Zhao Chunqing, Feng Xiaoyun, Tang Tao, Qiu Lihong
College of Science, China Agricultural University, Beijing, China College of Plant Protection, Nanjing Agricultural University, Nanjing, China *These authors contributed equally to this work.
College of Science, China Agricultural University, Beijing, China *These authors contributed equally to this work.
J Insect Sci. 2015 Aug 28;15(1). doi: 10.1093/jisesa/iev100. Print 2015.
Cytochrome P450 monooxygenases (CYPs), as an enzyme superfamily, is widely distributed in organisms and plays a vital function in the metabolism of exogenous and endogenous compounds by interacting with its obligatory redox partner, CYP reductase (CPR). A novel CYP gene (CYP9A11) and CPR gene from the agricultural pest insect Spodoptera exigua were cloned and characterized. The complete cDNA sequences of SeCYP9A11 and SeCPR are 1,931 and 3,919 bp in length, respectively, and contain open reading frames of 1,593 and 2,070 nucleotides, respectively. Analysis of the putative protein sequences indicated that SeCYP9A11 contains a heme-binding domain and the unique characteristic sequence (SRFALCE) of the CYP9 family, in addition to a signal peptide and transmembrane segment at the N-terminal. Alignment analysis revealed that SeCYP9A11 shares the highest sequence similarity with CYP9A13 from Mamestra brassicae, which is 66.54%. The putative protein sequence of SeCPR has all of the classical CPR features, such as an N-terminal membrane anchor; three conserved domain flavin adenine dinucleotide (FAD), flavin mononucleotide (FMN), and nicotinamide adenine dinucleotide phosphate (NADPH) domain; and characteristic binding motifs. Phylogenetic analysis revealed that SeCPR shares the highest identity with HaCPR, which is 95.21%. The SeCYP9A11 and SeCPR genes were detected in the midgut, fat body, and cuticle tissues, and throughout all of the developmental stages of S. exigua. The mRNA levels of SeCYP9A11 and SeCPR decreased remarkably after exposure to plant secondary metabolites quercetin and tannin. The results regarding SeCYP9A11 and SeCPR genes in the current study provide foundation for the further study of S. exigua P450 system.
细胞色素P450单加氧酶（CYPs）作为一个酶超家族，广泛分布于生物体中，并通过与其必需的氧化还原伴侣——CYP还原酶（CPR）相互作用，在外源和内源化合物的代谢中发挥着至关重要的作用。从农业害虫甜菜夜蛾中克隆并鉴定了一个新的CYP基因（CYP9A11）和CPR基因。SeCYP9A11和SeCPR的完整cDNA序列长度分别为1931和3919 bp，分别包含1593和2070个核苷酸的开放阅读框。对推测的蛋白质序列分析表明，SeCYP9A11除了在N端有一个信号肽和跨膜区段外，还包含一个血红素结合结构域和CYP9家族独特的特征序列（SRFALCE）。比对分析显示，SeCYP9A11与甘蓝夜蛾的CYP9A13序列相似性最高，为66.54%。SeCPR的推测蛋白质序列具有所有经典的CPR特征，如N端膜锚定；三个保守结构域：黄素腺嘌呤二核苷酸（FAD）、黄素单核苷酸（FMN）和烟酰胺腺嘌呤二核苷酸磷酸（NADPH）结构域；以及特征性结合基序。系统发育分析表明，SeCPR与HaCPR的一致性最高，为95.21%。在甜菜夜蛾的中肠、脂肪体和表皮组织以及所有发育阶段都检测到了SeCYP9A11和SeCPR基因。暴露于植物次生代谢物槲皮素和单宁后，SeCYP9A11和SeCPR的mRNA水平显著下降。本研究中关于SeCYP9A11和SeCPR基因的结果为进一步研究甜菜夜蛾P450系统提供了基础。
