CYP341B14: A cytochrome P450 involved in the specific epoxidation of pheromone precursors in the fall webworm Hyphantria cunea

• Female moths of the fall webworm utilize the epoxides of polyunsaturated hydrocarbons as sex pheromone components.
• We attempted to identify the enzyme involved in the epoxidation of pheromone precursors in the pheromone gland.
• A gene encoding cytochrome P450 (Hc_epo1), which was specifically expressed in the pheromone gland, was cloned.
• Hc_epo1 was determined to belong to a CYP341B clade in the CYP341 family and was assigned the name CYP341B14.
• Hc_epo1 specifically epoxidized the (Z)-double bond at the 9th position in the pheromone precursor.

Two of the four sex pheromone components in the fall webworm Hyphantria cunea (Lepidoptera: Arctiidae), cis-9,10-epoxy-(3Z,6Z)-3,6-henicosadiene and cis-9,10-epoxy-(3Z,6Z)-1,3,6-henicosatriene, possess an epoxy ring within their molecules. These compounds have been suggested to be biosynthesized from dietary linolenic acid via the following enzymatic reactions; chain elongation, terminal desaturation (in the case of the latter component), decarboxylation, and epoxidation. The last step of this biosynthesis, epoxidation, is known to occur specifically in the sex pheromone gland of females. We identified the enzyme involved in the epoxidation of pheromone precursors by focusing on cytochromes P450, which are known to catalyze the oxidation of various compounds. Three P450-like sequences (Hc_epo1, Hc_epo2, and Hc_epo3) were identified in the cDNA library prepared from the sex pheromone gland of H. cunea. Among these clones, only Hc_epo1 was specifically expressed in the pheromone gland. The full-length sequence of Hc_epo1 contained an ORF of 1527 bp, which encoded a protein of 509 amino acids with a predicted molecular weight of 57.9 kDa. The deduced Hc_epo1 amino acid sequence possessed the characteristics of P450. A phylogenetic analysis of the sequence indicated that Hc_epo1 belonged to the CYP341B clade in the CYP341 family. Therefore, it was named CYP341B14. A subsequent functional assay using Sf-9 cells transiently expressing CYP341B14 demonstrated that this P450 protein was able to specifically epoxidize a (Z)-double bond at the 9th position in the pheromone precursor, (3Z,6Z,9Z)-3,6,9-henicosatriene.

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