豚鼠CYP1A2 cDNA的分子克隆及其与豚鼠CYP1A1的比较

Molecular cloning of cDNA for guinea pig CYP1A2 comparison with guinea pig CYP1A1.

作者信息

Black V H, Wang A, Henry M, Shaw P

机构信息

Department of Cell Biology and Kaplan Cancer Center, New York University School of Medicine, New York 10016, USA.

出版信息

Arch Biochem Biophys. 1997 Aug 1;344(1):11-7. doi: 10.1006/abbi.1997.0189.

DOI:10.1006/abbi.1997.0189

PMID:9244376

Abstract

Guinea pig CYP1A2 cDNA was isolated by RT-PCR from liver tissue of 3,3'-methylcholanthrene-treated guinea pigs. It shares considerable sequence identity with guinea pig CYP1A1 (nt 77%, aa 65%), but differs in levels of constitutive expression, function, and inducibility. Western blot analysis of protein expressed by full-length cDNA in COS-1 cells identified CYP1A2 (56 kDa) and CYP1A1 (53 kDa) proteins in corun liver microsomes. CYP1A2 transfectants metabolized methoxyresorufin and ethoxyresorufin, while CYP1A1 transfectants metabolized only ethoxyresorufin. Constitutive expression of CYP1A2 mRNA (2.0 kb) and protein was much lower than that of CYP1A1 mRNA (2.6 kb) and protein, but the fold induction of CYP1A2 by 3,3'-methylcholanthrene was greater than that of CYP1A1. Changes in splicing of CYP1A2 pre-mRNA occur upon treatment with 3,3'-methylcholanthrene.

摘要

通过逆转录聚合酶链反应（RT-PCR）从经3,3'-甲基胆蒽处理的豚鼠肝脏组织中分离出豚鼠细胞色素P450 1A2（CYP1A2）cDNA。它与豚鼠细胞色素P450 1A1（CYP1A1）具有相当高的序列同一性（核苷酸水平为77%，氨基酸水平为65%），但在组成型表达水平、功能和诱导性方面存在差异。对COS-1细胞中全长cDNA所表达的蛋白质进行蛋白质印迹分析，在豚鼠肝脏微粒体中鉴定出CYP1A2（56 kDa）和CYP1A1（53 kDa）蛋白质。CYP1A2转染细胞代谢甲氧基试卤灵和乙氧基试卤灵，而CYP1A1转染细胞仅代谢乙氧基试卤灵。CYP1A2 mRNA（2.0 kb）和蛋白质的组成型表达远低于CYP1A1 mRNA（2.6 kb）和蛋白质，但3,3'-甲基胆蒽对CYP1A2的诱导倍数大于对CYP1A1的诱导倍数。用3,3'-甲基胆蒽处理后，CYP1A2前体mRNA的剪接发生变化。