Phosphorylation of the synaptic protein interaction site on N-type calcium channels inhibits interactions with SNARE proteins

Charles T. Yokoyama, Zu Hang Sheng, William A. Catterall

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131 Citations (Scopus)


The synaptic protein interaction (synprint) site on the N-type calcium channel α(1B) subunit binds to the soluble N-ethylmaleimide-sensitive attachment factor receptor (SNARE) proteins syntaxin and synaptosomal protein of 25 kDa (SNAP-25), and this association may be required for efficient fast synaptic transmission. Protein kinase C (PKC) and calcium and calmodulin- dependent protein kinase type II (CaM KII) phosphorylated a recombinant his- tagged synprint site polypeptide rapidly to a stoichiometry of 3-4 mol of phosphate/mol, whereas cAMP-dependent protein kinase (PKA) and cGMP-dependent protein kinase (PKG) phosphorylated the synprint peptide more slowly to a stoichiometry of <1 mol/mol. Two-dimensional phosphopeptide mapping revealed similar patterns of phosphorylation of synprint polypeptides and native rat brain N-type calcium channel α(1B) subunits by PKC and Cam KII. Phosphorylation of the synprint peptide with PKC or CaM KII, but not PKA or PKG, strongly inhibited binding of recombinant syntaxin or SNAP-25, even at a level of free calcium (15 μM) that stimulates maximal binding. In contrast, phosphorylation of syntaxin and SNAP-25 with PKC and CaM KII did not affect interactions with the synprint site. Binding assays with polypeptides representing the N- and C-terminal halves of the synprint site indicate that the PKC- and CaM KII-mediated inhibition of binding involves multiple, disperse phosphorylation sites. PKC or CaM KII phosphorylation of the synprint peptide also inhibited its interactions with native rat brain SNARE complexes containing syntaxin and SNAP-25. These results suggest that phosphorylation of the synprint site by PKC or CaM KII may serve as a biochemical switch for interactions between N-type calcium channels and SNARE protein complexes.

Original languageEnglish
Pages (from-to)6929-6938
Number of pages10
JournalJournal of Neuroscience
Issue number18
Publication statusPublished - 1997
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • General Neuroscience


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