The N-terminal domains of syntaxin 7 and vti1b form three-helix bundles that differ in their ability to regulate SNARE complex assembly.
The SNAREs syntaxin 7, syntaxin 8, vti1b, and endobrevin/VAMP8 function in the fusion of late endosomes. Although the core complex formed by these SNAREs is very similar to the neuronal SNARE complex, it differs from the neuronal complex in that three of the four SNAREs contain extended N-terminal regions of unknown structure and function. Here we show that the N-terminal regions of syntaxin 7, syntaxin 8, and vti1b contain well folded alpha-helical domains. Multidimensional NMR spectroscopy revealed that in syntaxin 7 and vti1b, the domains form three-helix bundles resembling those of syntaxin 1, Sso1p, and Vam3p. The three-helix bundle domain of vti1b is the first of its kind identified in a SNARE outside the syntaxin family. Only syntaxin 7 adopts a closed conformation, whereas in vti1b and syntaxin 8, the N-terminal domains do not interact with the adjacent SNARE motifs. Accordingly, the rate of SNARE complex assembly is retarded about 7-fold when syntaxin 7 contains its N-terminal domain, whereas the N-terminal domains of vti1b and syntaxin 8 have no influence on assembly kinetics. We conclude that three-helix bundles represent a common fold for SNARE N-terminal domains, not restricted to the syntaxin family. However, they differ in their ability to adopt closed conformations and thus to regulate the assembly of SNARE complexes.
Department of Neurobiology, Max-Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
Wolfram
W
Antonin
Irina
I
Dulubova
Demet
D
Arac
Stefan
S
Pabst
Juliane
J
Plitzner
Josep
J
Rizo
Reinhard
R
Jahn
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D000954:Antigens, Surface / Q000737:chemistry; D002352:Carrier Proteins / Q000737:chemistry*; D003001:Cloning, Molecular; D003593:Cytoplasm / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000737:chemistry; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D050765:Qa-SNARE Proteins; D050766:Qb-SNARE Proteins; D050600:SNARE Proteins; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D050827:Syntaxin 1; D013696:Temperature; D013997:Time Factors; D033921:Vesicular Transport Proteins
D000954:Antigens, Surface; D002352:Carrier Proteins; D005656:Fungal Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D050765:Qa-SNARE Proteins; D050766:Qb-SNARE Proteins; D050600:SNARE Proteins; C494212:STX1A protein, human; D029701:Saccharomyces cerevisiae Proteins; D050827:Syntaxin 1; C107706:VAM3 protein, S cerevisiae; C106736:VTI1 protein, S cerevisiae; C491416:VTI1B protein, human; D033921:Vesicular Transport Proteins; D005982:Glutathione Transferase