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Proc Natl Acad Sci U S A. 2009 Aug 25;106(34):14303-8. doi: 10.1073/pnas.0902976106. Epub 2009 Aug 10.

The N-terminal peptide of the syntaxin Tlg2p modulates binding of its closed conformation to Vps45p.

Author information

1
Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, MA 01605, USA.

Abstract

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.

PMID:
19667197
PMCID:
PMC2732825
DOI:
10.1073/pnas.0902976106
[Indexed for MEDLINE]
Free PMC Article

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