CYP2M1: Cloning, Sequencing, and Expression of a New Cytochrome P450 from Rainbow Trout Liver with Fatty Acid (ω-6)-Hydroxylation Activity

Abstract

A cDNA clone was isolated from a female rainbow trout liver λgτ11 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 calculatedMrof 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 (ω-6)-hydroxylation toward lauric acid previously observed with rainbow trout P450 LMC1.