Yang Y H, Wang J L, Miranda C L, Buhler D R
Genetics Program, Oregon State University, Corvallis 97331, USA.
Arch Biochem Biophys. 1998 Apr 15;352(2):271-80. doi: 10.1006/abbi.1998.0607.
A cDNA clone was isolated from a female rainbow trout liver lambda g tau 11 library using polyclonal antibodies raised against rainbow trout cytochrome P450 LMC1. This 2149-nucleotide clone contained an open reading frame encoding a protein of 499 amino acids with a calculated M(r) of 56,850 Da. On the basis of cytochrome P450 (P450) amino acid sequence comparisons, this rainbow trout P450 was assigned by the P450 Nomenclature Committee to a new P450 subfamily designated as CYP2M1. Northern blot results suggest that the expression of CYP2M1 at the transcriptional level was generally sex, tissue, and age specific. By use of a full-length CYP2M1 cDNA probe, it was observed that this cDNA hybridized strongly to a single 2.2-kb transcript in juvenile female rainbow trout trunk kidney and in liver from juvenile and sexually mature trout from both sexes. Negligible amounts of mRNA hybridizable to CYP2M1 cDNA were found in the juvenile and sexually mature male trunk kidney. cDNA-directed expression in COS-7 cells and of recombinant baculovirus in insect cells produced a protein that was reactive with rabbit anti-trout P450 LMC1 polyclonal antibody and exhibited the unique (omega-6)-hydroxylation toward lauric acid previously observed with rainbow trout P450 LMC1.
利用针对虹鳟鱼细胞色素P450 LMC1产生的多克隆抗体，从雌性虹鳟鱼肝λgt11文库中分离出一个cDNA克隆。这个2149个核苷酸的克隆包含一个开放阅读框，编码一个由499个氨基酸组成的蛋白质，计算分子量为56,850道尔顿。根据细胞色素P450（P450）氨基酸序列比较，这个虹鳟鱼P450被P450命名委员会指定为一个新的P450亚家族，命名为CYP2M1。Northern印迹结果表明，CYP2M1在转录水平的表达通常具有性别、组织和年龄特异性。使用全长CYP2M1 cDNA探针观察到，该cDNA与幼年雌性虹鳟鱼躯干肾以及幼年和性成熟虹鳟鱼（雌雄均有）肝脏中的一个单一2.2 kb转录本强烈杂交。在幼年和性成熟雄性躯干肾中发现与CYP2M1 cDNA可杂交的mRNA量极少。在COS-7细胞中进行cDNA指导表达以及在昆虫细胞中表达重组杆状病毒，产生了一种与兔抗虹鳟鱼P450 LMC1多克隆抗体反应的蛋白质，并表现出先前在虹鳟鱼P450 LMC1中观察到的对月桂酸独特的（ω-6）羟基化作用。
