Title: Syntaxin1a variants lacking an N-peptide or bearing the LE mutation bind to Munc18a in a closed conformation

Abstract

In neurons, soluble N-ethylmaleimide–sensitive factor attachment receptor (SNARE) proteins drive the fusion of synaptic vesicles to the plasma membrane through the formation of a four-helix SNARE complex. Members of the Sec1/Munc18 protein family regulate membrane fusion through interactions with the syntaxin family of SNARE proteins. The neuronal protein Munc18a interacts with a closed conformation of the SNARE protein syntaxin1a (Syx1a) and with an assembled SNARE complex containing Syx1a in an open conformation. The N-peptide of Syx1a (amino acids 1–24) has been implicated in the transition of Munc18a-bound Syx1a to Munc18a-bound SNARE complex, but the underlying mechanism is not understood. In addition, we report the X-ray crystal structures of Munc18a bound to Syx1a with and without its native N-peptide (Syx1aΔN), along with small-angle X-ray scattering (SAXS) data for Munc18a bound to Syx1a, Syx1aΔN, and Syx1a L165A/E166A (LE), a mutation thought to render Syx1a in a constitutively open conformation. We show that all three complexes adopt the same global structure, in which Munc18a binds a closed conformation of Syx1a. We also identify a possible structural connection between the Syx1a N-peptide and SNARE domain that might be important for the transition of closed-to-open Syx1a in SNARE complex assembly. Although the role of themore » N-peptide in Munc18a-mediated SNARE complex assembly remains unclear, our results demonstrate that the N-peptide and LE mutation have no effect on the global conformation of the Munc18a–Syx1a complex.« less

Authors:

Colbert, Karen N. ^[1]; Hattendorf, Douglas A. ^[1]; Weiss, Thomas M. ^[2]; Burkhardt, Pawel ^[3]; Fasshauer, Dirk ^[4]; Weis, William I. ^[1]

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+ Show Author Affiliations

1. Stanford Univ. School of Medicine, Stanford, CA (United States)
2. SLAC National Accelerator Lab., Menlo Park, CA (United States)
3. Max Planck Institute for Biophysical Chemistry, Gottingen (Germany); Univ. of California, Berkeley, CA (United States)
4. Max Planck Institute for Biophysical Chemistry, Gottingen (Germany); Univ. of Lausanne, Lausanne (Switzerland)

Publication Date:

Mon Jul 15 00:00:00 EDT 2013

Research Org.:

SLAC National Accelerator Laboratory (SLAC), Menlo Park, CA (United States)

Sponsoring Org.:

USDOE Office of Science (SC)

OSTI Identifier:

1132335

Report Number(s):

SLAC-REPRINT-2014-119
Journal ID: ISSN 0027-8424

Grant/Contract Number:  

AC02-76SF00515

Resource Type:

Accepted Manuscript

Journal Name:

Proceedings of the National Academy of Sciences of the United States of America

Additional Journal Information:

Journal Volume: 110; Journal Issue: 31; Journal ID: ISSN 0027-8424

Publisher:

National Academy of Sciences, Washington, DC (United States)

Country of Publication:

United States

Language:

English

Subject:

59 BASIC BIOLOGICAL SCIENCES; membrane trafficking; SM proteins; protein crystallography