Structure of the Munc18c/Syntaxin4 N-peptide complex defines universal features of the N-peptide binding mode of Sec1/Munc18 proteins

doi:10.1073/pnas.0701124104

Structure of the Munc18c/Syntaxin4 N-peptide complex defines universal features of the N-peptide binding mode of Sec1/Munc18 proteins

*Institute for Molecular Bioscience and Special Research Centre for Functional and Applied Genomics, University of Queensland, Brisbane, Queensland 4072, Australia; and
^§Garvan Institute of Medical Research, 384 Victoria Road, Darlinghurst, Sydney, New South Wales 2010, Australia

Edited by Thomas C. Südhof, The University of Texas Southwestern Medical Center, Dallas, TX, and approved April 3, 2007 (received for review February 7, 2007)

Abstract

Sec1/Munc18 proteins (SM proteins) bind to soluble NSF attachment protein receptors (SNAREs) and play an essential role in membrane fusion. Divergent modes of regulation have been proposed for different SM proteins indicating that they can either promote or inhibit SNARE assembly. This is in part because of discrete modes of binding that have been described for various SM/SNARE complexes. One mode suggests that SM proteins bind only to Syntaxins (Stx) preventing SNARE assembly, whereas in another they facilitate SNARE assembly and bind to SNARE complexes. The mammalian cell surface SM protein Munc18c binds to an N-peptide in Stx4, and this is compatible with its interaction with SNARE complexes. Here we describe the crystal structure of Munc18c in complex with the Stx4 N-peptide. This structure shows remarkable similarity with a yeast complex indicating that the mode of binding, which can accommodate SNARE complexes, is highly conserved throughout evolution. Modeling reveals the presence of the N-peptide binding mode in most but not all yeast and mammalian SM/Stx pairs, suggesting that it has coevolved to fulfill a specific regulatory function. It is unlikely that the N-peptide interaction alone accounts for the specificity in SM/SNARE binding, implicating other contact surfaces in this function. Together with other data, our results support a sequential two-state model for SM/SNARE binding involving an initial interaction via the Stx N-peptide, which somehow facilitates a second, more comprehensive interaction comprising other contact surfaces in both proteins.

Footnotes

^¶To whom correspondence should be addressed. E-mail: j.martin{at}imb.uq.edu.au
Author contributions: S.-H.H., D.E.J., and J.L.M. designed research; S.-H.H., C.F.L., C.L.G., and J.L.M. performed research; S.-H.H., C.F.L., C.L.G., D.E.J., and J.L.M. analyzed data; and S.-H.H., D.E.J., and J.L.M. wrote the paper.
↵ ^†Present address: School of Biomedical Sciences, University of Queensland, Brisbane, Queensland 4072, Australia.
↵ ^‡Present address: Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720-3206.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
Data deposition: The atomic coordinates and structure factors for Munc18c/Stx4_1–29 have been deposited in the Protein Data Bank, www.pdb.org (PDB ID code 2PJX).
This article contains supporting information online at www.pnas.org/cgi/content/full/0701124104/DC1.
Abbreviations:

SM protein,

Sec1/Munc18 protein;

SNARE,

soluble NSF attachment protein receptor;

Stx,

Syntaxin;

PDB,

Protein Data Bank.