摘要
目的在大肠杆菌中诱导表达以嗜酸乳杆菌克隆β-半乳糖苷酶基因lacZ,为研究其酶学特性做准备。方法从嗜酸乳杆菌ATCC4356中克隆lacZ基因,并构建表达载体pET22b-lacZ-H、pQE31-H-lacZ和pQE31-lacZ,然后在大肠杆菌中诱导表达,并测定表达产物的β-半乳糖苷酶活性。结果无标签的重组嗜酸乳杆菌β-半乳糖苷酶LacZ获得了功能性表达,表达量可达2.28kU/L.而融合His-Tag的重组嗜酸乳杆菌LacZ却失去了β-半乳糖苷酶活性,即使复性也不能恢复。结论克隆表达的成功为该酶的酶学性质研究和可能的应用打下了基础。
Objective To clone and express the lacZ gene of L. acidophilus in E. coli for research of the properties of the putative β-galactosidase. Methods The lacZ gene was cloned from L. acidophilus ATCC 4356 and was inserted into three recombinant vectors： pET22b-lacZ-H, pQE31-H-lacZ and pQE31-lacZ. The recombinant pro- teins were induced and were identified by SDS-PAGE and β-galactosidase assay. Results The recombinant lacZ with- out tag was expressed as functional β-galactosidase, which yield was 2. 28 kU/L, but the two recombinant LacZs with His-tag were expressed as proteins without activity and its activity could not recovery by renaturation. Conclusion The study make a base for research of enzymatic properties and probable application in the future.
出处
《成都医学院学报》
CAS
2009年第2期86-90,共5页
Journal of Chengdu Medical College
基金
四川省科技攻关项目(05SG022-019)
作者简介
潘渠（1973-），男，成都市人，博士，讲师。E-mail：ppqlove@126．com
通讯作者 李晋川，E-mail：ljc_cdm@126．com
