Structural analysis of the neuronal SNARE protein syntaxin-1A

Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544, USA.
Biochemistry (Impact Factor: 3.01). 08/2000; 39(29):8470-9. DOI: 10.1021/bi0003994
Source: PubMed

ABSTRACT Intracellular trafficking depends on the docking and fusion of transport vesicles with cellular membranes. Central to docking and fusion is the pairing of SNARE proteins (soluble NSF attachment protein receptors) associated with the vesicle and target membranes (v- and t-SNAREs, respectively). Here, the X-ray structure of an N-terminal conserved domain of the neuronal t-SNARE syntaxin-1A was determined to a resolution of 1.9 A using multiwavelength anomalous diffraction. This X-ray structure, which is in general agreement with an NMR structure of a similar fragment, provides new insight into the interaction surface between the N-terminal domain and the remainder of the protein. In vitro characterization of the intact cytoplasmic domain of syntaxin revealed that it forms dimers, and probably tetramers, at low micromolar concentrations, with concomitant structural changes that can be detected by limited proteolysis. These observations suggest that the promiscuity characteristic of pairing between v-SNAREs and t-SNAREs extends to the formation of homo-oligomeric t-SNARE complexes as well. They also suggest a potential role for the neuronal Sec1 protein (nSec1) in preventing the formation of syntaxin multimers.

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Available from: Robert Fairman, Jul 15, 2015
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    • "This structure, denoted Habc, may fold back onto the SNARE domain and enable the molecule to adopt a 'closed' conformation that prevents assembly of the core fusion complex (i.e. Habc could act as an autoinhibitor of SNARE function) (Lerman et al., 2000; Teng et al., 2001). A second type of autonomous N-terminal structure, a profilin-like domain, is present within some R-SNAREs as well as putatively unrelated proteins (Rossi et al., 2004). "
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    • "Syntaxin was reconstituted into vesicles at submicromolar concentrations, which is below the reported micromolar K D for self-interaction (Lerman et al., 2000)). In order to achieve lateral separation between dye labeled molecules in the bilayer greater than the optical resolution of the microscope, vesicles containing syntaxin were diluted with protein-free vesicles before bilayer deposition. "
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    • "For example, only higher-molecular-weight complexes of AtSyp122, potentially including SNARE homodimers, were suppressed when incubated with the Sp3 fragment of AtSyp122 in cell extracts including each of the plasma membrane SNAREs (Fig. 7). The fact that these proteins pair and bind, even after solubilisation, underscores a stability of these complexes that accords with previous analyses of these extra-canonical SNARE interactions (Antonin et al., 2002a; Lerman et al., 2000). Of course, the ability for these protein domains to interact in vitro does not confirm a similar behaviour in vivo, nor does it preclude other interactions that might compete for SNARE complex elements and thereby suppress vesicle traffic. "
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