利用易错PCR技术,建立脂肪酶基因随机突变文库,将随机突变脂肪酶基因转化毕赤酵母GS115,初步筛选了4000株突变菌株,对300株较优突变株进行酶的活力、耐热性、耐酸性的摇管复筛,进一步摇瓶复筛后获得优良突变株ep3,所产突变脂肪酶(ep3-GS)的适宜pH为9.4,适宜作用温度为35℃,与野生重组脂肪酶(PEL-GS)一致,该温度下酶的比活为3440 U/mg,比野生型脂肪酶提高17%。

A random mutated lipase gene library was constructed by error prone PCR technique.4000 strains of GS115 containing random mutant lipases were primary screened.300 strains were secondary screened based on lipase activity,thermostability and acid resistance,one mutant strain named ep3 was finally selected by flask fermentation.The optimum pH and optimum working temperature of ep3 lipase was pH 9.4 and 35℃,respectively,which was the same as that of the wild type recombinant lipase(PEL-GS),and the specific activity of the lipase at pH 9.4 and 35℃ was 3 440 U/mg,which was 17% higher than that of the wild type lipase.