Complementary DNA cloning and organ expression of cytochrome P450 1C2 in carp (Cyprinus carpio).

Yoshio Kaminishi, Mohamed A H El-Kady, Ryoichi Mitsuo, Takao Itakura
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Abstract

Cytochrome P450 (CYP) genes, which make up a large gene superfamily, are known to play an important role in drug metabolism. The CYP1 family, one of the gene families of the CYP superfamily, has three subfamilies of genes whose sequences have been deposited in the GenBank/EMBL thus far: CYP1A, CYP1B, and CYP1C. Mammals as well as fish confront numerous foreign chemicals in the environment that may accumulate to toxic levels unless they are metabolized and eliminated by processes largely mediated by CYP enzymes. A new complementary DNA of the CYP1C subfamily encoding CYP1C2 was isolated from the carp liver after a single intraperitoneal injection of beta-napthoflavone (BNF). The full-length cDNA obtained contained a 5' noncoding region of 198 bp, an open reading frame of 1575 bp coding for 524 amino acids and a stop codon, and a 3' noncoding region of 531 bp. The predicted molecular weight of the protein was approximately 59.3 kDa. The amino acid sequence deduced from the carp CYP1C2 sequence showed a similarity of 76.6% with that deduced from our previously reported carp CYP1C1. It exhibited similarities of 77.3, 73.7, and 76.4% with those deduced from scup CYP1C2, scup CYP1C1, and Japanese eel CYP1C1 sequences, respectively. Carp CYP1C2 cDNA showed similarities with reported CYP1Bs of teleosts and mammals, namely, 47.6, 45.3, 45.7, 44.0, and 44.6% for carp, plaice, human, rat, and mouse CYP1B1s, respectively, while it exhibited a similarity of 49.0% with carp CYP1B2. The carp CYP1C2 sequence was aligned with the CYP1 sequences and has been deposited in the GenBank/EMBL data bank with the accession number AY437777. The phylogenetic tree constructed using fish and mammalian CYP1 sequences suggested a closer relationship of CYP1C with CYP1B than with CYP1A. The tree showed possibile existence of CYP1C subfamily genes in mammalian species. Northern blot analysis of the liver, intestines, kidneys, and gills revealed a distinct induced expression only in the kidneys, with no detectable constitutive expression in the other organs studied.

鲤鱼细胞色素P450 - 1C2的互补DNA克隆及器官表达。
细胞色素P450 (CYP)基因是一个庞大的基因超家族,在药物代谢中起重要作用。CYP1家族是cypp超家族的基因家族之一,有三个亚家族的基因,其序列迄今已存放在GenBank/EMBL中:CYP1A、CYP1B和CYP1C。哺乳动物和鱼类都要面对环境中大量的外来化学物质,这些化学物质可能积累到有毒水平,除非它们被主要由CYP酶介导的过程代谢和消除。经单次腹腔注射β -napthoflavone (BNF)后,从鲤鱼肝脏中分离出一个新的CYP1C亚家族编码CYP1C2的互补DNA。获得的全长cDNA包含一个198 bp的5'非编码区,一个1575 bp的开放阅读框,编码524个氨基酸和一个停止密码子,以及一个531 bp的3'非编码区。该蛋白的预测分子量约为59.3 kDa。从鲤鱼CYP1C2序列推断出的氨基酸序列与我们先前报道的鲤鱼CYP1C1序列的相似性为76.6%。其与scup CYP1C2、scup CYP1C1和日本鳗鲡CYP1C1序列的相似性分别为77.3、73.7和76.4%。鲤鱼CYP1C2 cDNA与已报道的鱼类和哺乳动物cyp1b1相似度分别为47.6%、45.3%、45.7%、44.0%和44.6%,与鲤鱼CYP1B2相似度为49.0%。鲤鱼CYP1C2序列与CYP1序列一致,已存入GenBank/EMBL数据库,登录号为AY437777。利用鱼类和哺乳动物CYP1序列构建的系统发育树表明,CYP1C与CYP1B的关系比与CYP1A的关系更密切。该树显示哺乳动物中可能存在CYP1C亚家族基因。对肝脏、肠道、肾脏和鳃的Northern blot分析显示,仅在肾脏中有明显的诱导表达,在其他器官中没有检测到组成性表达。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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