Isolation and Characterization of P450 Gene from the Formosan Subterranean Termite, Coptotermes formosanus (Isoptera: Rhinotermitidae)

Jianchu Mo,Qinggang Xu,Yue Zhou,Keping Chen

doi:10.13102/sociobiology.v59i4.493

Abstract

A cytochrome P450 gene belonging to family9 was isolated from the midgut transcriptome of the termite Coptotermes formosanus Shiraki, for screening enzymes related to biomass degeneration. Some studies show that insect P450 enzymes have ligninase activities for catalyzing lignin degradation. We employed the RACE method to clone this cytochrome P450 gene, named CYP9AX1 (GenBank accession No.JN969113). To the best of our knowledge, CYP9AX1 is the first member of the CYP9 family cloned from this termite. The full-length CYP9AX1 cDNA was 2242 bp long and included a 1599bp open-reading-frame (ORF), a 61-bp 5’-untranslated region (UTR) and a 592-bp 3’-UTR (excluding the poly-A tail). The CYP9AX1 protein deduced from the ORF contains 532 amino acids with a predicted signal peptide composed of 20 amino acid at its N-terminal and the classic heme-binding domain FXXGXXXCXG (residues 468-477). At position 473, residue Arg (R) changes to Gln (Q), this suggests that CYP9AX1 is a new type of CYP subfamily 9A. The phylogenetic tree showed that C. formosanus has high genetic relationship with Blattella germanica and Diploptera punctata. Quantitative RT-PCR assays demonstrated that CYP9AX1 was expressed most abundantly in malpighian tubules, and slightly lower in the head, foregut, midgut and hindgut. The results suggested that CYP9AX1 may be involved in enzymatic detoxification systems of the delignification process in C. formosanus.

Highlights

The cytochrome P450 (CYP) monooxygenases have numerous functional roles in oxidative transformation of endogenous substrates and xenobiotics (Nelson et al 1996)
The comparative analysis of the C. formosanus P450 gene (CfP450) protein sequence with other insect CYP genes showed that CfP450 has the highest homology to the cytochrome P450 family 9 (CYP9) family
The members of cytochrome P450 family 9 (CYP9) were well known to be related in the oxidative metabolism of insecticides (Feyereisen 1999)

Summary

Introduction

The cytochrome P450 (CYP) monooxygenases have numerous functional roles in oxidative transformation of endogenous substrates and xenobiotics (Nelson et al 1996). Many studies showed that CYP9 genes such as CYP9A2,4,5, CYP9M10, CYP9A12,17, CYP9Q, are mainly involved in detoxification of plant allelochemicals and pesticides (Hardstone et al 2010; Itokawa et al 2010; Mao et al 2011; Stevens et al 2000; Zhou et al 2010). Some insect P450 enzymes show ligninase activities for catalyzing lignin degradation (Geib et al 2008; Scharf & Boucias 2010). The termite cytochrome P450 enzyme may be involved in lignin degradation. It is a candidate pre-treatment enzyme for delignification of lignocelluloses (Scharf & Boucias 2010). Revealing the nature and roles of the P450 monooxoygenases of C. formosanus will facilitate the understanding of the mechanism of this enzyme in lignocellulose pre-treatment

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Conclusion