Direct interaction of the calcium sensor protein synaptotagmin I with a cytoplasmic domain of the alpha(1)A subunit of the P/Q-type calcium channel

被引:104
作者
Charvin, N
Leveque, C
Walker, D
Berton, F
Raymond, C
Kataoka, M
ShojiKasai, Y
Takahashi, M
DeWaard, M
Seagar, MJ
机构
[1] FAC MED SECTEUR NORD MARSEILLE,INST JEAN ROCHE,INSERM,UNITE 464,F-13916 MARSEILLE 20,FRANCE
[2] MITSUBISHI KASEI INST LIFE SCI,MACHIDA,TOKYO 194,JAPAN
关键词
alpha(1A) subunit; calcium channel; omega-conotoxin MVIIC; synaptotagmins;
D O I
10.1093/emboj/16.15.4591
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Synaptotagmins are synaptic vesicle proteins containing two calcium-binding C2 domains which are involved in coupling calcium influx through voltage-gated channels to vesicle fusion and exocytosis of neurotransmitters. The interaction of synaptotagmins with native P/Q-type calcium channels was studied in solubilized synaptosomes from rat cerebellum, Antibodies against synaptotagmins I and II, but not TV co-immunoprecipitated [I-125]omega-conotoxin MVIIC-labelled calcium channels, Direct interactions were studied between in vitro-translated [S-35]synaptotagmin I and fusion proteins containing cytoplasmic loops of the alpha(1)A subunit (BI isoform), Gel overlay revealed the association of synaptotagmin I with a single region (residues 780-969) located in the intracellular loop connecting homologous domains II and III, Saturable calcium-independent binding occurred with equilibrium dissociation constants of 70 nM and 340 nM at 4 degrees C and pH 7.4, and association was blocked by addition of excess recombinant synaptotagmin I, Direct synaptotagmin binding to the pore-forming subunit of the P/Q-type channel may optimally locate the calcium-binding sites that initiate exocytosis within a zone of voltage-gated calcium entry.
引用
收藏
页码:4591 / 4596
页数:6
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