【摘要】 利用含有三油酸甘油酯的脂肪酶选择性筛选培养基,从奶牛瘤胃微生物元基因组文库15360个克隆中,筛选得到了18个脂肪酶阳性克隆,其插入片段大约为60kb,并且各个克隆的插入片段各不一样。利用p-NPP法对脂肪酶克隆的脂肪酶活性分析,表明均具有大小不等的脂肪酶活性。底物特异性分析表明Lipase6、Lipase7和Lipase8分别对C16底物(对硝基苯棕榈酸酯)、C12底物(对硝基苯月桂酸酯)和C16底物(对硝基苯棕榈酸酯)水解能力最强。Lipase6、Lipase7、Lipase8的脂肪酶最适pH为7.5;Lipase8的脂肪酶活性半衰期随反应温度的升高而缩短,70oC时能达到30min。本研究所筛选的脂肪酶具有不同的底物特异性和较好的热稳定性,这对于工业化生产具有一定的应用潜力。更多还原

【Abstract】 Using lipase segregation agar containing trioleoylglycerol,we obtained 18 lipase positive clones by screening from a metagenome library of dairy rumen microflora containing 15 360 clones.The average insert size of lipase positive clones was about 60 kb.Lipase enzyme activity analysis by p-NPP method indicated that Lipase6,Lipase7 and Lipase8 had higher lipolytic activities to substrates of p-nitrophenyl palmitate(C16),p-nitrophenyl alaurate(C12) and p-nitrophenyl palmitate(C16) respectively.The optimum pH of Lipase 6,Lipase 7 and Lipase 8 were 7.5.The halflife period of Lipase 8 with the value of 15 min in 70oC decreased with the increase of temperature.In conclusion,the lipases screened in this study had different substrates specificity and good thermo stability,which laid a basis for large-scale industrial application.更多还原