Isolation and sequence of cDNA encoding a cytochrome P-450 from an insecticide-resistant strain of the house fly, Musca domestica

Abstract

A cDNA expression library from phenobarbital-treated house fly (Musca domestica) was screened with rabbit antisera directed against partially purified house fly cytochrome P-450. Two overlapping clones with insert lengths of 1.3 and 1.5 kilobases were isolated. The sequence of a 1629-base-pair (bp) cDNA was obtained, with an open reading frame (nucleotides 81-1610) encoding a P-450 protein of 509 residues (Mr = 58,738). The insect P-450 protein contains a hydrophobic NH2 terminus and a 22-residue cysteine-containing fragment near the COOH terminus that is known to bind the heme; both of these features have been found in the more than five dozen vertebrate P-450 proteins of which the sequences are presently known. Interestingly, the termination codon UAA may be contained in a putative polyadenylylation signal (AAUAAA) of the mRNA. This P-450 protein exhibits the most similarity (27% amino acid positional identity) with mammalian proteins of the P450III family but qualifies as a member of another family, which we propose to designate the P450VI gene family. This cDNA and deduced protein sequence should provide important information in the study of evolution of the P-450 gene superfamily, as well as provide an important probe for studying the regulation of insect P-450 and understanding the molecular genetics of insecticide resistance.