葡激酶基因的分离及其在大肠杆菌中的高效表达  

Isolation of the gene encoding staphylokinase and its high expression in E.coli

作　　者：邹民吉[1] 王嘉玺[1] 赵春文[1] 王利红[1] 段聚宝[1] 

机构地区：[1]军事医学科学院基础医学研究所

出　　处：《军事医学科学院院刊》1998年第4期257-59,292,共1页Bulletin of the Academy of Military Medical Sciences

摘　　要：目的：分离葡激酶基因并使其在Ｅ．ｃｏｌｉ中高效表达。方法：利用ＰＣＲ技术自溶源性金黄色葡萄球菌ＦＲ６１０株的染色体ＤＮＡ中分离葡激酶编码序列，ＤＮＡ序列分析后，组入高效表达载体ｐＢＶ２２０中，转化至大肠杆菌。结果和结论：分离的葡激酶基因与文献报道相比，第１０８位核苷酸由腺嘌呤核苷酸变异为鸟嘌呤核苷酸，导致了成熟葡激酶第３７位氨基酸由精氨酸变异为甘氨酸。表达结果显示，表达产物占菌体总蛋白的７０％，获得了高效表达菌株。初步纯化表达蛋白，以链激酶为标准品测定其溶栓活性。Objective: To isolate the coding region of the staphylokinase(SAK) gene, and express it in E.coli. Methods: Amplifying the SAK coding sequence from the chromosomal DNA of the lysogen of Staphylococcus aureus strain FR610. And then the isolated fragment was sequenced by dideoxy method, recombined in vitro with plasmid pBV220, and transformed E.coli DH5α cells. Results and Conclusion: An high expression strain was obtained. The expressed SAK amounted to 70% of the total bacterial protein. The specific activity of the SAK was 3×10 6 units/mg, measured by comparison with a standard preparation. A high expression [WTBX]E.coli strain for SAK was successfully obtained.

关 键 词：葡萄球菌激酶 克隆表达 溶栓活性 大肠杆菌 分离 

分 类 号：Q78[生物学—分子生物学]