睾丸酮丛毛单胞菌teiR基因插入失活突变体构建及降解特性分析  

teiR-disrupted Mutant of Comamonas testosteroni Construction and Its Characteristics in Steroid Biodegradation

作　　者：陈建秋[1] 潘大仁[2] 艾育芳[2] 周以飞[1] 潘菲[2] 林彬辉[2] 

机构地区：[1]福建农林大学作物科学学院,福州35002 [2]福建农林大学生命科学学院,福州350002

出　　处：《应用与环境生物学报》2009年第6期830-834,共5页Chinese Journal of Applied and Environmental Biology

基　　金：教育部科学技术重点项目(No.207056);福建省自然科学基金计划项目(No.2008J0041)资助~~

摘　　要：睾丸酮丛毛单胞菌(Comamonas testosteroni)是革兰氏阴性菌,能够以各种类固醇化合物作为唯一碳源和能源,生物信息学分析发现睾丸酮丛毛单胞菌teiR基因编码蛋白属于LuxR-type转录调控蛋白.利用同源重组原理构建teiR基因插入失活突变体C.mut,初步研究了teiR基因在类固醇代谢途径中的作用,以及对睾丸酮丛毛单胞菌关键降解酶3a-HSD/CR的表达影响.结果发现,teiR基因插入失活突变体C.mut在LB培养基中的生长速率不受影响,突变体C.mut抑制了teiR基因和关键降解酶3a-HSD/CR的表达,同时突变体C.mut丧失了以睾丸酮作为碳源进行生长的能力.结果表明teiR基因调节睾丸酮丛毛单胞菌类固醇代谢,同时teiR基因表达对3a-HSD/CR基因的表达是必需的.Comamonas testosteroni is a Gram-negative bacterium,and has the ability to use various steroids as sole carbon and energy sources. Bioinformatics analysis indicates that teiR encoding protein is a LuxR-type regulator. In this paper,recombinant plasmid pKmut was integrated into C. testosteroni chromosome DNA through homologous recombination by electroporation to generate teiR gene knock-out mutant C. mut. Protein expression levels and its degradation ability were determined by enzyme-linked immunosorbent assay and high performance liquid chromatography, respectively. The growth of teiR-disrupted mutant C. mut was not afeected in LB medium, but it inhibited the 3a-HSD/CR and teiR gene expression. Finally, the mutant lost the ability to use testosterone as carbon and energy sources. Our results confirmed that the teiR gene mediated the steroid metabolism in C. testosteroni and was necessary for the 3a-HSD/CR expression. Fig 7, Ref 14

关 键 词：睾丸酮丛毛单胞菌 teiR基因 同源重组 降解酶3a-HSD/CR类固醇 类固醇 

分 类 号：X172[环境科学与工程—环境科学] Q78[生物学—分子生物学]