甘薯茎线虫乙酰胆碱酯酶基因ace-3全长cDNA的克隆和序列分析

利用RT- PCR结合RACE技术克隆了甘薯茎线虫（Ditylenchus destructor）乙酰胆碱酯酶基因cDNA，Dd-ace-3，提交GenBank登录号EF583057。该cDNA序列5’端存在反式剪接引导序列SL1,全长2517bp ,包含一个1836bp的开放阅读框；在推导出的611个氨基酸残基的前体蛋白中, N 端的前21个氨基酸残基为信号肽, C-28的位置存在糖基磷脂酰肌醇(GPI)锚定位点，除此之外的562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64396.81D。在一级结构中,形成催化活性中心的3个氨基酸残基 (Ser232 ,Glu360 和His479) ,以及在亚基内形成二硫键的6个半胱氨酸完全保守；在电鳐乙酰胆碱酯酶分子的催化功能域中存在14个保守的芳香族氨基酸残基,其中11个在甘薯茎线虫乙酰胆碱酯酶中完全保守。该酶的氨基酸序列与秀丽小杆线虫（Caenorhabditis elegans）ACE-3和ACE-4的同源性达56.1%和50.0%。与其它线虫和物种乙酰胆碱酯酶的聚类分析显示，甘薯茎线虫的乙酰胆碱酯酶与Ш型乙酰胆碱酯酶ACE-3同属一个支系。

Cloning and Sequence Analysis of a New Acetylcholinesterase Gene ace-3 from Ditylenchus destructor

A new cDNA, named Dd-ace-1, encoding an acetylcholinesterase (AChE, EC3.1.1.7) was isolated from the sweet potato stem nematode, Ditylenchus destructor. The full-length cDNA, carrying the trans-spliced SL1 lea- der sequence, is 2517bp long with an open reading frame of 836bp encoding 611 amino acid residues (GenBank a- cession no. EF583057). The complete amino acid sequence of AChE deduced from the cDNA consists of 21 resi- dues for the putative signal peptide and possesses a potential cleavage-addition site(ω) for a glycophosphatidylin- ositol anchor(GPI) at position 583 and 562 residues for the mature protein with a predicted molecular weight of 70144.12D. The conserved motifs involved in the catalytic triad, the choline binding sit and 11 aromatic residues lining the catalytic gorge were present in the Dd-ACE-1 deduced protein. The prediected protein shared with str- ong homology with Caenorhabditis elegans ACE-3 and C. elegans ACE-4. Phylogenetic analysis based on other nematodes and species AChEs showed that the deduced AChE formed a cluster with ACE-3s.