Lin C G, Lin Y C, Liu H W, Kao L S
Institute of Biomedical Sciences, Academia Sinica, Taipei 115, Taiwan, Republic of China.
Biochem J. 1997 May 15;324 ( Pt 1)(Pt 1):85-90. doi: 10.1042/bj3240085.
In this study we examined the biochemical properties and subcellular localization of Rab3A, Rab3B and Rab3C in bovine adrenal chromaffin cells. The Kd for guanosine 5'-[gamma-thio]triphosphate (GTP[S]) of the three Rab3 proteins was 15, 2700 and 204 nM for Rab3A, Rab3B and Rab3C respectively. The intrinsic GTPase activity of the three Rab3 proteins seemed similar and was increased approx. 3-fold by bovine chromaffin cell lysate. Truncation of the C-terminal 31 amino acid residues decreased the binding affinity for GTP[S] of the three Rab3 proteins. When the C-terminus of Rab3C was replaced with that of Rab3A, the binding affinity of Rab3C for GTP[S] was decreased, but the replacement did not affect the affinity of Rab3B for GTP[S]. Immunostaining experiments showed that Rab3A, Rab3B and Rab3C are localized separately within chromaffin cells. Anti-Rab3A and anti-Rab3C antibodies stained vesicle-like structures, whereas anti-Rab3B antibody distinctly stained the plasma membrane. In summary, bovine chromaffin cells express the three Rab3 proteins but the subcellular localization and biochemical properties of the three Rab3 proteins are distinct.
在本研究中，我们检测了牛肾上腺嗜铬细胞中Rab3A、Rab3B和Rab3C的生化特性及亚细胞定位。三种Rab3蛋白对鸟苷5'-[γ-硫代]三磷酸（GTP[S]）的解离常数（Kd），Rab3A为15 nM，Rab3B为2700 nM，Rab3C为204 nM。三种Rab3蛋白的内在GTP酶活性似乎相似，且经牛嗜铬细胞裂解液处理后活性增加约3倍。C末端31个氨基酸残基的截短降低了三种Rab3蛋白对GTP[S]的结合亲和力。当Rab3C的C末端被Rab3A的C末端取代时，Rab3C对GTP[S]的结合亲和力降低，但该取代不影响Rab3B对GTP[S]的亲和力。免疫染色实验表明，Rab3A、Rab3B和Rab3C在嗜铬细胞内分别定位。抗Rab3A和抗Rab3C抗体染色呈囊泡样结构，而抗Rab3B抗体则明显染色质膜。总之，牛嗜铬细胞表达三种Rab3蛋白，但三种Rab3蛋白的亚细胞定位和生化特性各不相同。
