产甲酸草酸杆菌oxc基因的克隆及其蛋白结构预测

Cloning and structure prediction of oxc gene of oxalyl-coenzyme A decarboxylase from Oxalobacter formigenes

参考GenBank中已公布的产甲酸草酸杆菌(Oxalobacter formigenes,OxB)的参考序列(M77128)设计了一对扩增OXC酶蛋白基因的引物,对产甲酸草酸杆菌(ATCC 35274)的DNA抽提物进行扩增,获得了特异性的扩增片段.运用Lnternet网络上的数据库及程序,对OXC酶蛋白的理化性质、结构和功能进行生物信息学分析.结果显示克隆序列与参考序列的核苷酸序列以及推导的氨基酸序列的同源性达到99.9%和99.6%,预测该酶蛋白是分泌到胞外行使生物学功能的胞外酶,具有良好的抗原性和亲水性,二级结构是以α-螺旋为主的混合型蛋白而且其二级结构预测显示OXC酶蛋白属于硫胺素焦磷酸依赖酶家族,和其它酶家族成员一样能够发挥降解草酸的功能.

The purpose of this study is to clone the oxc gene encoding the oxalic acid-degrading enzyme oxalyl-coenzyme A decarboxylase from Oxalobacter formigenes（OxB,ATCC 35274）and to obtain some clues of OXC protein structure.A pair of primers for amplification of oxc gene was designed and synthesized according to the published oxalobacter formigenes oxc gene sequence（GenBank M77128）.The specific PCR product was successfully obtained and cloned from the Oxalobacter formigenes（OxB,ATCC 35274）.The results showed that the cloned oxc gene had a 99.9 % homology with the reference sequence（GenBank M77128） and the deduced amino acid sequence showed a significant homology 99.6 %.The expressed enzyme belonged to the exocellular enzyme and had good antigenicity and hydrophilicity.Many of α-helix was found in the OXC protein second structure which was mixed protein.The prediction of OXC protein second structure showed that it belong to thiamin diphosphate dependent enzyme family.The OXC protein can catabolize the highly toxic compound oxalate like the other enzyme of thiamin diphosphate dependent enzyme.