嗜水气单胞菌丝氨酸蛋白酶基因克隆与序列分析  

Cloning and Sequence Analysis of Serine Protease Gene from Aeromonas hydrophila

作　　者：胡秀彩[1] 李雪[2] 兰云[2] 张培[2] 沈晓静[2] 吕爱军[1,2] 朱爱华[1,2] 

机构地区：[1]江苏省药用植物生物技术重点实验室 [2]江苏师范大学生命科学学院,江苏徐州221116

出　　处：《生物技术》2014年第3期5-8,共4页Biotechnology

基　　金：国家自然科学基金项目(No.31272692;No.30800847)资助

摘　　要：目的:克隆嗜水气单胞菌丝氨酸蛋白酶(Ahp)基因,进一步分析序列、预测三级结构。方法:设计Ahp基因特异性引物,利用PCR方法扩增基因序列,亚克隆至pMD18-T载体,进行DNA测序及生物信息学分析。结果:获得Ahp基因大小为1 645bp,推导编码548aa,含有天冬氨酸(82-93aa)、组氨酸(123-133aa)和丝氨酸活性位点(342-352aa)。Ahp蛋白三级结构与模板(PDB:3HJR_A)相似性达81.48%,预测60-392aa区域为肽酶S8结构域(PF00082)、480-548aa区域为前蛋白转化酶P结构域(PF01483),其中3个氨基酸活性位点分布于三级结构N端,与拉马钱德兰图检测分析Ahp蛋白的空间结构基本一致。结论:该研究对嗜水气单胞菌Zf1菌株Ahp基因编码蛋白进行结构预测,有助于理解嗜水气单胞菌丝氨酸蛋白酶的作用机理。Objective： Serine protease gene of Aeromonas hydrophila Zfl was cloned and the structure of the protein was further predicted. Method： The serine protease gene （Ahp） was amplified by PCR and then was subcloned into pMD18 -T vector. It was analyzed by DNA sequencing and bioinformatics methods. Result： The sequencing analysis of Ahp gene fragment was 1 645bp long, encoding a protein of 548 amino acids which has aspartic acid active site （82 -93aa）, histidine active site （123 - 133aa） and serine active site （342 -352aa）. Homology modeling was used to construct 3D structure of Ahp protein with the standard model （PDB： 3HJR_A） , in which N - terminus the three active sites were located. Conclusion ： The sequences analysis and structure predictions of Ahp protein help to understand Aeromonas hydrophila pathogenesis mechanisms.

关 键 词：嗜水气单胞菌 丝氨酸蛋白酶 序列分析 三级结构 

分 类 号：Q785[生物学—分子生物学]