褐飞虱乙酰胆碱酯酶基因全长cDNA的克隆及序列分析  

Molecular cloning and characterization of an acetylcholinesterase cDNA in the brown planthopper,Nilaparvata lugens(Homoptera:Delphacidae)

作　　者：杨之帆[1] 何光存[2] 

机构地区：[1]湖北大学生命科学学院,武汉430062 [2]武汉大学生命科学学院,武汉430072

出　　处：《昆虫学报》2006年第6期1034-1041,共8页Acta Entomologica Sinica

基　　金：国家自然科学基金项目(30500328;30570140)

摘　　要：利用反转录聚合酶链式反应(RT_PCR)结合快速扩增cDNA末端(RACE)技术克隆了褐飞虱Nilaparvata lugens乙酰胆碱酯酶基因cDNA。该cDNA全长2467bp,包含一个1938bp的开放阅读框(GenBank登录号AJ852420);在推导出的646个氨基酸残基的前体蛋白中,N端的前30个氨基酸残基为信号肽,随后的616个氨基酸残基是成熟的乙酰胆碱酯酶序列,其预测的分子量为69418D。在一级结构中,形成催化活性中心的3个氨基酸残基(Ser242,Glu371和His485),以及在亚基内形成二硫键的6个半胱氨酸完全保守;位于催化功能域的14个芳香族氨基酸中有10个完全保守。该酶的氨基酸序列与黑尾叶蝉的同源性最高,达83%。对来自23种昆虫(包括褐飞虱)的30个乙酰胆碱酯酶的聚类分析显示,褐飞虱的乙酰胆碱酯酶与其中6个Ⅱ型乙酰胆碱酯酶(AChE2)同属一个支系;此外,只存在于昆虫AChE2中的超变区及特异的氨基酸残基,也存在于褐飞虱的乙酰胆碱酯酶中。以上结果表明,所克隆的褐飞虱的乙酰胆碱酯酶基因是一个与黑腹果蝇的orthologous型基因同源的AChE2基因。A full cDNA encoding an acetylcholinesterase （ACHE, EC3.1.1.7） from the brown planthopper, Nilaparvata lugens （Homoptera： Delphacidae） was cloned and characterized. The complete cDNA （2 467 bp） contains a 1 938 bp open reading frame encoding 646 amino acid residues （GenBank accession no. AJ852420） . The complete amino acid sequence of AChE deduced from the cDNA consists of 30 residues for the putative signal peptide and 616 residues for the mature protein with a predicted molecular weight of 69 418 D. The three residues （Ser242, Glu371 and His485） that putatively form the catalytic triad and six Cys that form intra-subunit disulfide bonds are completely conserved, and 10 out of the 14 aromatic residues lining the active site gorge of the AChE are also conserved. The deduced amino acid sequence is most similar to AChE of Nephotettix cincticeps with 83 % amino acid identity. Phylogenetic analysis based on 30 AChEs from 23 insect species showed that the deduced AChE formed a cluster with 6 AChE2s. Additionally, the hypervariable region and amino acids specific to insect ACHE2 also exist in N. lugens ACHE. The results indicate that the AChE gene cloned in this work belongs to insect ACHE2 subgroup, which are homologous to Drosophila ACHE2.

关 键 词：褐飞虱 乙酰胆碱酯酶 CDNA克隆 聚类分析 

分 类 号：Q966[生物学—昆虫学]