| mdlA基因在毕赤酵母中的高效表达及表达产物性质研究 |
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| 引用本文： | 李琦,王雅琴,谭天伟,王子镐. mdlA基因在毕赤酵母中的高效表达及表达产物性质研究[J]. 微生物学通报, 2006, 33(1): 18-23 |
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| 作者姓名： | 李琦 王雅琴 谭天伟 王子镐 |
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| 作者单位： | 1. 北京化工大学生命科学与技术学院,北京,100029
2. 北京市生物加工过程重点实验室,北京,100029 |
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| 基金项目： | 北京市生物加工过程重点实验室开放项目(No.SYS100100421) |
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| 摘 要： | 将编码甘油单-二酰酯脂肪酶(MDGL)的基因mdlA插入到分泌表达质粒pPIC9K中,通过电激将线性化的重组质粒整合到毕赤酵母(Pichia pastoris)GS115中,筛选出H is Mut 表型菌株,进一步用G418筛选获得高拷贝转化子,并用PCR方法鉴定。诱导培养后,SDS-PAGE表明MDGL在毕赤酵母中得到有效表达。表达产物在温度40℃,pH7.5具有最高活性,其发酵液酶活可达到325U/mL,以橄榄油为底物时没有检测到活性。表达产物与甘油三酰酯脂肪酶共同作用时产生的脂肪酸量比甘油三酰酯脂肪酶单独作用提高了93.5%。
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| 关 键 词： | 甘油单-二酰酯脂肪酶 毕赤酵母 表达 |
| 文章编号： | 0253-2654（2006）01-0018-06 |
| 收稿时间： | 2005-03-17 |
| 修稿时间： | 2005-04-25 |
High Expression of mdlA Gene in Pichia pastoris and Study of Properties of the Recombinant Lipase |
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| LI Qi,WANG Ya-Qin,TAN Tian-Wei,WANG Zi-Hao. High Expression of mdlA Gene in Pichia pastoris and Study of Properties of the Recombinant Lipase[J]. Microbiology China, 2006, 33(1): 18-23 |
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| Authors: | LI Qi WANG Ya-Qin TAN Tian-Wei WANG Zi-Hao |
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| Affiliation: | 1. College of Life Science and Technology, Beijing University of Chemical Technology, Beijing 100029 ; 2. Beijing Key Laboratory of Bioprocess, Beijing 100029 |
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| Abstract: | The mono-and diacylglycerol lipase(MDGL)-encoding gene(mdlA) was inserted into methanol-inducible expression vector pPIC9K.The linearized recombinant plasmid was uransformed into chromosome of Pichia pastoris GS115 strain by electroporation.Some high-copy transformants(His~( )Mut~( )) were picked up by G-418 and conformed by PCR.SDS-PAGE analysis indicated efficient expression of recombinant MDGL.Some enzymatic properties of the recombinant MDGL were also determined.The activity of the recombinant MDGL was up to 325U/mL under the optimal conditions and no activity was detected towards olive oil.The amount of fatty acid produced by the catalysis of recombinant MDGL and triacylglycerol lipase had a increase of 93.5% over triacylglycerol lipase lonely. |
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| Keywords: | Mono-and diacylglycerol lipase Pichia pastoris Expression |
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