Human CYP2B6: expression, inducibility and catalytic activities
Human CYP2B6: expression, inducibility and catalytic activities
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DOI：
10.1097/00008571-199906000-00004
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发表时间：
1999-06-01
期刊：
影响因子：
--
通讯作者：
de Waziers, I
中科院分区：
文献类型：
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作者：
Gervot, L;Rochat, B;de Waziers, I
克隆了人细胞色素2B 6 cDNA，并在细菌和酵母中表达。它在酿酒酵母中的表达使我们能够获得高水平的活性酵母表达的CYP 2B 6蛋白，以评估其在乙氧基试卤灵、五氧试卤灵、苄氧试卤灵、乙氧基香豆素、睾酮和环磷酰胺代谢中的作用。动力学分析表明，人CYP 2B 6优先代谢苄氧试卤灵和五氧试卤灵，尽管其他CYP也在人肝微粒体中代谢这些底物。CYP 2B 6也表现出较强的4-羟基环磷酰胺活性。它在大肠杆菌中的表达使我们能够产生一个非常特异性的抗人CYP 2B 6抗体。未观察到该抗体与CYP 1A 1、1A 2、3A 4、3A 5、2C 8、2C 9、2C 18、2C 19、2D 6或2 E1的交叉反应。该抗体使我们能够研究CYP 2B 6在人类中的肝和肝外表达，以及其在原代培养的人肝细胞和不同的人细胞系中的表达和诱导。免疫印迹分析显示，CYP 2B 6蛋白在检测的48份人肝脏样品中的43份中表达，水平范围为0.4 - 8 pmol/mg微粒体蛋白，平均值为1.7 pmol/mg蛋白，CYP 2B也在人脑、肠和肾脏中表达，在肺中表达水平较低。在人肝、肾、肺、气管和肠中检测到CYP 2B mRNA。我们还发现，CYP 2B 6在蛋白质和mRNA水平上被苯巴比妥（2 mM）和环磷酰胺（1 mM）诱导，苯巴比妥是一种已知由CYP 2B 6代谢的抗癌药物。在任何测试的人细胞系中均未观察到CYP 2B 6的表达或诱导，Pharmacogenetics 9：295-306（C）1999 Lippincott威廉姆斯和威尔金斯。
Human cytochrome (CYP)2B6 cDNA was cloned and expressed in bacteria and in yeast. Its expression in Saccharomyces cerevisiae enabled us to obtain, at a high level, an active yeast-expressed CYP2B6 protein, so as to assess its role in the metabolism of ethoxyresorufin, pentoxyresorufin, benzyloxyresorufin, ethoxycoumarin, testosterone and cyclophosphamide, Kinetic analysis showed that human CYP2B6 preferentially metabolized benzyloxyresorufin and pentoxyresorufin, although other CYPs also metabolized these substrates in human liver microsomes. CYP2B6 also manifested a strong 4-hydroxycyclophosphamide activity. Its expression in Escherichia coli enabled us to produce a very specific anti-human CYP2B6 antibody. No cross reactivity of this antibody was observed with CYPs1A1, 1A2, 3A4, 3A5, 2C8, 2C9, 2C18, 2C19, 2D6 or 2E1. This antibody enabled us to study the hepatic and extrahepatic expression of CYP2B6 in man, as well as its expression and inducibility in primary cultured human hepatocytes and in different human cell lines. Immunoblot analysis revealed that the CYP2B6 protein was expressed in 43 of the 48 human liver samples tested, with levels ranging from 0.4 to 8 pmol/mg of microsomal protein with a mean of 1.7 pmol/mg protein, CYP2B was also expressed in human brain, intestine and kidney, and at a lower level in the lung. CYP2B mRNA was detected in human liver, kidney, lung, trachea and intestine, We also found that CYP2B6 is induced at protein and mRNA levels by phenobarbital (2 mM) and cyclophosphamide (1 mM), an anticancer drug known to be metabolized by CYP2B6, No expression or inducibility of CYP2B6 was observed in any of the human cell lines tested, Pharmacogenetics 9:295-306 (C) 1999 Lippincott Williams & Wilkins.