Barbier O, Bélanger A, Hum D W
Laboratory of Molecular Endocrinology, CHUL Research Center, Laval University, 2705 Laurier Boulevard, Quebec, Canada G1V 4G2.
Biochem J. 1999 Feb 1;337 ( Pt 3)(Pt 3):567-74.
Steroid glucuronidation by UDP-glucuronosyltransferase (UGT) enzymes is a mechanism leading to catabolism and elimination of steroid hormones. To establish an animal model to investigate the conjugation of steroids by UGT enzymes, previous results revealed that simian and human are unique in having high levels of circulating androsterone glucuronide and androstane-3alpha, 17beta-diol (3d-Diol) glucuronide. A cDNA, UGT2B20, was isolated from cynomolgus monkey liver and prostate libraries. The cDNA was 2075 bp in length and contained an open reading frame of 1590 bp, encoding a protein of 530 amino acid residues. The UGT2B20 clone was transfected and stably expressed in the human embryo kidney HK293 cell line, and the transferase activity of UGT2B20 was tested with 73 compounds. This enzyme was shown to be active with androgens, such as testosterone, dihydrotestosterone (DHT) and 3alpha-Diol, and on catecholoestrogens including 1,3,5,10-oestratriene-3, 4-diol-17-one. Kinetic analysis performed with intact cells yielded apparent Km values of 1.1, 2.3 and 4.6 microM for 3alpha-Diol, DHT and testosterone respectively. Reverse transcriptase-PCR analysis demonstrated that UGT2B20 transcript is expressed in several tissues including the liver, prostate, kidney, epididymis and adrenal of the cynomolgus monkey. Amino acid sequence alignment shows that the UGT2B20 protein is 92% identical with UGT2B15. Both enzymes have similar apparent Km values for DHT and 3alpha-Diol, and demonstrate similar transcript tissue distribution. The characterization of simian UGT2B20 as a structural and functional homologue of human UGT2B15 further demonstrates the similarities of steroid glucuronidation in these two species, and indicates the relevance of using the monkey as an animal model to study and understand steroid glucuronidation in extrahepatic-steroid target tissues.
UDP-葡萄糖醛酸基转移酶（UGT）催化的类固醇葡萄糖醛酸化是类固醇激素分解代谢和消除的一种机制。为建立一个用于研究UGT酶对类固醇进行结合作用的动物模型，先前的研究结果表明，猿猴和人类在循环中雄酮葡萄糖醛酸苷和雄甾烷-3α,17β-二醇（3d-二醇）葡萄糖醛酸苷水平较高方面具有独特性。从食蟹猴肝脏和前列腺文库中分离出一个cDNA，即UGT2B20。该cDNA长度为2075 bp，包含一个1590 bp的开放阅读框，编码一个由530个氨基酸残基组成的蛋白质。将UGT2B20克隆转染并稳定表达于人胚肾HK293细胞系中，并用73种化合物测试UGT2B20的转移酶活性。结果显示该酶对雄激素如睾酮、双氢睾酮（DHT）和3α-二醇有活性，对儿茶酚雌激素包括1,3,5,10-雌三烯-3,4-二醇-17-酮也有活性。对完整细胞进行的动力学分析得出，3α-二醇、DHT和睾酮的表观Km值分别为1.1、2.3和4.6 microM。逆转录酶-PCR分析表明，UGT2B20转录本在食蟹猴的多个组织中表达，包括肝脏、前列腺、肾脏、附睾和肾上腺。氨基酸序列比对显示，UGT2B20蛋白与UGT2B15的同源性为92%。两种酶对DHT和3α-二醇的表观Km值相似，且转录本组织分布也相似。将猿猴UGT2B20鉴定为人类UGT2B15的结构和功能同源物，进一步证明了这两个物种在类固醇葡萄糖醛酸化方面的相似性，并表明使用猴子作为动物模型来研究和理解肝外类固醇靶组织中的类固醇葡萄糖醛酸化具有相关性。
