裂殖壶菌脂肪酶基因STGL2克隆及底物特异性研究
首发时间:2018-01-16
摘要:裂殖壶菌甘油三酯合成和水解的过程中,脂肪酶起着重要的作用。本文根据同源性分析结果,筛选得到裂殖壶菌脂肪酶基因STGL2,以cDNA为模板获得STGL2脂肪酶基因,将其连接到pPIC9K载体上,得到重组质粒pPIC9K-STGL2,并在毕赤酵母中成功表达。系统分析了重组脂肪酶(Stgl2p)的底物特异性,结果表明:裂殖壶菌脂肪酶Stgl2p的最适底物为对硝基苯酚月桂酸酯(p-NPL),其最适pH为7.5,最适反应温度为35oC。综上所述,Stgl2p脂肪酶在裂殖壶菌细胞内对甘油三酯分子上中长碳链脂肪酸的水解起着重要的作用
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Gene Cloning of Lipase from Schizochytrium sp. and Study of Substrate Specificity
Abstract:Lipases play an important role in the synthesis and hydrolysis of triglycerides in Schizochytrium. According to the results of homology analysis, Schizochytrium lipase gene STGL2 was screened out. The lipase of STGL2 was obtained as from cDNA , was inserted into the expression vector pPIC9K and transformed into Pichia pastoris SMD1168. The recombinant plasmid pPIC9K-STGL2 was obtained. The results showed that the lipase gene STGL2 was successfully expressed in Pichia pastoris. The optimal substrate for Stgl2p was p-laurate (C12). The optimum pH was 7.5. Suitable reaction temperature was 35oC. In conclusion, lipase of Stgl2p plays an important role in the hydrolysis of medium-long chain fatty acids in triglyceridesv of Schizochytrium lipolytica.
Keywords: Schizochytrium lipase gene Pichia pastoris substrate specificity
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