宇佐美曲霉环氧化物水解酶基因的克隆与生物信息学分析  

Cloning and Bioinformatics Analysis of Epoxide Hydrolase Gene from Aspergillus usamii

作　　者：胡蝶[1] 朱利娟[2] 邬敏辰[3] 汪俊卿[1] 唐存多[1] 冯峰[2] 余涛[2] 

机构地区：[1]江南大学生物工程学院,江苏无锡214122 [2]江南大学药学院,江苏无锡214122 [3]江南大学无锡医学院,江苏无锡214122

出　　处：《食品与生物技术学报》2013年第12期1244-1252,共9页Journal of Food Science and Biotechnology

基　　金：国家自然科学基金项目(31101229);国家自然科学基金项目(31271811)

摘　　要：环氧化物水解酶(Epoxide hydrolase,EH)是酶法拆分消旋体环氧化物,制备光学活性环氧化物和邻二醇的重要酶之一。通过RT-PCR和新构建的THSO-PCR侧翼未知DNA序列扩增技术,克隆了一种来源于宇佐美曲霉(Aspergillus usamii)E001的EH基因,命名为Aueh2(GenBank No.KF061095),并对该基因进行了相关生物信息学分析。Aueh2的DNA序列长度为2 481 bp,其中包含了5′端和3′端侧翼调控序列,6个内含子序列和编码cDNA序列。开放阅读框序列长度为1 188 bp,编码395个氨基酸,对应的蛋白质命名为AuEH2;该蛋白质为无信号肽的亲水蛋白质,其理论相对分子质量44.6 kD;其三维结构包含EH典型的"α/β"核心催化结构域和"帽子"结构域,活性中心由催化三联体Asp191、His369和Glu343组成。本课题研究成果为深入研究AuEH2及其应用奠定了基础。Epoxide hydrolase can be effectively used in the resolution of epoxides for producting optically active epoxides and vicinal diols. A gene encoding a novel epoxide hydrolase from Aspergillus usamii E001 was cloned by using reverse transcription PCR and newly constructed T- hairpin structure-mediated PCR amplification （THSO-PCR） techniques. The cloned gene（named Aueh2） is 2,481 bp in length,harboring 5＇ and 3＇ flanking regulatory regions and the encoded cDNA sequence interrupted by six introns. The open reading frame of A ueh2 encodes a protein of 395-aa（designated AuEH2） with the calculated molecular weight of 44.6 kD. The primary structure analysis of AuEH2 demonstrated that it is a hydrophilic protein,and belongs to the α/β hydrolase fold family. The structure of AuEH2 displays that the catalytic center is situated between the α/β core domain and the lid domain with a catalytic triad consisting of Asp^191,His^369,and Glu^343. The results will lay a foundation for further research of the AuEH2 and its application in depth.

关 键 词：宇佐美曲霉 环氧化物水解酶 侧翼未知DNA序列 基因克隆 生物信息学分析 

分 类 号：Q78[生物学—分子生物学]