烟草节杆菌肌酸酶的高效异源表达与应用分析

Expression and Application Analysis of Creatinase from Arthrobacter nicotianae

肌酸酶(CRE)是临床检测血液和尿液中肌酐的关键酶之一。作者成功地从烟草节杆菌23710(Arthrobacter nicotianae 23710)中克隆了CRE基因,并实现了CRE在Escherichia coli BL21(DE3)中的表达。经过硫酸铵分级沉淀、阴离子交换色谱、分子筛后得到了较纯的重组CRE,比酶活达到了20.25 U/mg。重组CRE对于螯合剂EDTA、表面活性剂(Tween20,和Triton X-100)以及常用的防腐剂Na N3有很好的耐受性。

Creatine amidinohydrolase（CRE） is one of the key enzymes for clinical determination of creatinine in serum and urine. In this study,the gene encoding for CRE was successfully amplified from Arthrobacter nicotianae 23710 and functionally overexpressed in Escherichia coli BL21（DE3）.The recombinant CRE was purified through a series of operation including ammonium sulfate precipitation,anion exchange chromatography and gel filtration chromatography. The specific activity of the purified enzyme reached 20.25 U/mg. The recombinant enzyme shows excellent resistance to the chelating agent EDTA,the surfactants（Tween20 and Triton X-100） and the common preservative NaN3.