Rabconnectin-3, a novel protein that binds both GDP/GTP exchange protein and GTPase-activating protein for Rab3 small G protein family
Rabconnectin-3, a novel protein that binds both GDP/GTP exchange protein and GTPase-activating protein for Rab3 small G protein family
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DOI：
10.1074/jbc.c100730200
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发表时间：
2002-03-22
影响因子：
4.8
通讯作者：
Takai, Y
中科院分区：
文献类型：
--
作者：
Nagano, F;Kawabe, H;Takai, Y
Rab3A是Rab3小G蛋白家族的一员，调节神经递质的钙依赖性胞吐。Rab3A的周期性激活和失活是Rab3A在胞吐作用中所必需的。GDP-Rab3A被Rab3 GDP/GTP交换蛋白(Rab3 GEP)激活为GTP-Rab3A，GTP-Rab3A被Rab3 GTP酶激活蛋白(Rab3 GAP)灭活为GDP-Rab3A。目前尚不清楚这些调节因子是如何或在多重胞吐作用的哪个步骤中被激活和失活的。我们从大鼠脑突触泡粗提物中分离到一种与Rab3、GEP和GAP抗体免疫共沉淀的新蛋白。该蛋白命名为RabConnectin-3，可与Rab3 GEP和GAP结合。从人的cDNA文库中克隆了RabConnectin-3的基因，并对其一级结构进行了测定。人RabConnectin-3由3,036个氨基酸组成，计算的M-r为339,753。它有12个WD域名。大鼠的组织和亚细胞分布分析表明，RabConnectin-3在脑内大量表达，并在突触小泡部分丰富表达。免疫荧光和免疫电子显微镜显示RabConnectin-3主要集中在突触的突触小泡上。这些结果表明，RabConnectin-3是突触小泡上Rab3GEP和GAP的支架分子。
Rab3A, a member of the Rab3 small G protein family, regulates Ca2+-dependent exocytosis of neurotransmitter. The cyclical activation and inactivation of Rab3A are essential for the Rab3A action in exocytosis. GDP-Rab3A is activated to GTP-Rab3A by Rab3 GDP/GTP exchange protein (Rab3 GEP), and GTP-Rab3A is inactivated to GDP-Rab3A by Rab3 GTPase-activating protein (Rab3 GAP). It remains unknown how or in which step of the multiple exocytosis steps these regulators are activated and inactivated. We isolated here a novel protein that was co-immunoprecipitated with Rab3 GEP and GAP by their respective antibodies from the crude synaptic vesicle fraction of rat brain. The protein, named rabconnectin-3, bound both Rab3 GEP and GAP. The cDNA of rabconnectin-3 was cloned from a human cDNA library and its primary structure was determined. Human rabconnectin-3 consisted of 3,036 amino acids and showed a calculated M-r of 339,753. It had 12 WD domains. Tissue and subcellular distribution analyses in rat indicated that rabconnectin-3 was abundantly expressed in the brain where it was enriched in the synaptic vesicle fraction. Immunofluorescence and immunoelectron microscopy revealed that rabconnectin-3 was concentrated on synaptic vesicles at synapses. These results indicate that rabconnectin-3 serves as a scaffold molecule for both Rab3 GEP and GAP on synaptic vesicles.