MOLECULAR AND FUNCTIONAL-CHARACTERIZATION OF AN ORGANIC ANION TRANSPORTING POLYPEPTIDE CLONED FROM HUMAN LIVER
MOLECULAR AND FUNCTIONAL-CHARACTERIZATION OF AN ORGANIC ANION TRANSPORTING POLYPEPTIDE CLONED FROM HUMAN LIVER
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DOI：
10.1016/0016-5085(95)90588-x
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发表时间：
1995-10-01
期刊：
影响因子：
29.4
通讯作者：
MEIER, PJ
中科院分区：
文献类型：
--
作者：
KULLAKUBLICK, GA;HAGENBUCH, B;MEIER, PJ
背景和目标：在最近克隆的大鼠肝有机阴离子转运蛋白的基础上，我们尝试克隆相应的人肝有机阴离子转运多肽。方法：用特异性大鼠肝互补DNA探针筛选人肝互补DNA文库。通过与大鼠蛋白质的同源性克隆了人肝转运蛋白，并在非洲爪蟾卵母细胞中进行了功能表征。结果如下：克隆的人肝有机阴离子转运多肽由670个氨基酸组成，与相应的大鼠肝蛋白的氨基酸同源性为67%。将体外转录的互补RNA注入蛙卵母细胞导致[S-35]溴磺酞非钠依赖性摄取的表达（米氏常数[K-m]，类似于20 μ mol/L），[H-3]胆酸盐（K-m，类似于93 μ mol/L），[H-3]-牛磺胆酸盐（K-m，类似于60 μ mol/L）、[C-14]甘氨胆酸盐、[H-3]牛磺鹅脱氧胆酸盐和[H-3]牛磺熊脱氧胆酸盐（K-m类似于19 μ mol/L）。北方印迹分析显示与来自人肝、脑、肺、肾和睾丸的信使RNA种类的交叉反应性。对一组人-啮齿动物体细胞杂交体的基因组DNA进行聚合酶链反应分析，将克隆的人有机阴离子转运蛋白定位于12号染色体。结论：这些研究表明，克隆的人肝有机阴离子转运蛋白与先前克隆的大鼠肝转运蛋白密切相关，但可能不相同。此外，其在各种肝外组织中的额外定位表明其在人体的整体经上皮有机阴离子转运中起着重要作用。
Background & Aims: Based on a recently cloned rat liver organic anion transporter, we attempted to clone the corresponding human liver organic anion transporting polypeptide. Methods: A human liver complementary DNA library was screened with a specific rat liver complementary DNA probe. The human liver transporter was cloned by homology with the rat protein and functionally characterized in Xenopus laevis oocytes. Results: The cloned human liver organic anion transporting polypeptide consists of 670 amino acids and shows a 67% amino acid identity with the corresponding rat liver protein. Injection of in vitro transcribed complementary RNA into frog oocytes resulted in the expression of sodium-independent uptake of [S-35]bromosulfophthalein (Michaelis constant [K-m], similar to 20 mu mol/L), [H-3]cholate (K-m, similar to 93 mu mol/L), [H-3]- taurocholate (K-m, similar to 60 mu mol/L), [C-14]glycocholate, [H-3]taurochenodeoxycholate, and [H-3]tauroursodeoxycholate (K-m similar to 19 mu mol/L). Northern blot analysis showed cross-reactivity with messenger RNA species from human liver, brain, lung, kidney, and testes. Polymerase chain reaction analysis of genomic DNA from a panel of human-rodent somatic cell hybrids mapped the cloned human organic anion transporter to chromosome 12. Conclusions: These studies show that the cloned human liver organic anion transporter is closely related to, but probably not identical to, the previously cloned rat liver transporter. Furthermore, its additional localization in a variety of extrahepatic tissues suggests that it plays a fundamental role in overall transepithelial organic anion transport of the human body.