Cloning, sequencing and expression of the major rainbow trout constitutive cytochrome P450 (CYP2K1). Identification of a new cytochrome P450 gene subfamily

Abstract

A full-length cDNA has been cloned and sequenced for the major constitutive rainbow trout liver cytochrome P450 that had earlier been designated as cytochrome P450 LMC2. This cDNA clone encodes for a protein of 504 amino acids with a calculated M r of 56,795. While the cDNA nucleotides for trout P450 LMC2 shared a 49 and 47% homology, respectively, with the two rainbow trout P450s previously sequenced (hepatic P4501A1 and an ovarian 17-hydroxylase), it showed a 54–56% nucleotide identity with several human or other mammalian P450 forms from family 2. From amino acid sequence comparisons, the trout P450 LMC2 has been assigned to a new cytochrome P450 gene family designated P4502K1 (CYP2K1). Northern blots of total RNA isolated from sexually mature male and female rainbow trout examined with a cDNA probe (2K1,7c), indicated that mRNA levels were substantially higher in the trunk kidney of male trout compared to that seen in females. Multiple 2K1,7c hybridizable bands at about 2.8 kb and 1.6–1.9 kb were found in the trunk kidney preparations, with the 2.8 kb band predominating. A major 1.9 kb mRNA band, detectable with the 2K1,7c probe, was present at similar concentrations in livers of both sexes.