几种南海芋螺二硫键异构酶的克隆及进化分析

摘要

目的:从来自中国南海的4种芋螺中克隆出包含完整3′和5′非翻译区的蛋白质二硫键异构酶(PDI)全基因序列,并对其进行序列及进化分析.方法:根据各种生物PDI基因的保守区域设计引物,利用3′和5′cDNA末端快速扩增(RACE)方法克隆出PDI全基因序列,并通过生物信息学方法对各芋螺PDI序列进行分析.结果与结论:从中国南海玉女芋螺、黑星芋螺、堂皇芋螺、桶形芋螺cDNA中克隆出包含有完整3′和5′非翻译区的PDI全基因序列;分析结果表明各芋螺之间的同源性大于90%,而与对虾、人类、酿酒酵母的同源性均小于60%;各芋螺PDI与其他生物的2个活性位点序列高度保守,而底物结合位点具有物种特异性,进化树显示各芋螺PDI的特征可能受其捕食食性影响.%Objective: To clone and analyze entire nucleotide sequences of protein disulfide isomerase(PDI) of four Conus from South China Sea.Methods: The entire nucleotide sequences of PDI of Conus from the South China Sea were cloned by 3' and 5' rapid amplification of cDNA ends(RACE), which primers were designed on conservative amino acid sequences of other PDI, and the sequences were analyzed by bioinformatics.Results & Conclution: The homology of the deduced amino acid sequences among these Conus species was more than 90％, and higher than that of the Litopenaeus vannamei, Homo sapiens and Saccharomyces cerevisiae.Two active sites of PDI were highly conservative between Conus and other organisms, but the substrate binding sites were species specific.Phylogenetic tree analysis showed that the mutants of PDI from Conus might be affected by their predator diet.