARE Proteins; Recombinant Proteins; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Vesicular Transport Proteins
Life Sciences | Medicine and Health Sciences
The Sec1/Munc18 (SM) protein family regulates intracellular trafficking through interactions with individual SNARE proteins and assembled SNARE complexes. Revealing a common mechanism of this regulation has been challenging, largely because of the multiple modes of interaction observed between SM proteins and their cognate syntaxin-type SNAREs. These modes include binding of the SM to a closed conformation of syntaxin, binding to the N-terminal peptide of syntaxin, binding to assembled SNARE complexes, and/or binding to nonsyntaxin SNAREs. The SM protein Vps45p, which regulates endosomal trafficking in yeast, binds the conserved N-terminal peptide of the syntaxin Tlg2p. We used size exclusion chromatography and a quantitative fluorescent gel mobility shift assay to reveal an additional binding site that does not require the Tlg2p N-peptide. Characterization of Tlg2p mutants and truncations indicate that this binding site corresponds to a closed conformation of Tlg2p. Furthermore, the Tlg2p N-peptide competes with the closed conformation for binding, suggesting a fundamental regulatory mechanism for SM-syntaxin interactions in SNARE assembly and membrane fusion.
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Citation: Proc Natl Acad Sci U S A. 2009 Aug 25;106(34):14303-8. Epub 2009 Aug 10. Link to article on publisher's site
DOI of Published Version
Proceedings of the National Academy of Sciences of the United States of America
Furgason, Melonnie Lynn Marie; MacDonald, Chris; Shanks, Scott G.; Ryder, Sean P.; Bryant, Nia J.; and Munson, Mary, "The N-terminal peptide of the syntaxin Tlg2p modulates binding of its closed conformation to Vps45p" (2009). Open Access Articles. 2203.