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PMID: 9144250 Published · ppublish English Comparative Study Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Interaction of the synprint site of N-type Ca2+ channels with the C2B domain of synaptotagmin I.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 94 ·No. 10 ·1997-05-13 ·Pages 5405-10

Sheng ZH, Yokoyama CT, Catterall WA

Abstract

N-type Ca2+ channels mediate Ca2+ influx, which initiates fast exocytosis of neurotransmitters at synapses, and they interact directly with the SNARE proteins syntaxin and SNAP-25 (synaptosome-associated protein of 25 kDa) through a synaptic protein interaction (synprint) site in the intracellular loop connecting domains II and III of their alpha1B subunits. Introduction of peptides containing the synprint site into presynaptic neurons reversibly inhibits synaptic transmission, confirming the importance of interactions with this site in synaptic transmission. Here we report a direct interaction of the synprint peptide from N-type Ca2+ channels with synaptotagmin I, an important Ca2+ sensor for exocytosis, as measured by an affinity-chromatography binding assay and a solid-phase immunoassay. This interaction is mediated by the second C2 domain (C2B) of synaptotagmin I, but is not regulated by Ca2+. Using both immobilized recombinant proteins and native presynaptic membrane proteins, we found that the synprint peptide and synaptotagmin competitively interact with syntaxin. This interaction is Ca2+-dependent because of the Ca2+ dependence of the interactions between syntaxin and these two proteins. These results provide a molecular basis for a physical link between Ca2+ channels and synaptotagmin, and suggest that N-type Ca2+ channels may undergo a complex series of Ca2+-dependent interactions with multiple presynaptic proteins during neurotransmission.

MeSH Terms
Binding Sites Binding, Competitive Calcium/metabolism Calcium Channels/chemistry,metabolism Calcium Channels, L-Type Calcium-Binding Proteins Chromatography, Affinity Escherichia coli Glutathione Transferase Kinetics Membrane Glycoproteins/chemistry,metabolism Membrane Proteins/metabolism Nerve Tissue Proteins/chemistry,metabolism Peptide Fragments/metabolism Qa-SNARE Proteins Recombinant Fusion Proteins/chemistry,metabolism Sequence Tagged Sites Synaptic Transmission Synaptosomal-Associated Protein 25 Synaptotagmin I Synaptotagmins
Chemicals
Calcium Channels Calcium Channels, L-Type Calcium-Binding Proteins Membrane Glycoproteins Membrane Proteins Nerve Tissue Proteins Peptide Fragments Qa-SNARE Proteins Recombinant Fusion Proteins Synaptosomal-Associated Protein 25 Synaptotagmin I Synaptotagmins Glutathione Transferase Calcium
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Sheng Z H
Department of Pharmacology, Box 357280, University of Washington, Seattle, WA 98195-7280, USA.
Yokoyama C T
Catterall W A
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
0027-8424
DOI
10.1073/pnas.94.10.5405
Published
1997-05-13
Pages
5405-10
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC24691
Subset
IM
Grants
NIGMS NIH HHS · T32 GM007108 · United States
NIMH NIH HHS · F32 MH010775 · United States
NIGMS NIH HHS · T32 GM07108-19 · United States
NIMH NIH HHS · 1F32 MH10775-01 · United States
NINDS NIH HHS · R01 NS022625 · United States
NINDS NIH HHS · NS22625 · United States
External Links
PubMed source DOI: 10.1073/pnas.94.10.5405 PMC Full Text View journal details
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