Human ethanol-inducible P450IIE1: complete gene sequence, promoter characterization, chromosome mapping, and cDNA-directed expression.
Human ethanol-inducible P450IIE1: complete gene sequence, promoter characterization, chromosome mapping, and cDNA-directed expression.
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DOI：
10.1021/bi00425a019
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发表时间：
1988-12
期刊：
影响因子：
2.9
通讯作者：
M. Umeno;O. McBride;Chung S. Yang;H. Gelboin;F. J. Gonzalez
中科院分区：
文献类型：
--
作者：
M. Umeno;O. McBride;Chung S. Yang;H. Gelboin;F. J. Gonzalez
从λ EMBL 3基因组文库中分离出编码乙醇诱导的亚硝胺代谢P-450的人P450 IIE 1基因，并进行了全序列测定。人类基因跨越11，413个碱基对，包含9个外显子和一个典型的TATA盒。还对2788和559碱基对的上游和下游DNA进行了测序，并与大鼠基因进行了比较。在大鼠和人类基因转录起始位点上游140个碱基对内观察到序列相似性的显著区域。将转录起始位点上游539个碱基对的人DNA插入表达载体pSVOAL δ 5 ′，当将构建体导入大鼠肝癌细胞系时检测到荧光素酶活性。活性比劳斯肉瘤病毒LTR驱动的荧光素酶基因pRSVL低100倍以上。通过使用啮齿动物-人细胞杂交组合，将该基因定位于10号染色体（CYP 2 E位点）。用基因组克隆的第一外显子和部分cDNA克隆构建的全长cDNA在COS细胞中表达，发现其编码N-亚硝基二甲胺脱甲基酶活性。
The human P450IIE1 gene, coding for an ethanol-inducible nitrosamine-metabolizing P-450, was isolated from a lambda EMBL3 genomic library and completely sequenced. The human gene spanned 11,413 base pairs and contained nine exons and a typical TATA box. Upstream and downstream DNAs of 2788 and 559 base pairs were also sequenced and compared to the rat gene. Significant areas of sequence similarity were observed within 140 base pairs upstream of the transcription start site in the rat and human genes. Human DNA 539 base pairs upstream of the transcription start site was inserted into the expression vector pSVOAL delta 5', and luciferase activity was detected when the constructs were introduced into a rat hepatoma cell line. The activity was over 100-fold lower than that of pRSVL, a Rous sarcoma virus LTR-driven luciferase gene. By use of panels of rodent-human cell hybrids, the gene was mapped to chromosome 10 (CYP2E locus). A full-length cDNA, constructed with the first exon of the genomic clone and a partial cDNA clone, was expressed in COS cells and found to code for N-nitrosodimethylamine demethylase activity.