title: Cloning, functional characterization, and expression profiles of NADPH-cytochrome P450 reductase gene from the Asiatic rice striped stem borer, Chilo suppressalis (Lepidoptera: Pyralidae).
abstract: NADPH-cytochrome P450 reductase (CPR) is one of the most important components of the cytochrome P450 enzyme system. It catalyzes electron transfer from NADPH to all known P450s, thus plays central roles not only in the metabolism of exogenous xenobiotics but also in the regulation of endogenous hormones in insects. In this study, a full-length cDNA encoding of a CPR (named CsCPR) was isolated from the Asiatic rice striped stem borer, Chilo suppressalis, by using reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) methods. The cDNA contains a 2061 bp open reading frame, which encodes an enzyme of 686 amino acid residues, with a calculated molecular mass of 77.6 kDa. The deduced peptide has hallmarks of typical CPR, including an N-terminal membrane anchor and the FMN, FAD and NADPH binding domains. The N-terminal-truncated protein fused with a 6 × His·tag was heterologously expressed in Escherichia coli Rosetta (DE3) cells and purified, specific activity and the Km values of the recombinant enzyme were determined. Tissue- and developmental stage-dependent expression of CsCPR mRNA was investigated by real-time quantitative PCR. The CsCPR mRNA was noticeably expressed in the digestive, metabolic, and olfactory organs of the larvae and adults of C. suppressalis. Our initial results would provide valuable information for further study on the interactions between CPR and cytochrome P450 enzyme systems.
authors.affiliation: State Key Laboratory of Rice Biology, Key Laboratory of Agricultural Entomology, Ministry of Agriculture, Institute of Insect Sciences, Zhejiang University, Hangzhou, China.
authors.forename: Su
authors.initials: S
authors.lastname: Liu
authors.forename: Qing-Mei
authors.initials: QM
authors.lastname: Liang
authors.forename: Yuan-Jie
authors.initials: YJ
authors.lastname: Huang
authors.forename: Xin
authors.initials: X
authors.lastname: Yuan
authors.forename: Wen-Wu
authors.initials: WW
authors.lastname: Zhou
authors.forename: Fei
authors.initials: F
authors.lastname: Qiao
authors.forename: Jiaan
authors.initials: J
authors.lastname: Cheng
authors.forename: Geoff M
authors.initials: GM
authors.lastname: Gurr
authors.forename: Zeng-Rong
authors.initials: ZR
authors.lastname: Zhu
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D007915:Lepidoptera / Q000201:enzymology* / Q000235:genetics* / Q000254:growth & development; D008969:Molecular Sequence Data; D009251:NADPH-Ferrihemoprotein Reductase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D009928:Organ Specificity; D010802:Phylogeny; D017421:Sequence Analysis
chemical_list: D009251:NADPH-Ferrihemoprotein Reductase
keywords: Chilo suppressalis; Cytochrome c; NADPH; P450 reductase; Relative mRNA level