大鼠有机阳离子转运体2（rOCT2）第475位氨基酸的突变会损害有机阳离子转运。
Mutation of amino acid 475 of rat organic cation transporter 2 (rOCT2) impairs organic cation transport.
作者信息
Bahn Andrew, Hagos Yohannes, Rudolph Tanja, Burckhardt Gerhard
机构信息
Zentrum für Physiologie und Pathophysiologie, Abt Vegetative Physiologie und Pathophysiologie, Universität Göttingen, Humboldtallee 23, 37073 Göttingen, Germany.
出版信息
Biochimie. 2004 Feb;86(2):133-6. doi: 10.1016/j.biochi.2003.12.001.
Protein sequence alignments revealed one amino acid position, where organic cation transporters (OCTs, aspartate (D) at position 475 of rOCT2) and organic anion transporters (OATs, arginine (R) at position 466 of rOAT1) are charged oppositely. To address the impact of this amino acid for protein function we cloned rat organic cation transporter 2 (rOCT2), the renal electrogenic cation transporter of the basolateral side of proximal tubule cells. Site-directed mutagenesis was used to generate rOCT2-D475R (rOCT2-mut). Heterologous expression of rOCT2 wild-type (rOCT2-wt) in A6 cells resulted in a significant uptake of the fluorescent organic cation 4-(4-dimethylaminostyryl)-N-methylpyridinium (ASP(+)). Accordingly, rOCT2-wt-transfected COS 7 cells showed an almost fourfold uptake of 25 microM [(14)C]-TEA, whereas rOCT2-mut did not exhibit any uptake of [(14)C]-TEA. These data indicate that rOCT2 transports both ASP(+) and TEA and that aspartate at position 475 of rOCT2 plays a critical role in transport function.
蛋白质序列比对显示出一个氨基酸位置，在此位置上，有机阳离子转运体（OCTs，大鼠OCT2第475位的天冬氨酸（D））和有机阴离子转运体（OATs，大鼠OAT1第466位的精氨酸（R））带有相反电荷。为了研究该氨基酸对蛋白质功能的影响，我们克隆了大鼠有机阳离子转运体2（rOCT2），它是近端小管细胞基底外侧的肾生电阳离子转运体。采用定点诱变技术构建了rOCT2-D475R（rOCT2-突变体）。rOCT2野生型（rOCT2-wt）在A6细胞中的异源表达导致荧光有机阳离子4-(4-二甲基氨基苯乙烯基)-N-甲基吡啶鎓（ASP(+)）的显著摄取。相应地，转染rOCT2-wt的COS 7细胞对25微摩尔[(14)C]-TEA的摄取几乎增加了四倍，而rOCT2-突变体对[(14)C]-TEA没有任何摄取。这些数据表明，rOCT2既能转运ASP(+)也能转运TEA，并且rOCT2第475位的天冬氨酸在转运功能中起关键作用。
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