| 两种策略实现1,3-丙二醇关键酶基因的共表达 |
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| 引用本文： | 曲荟锦,王凤寰,田平芳,谭天伟.两种策略实现1,3-丙二醇关键酶基因的共表达[J].北京化工大学学报(自然科学版),2007,34(4):421-424. |
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| 作者姓名： | 曲荟锦 王凤寰 田平芳 谭天伟 |
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| 作者单位： | 北京化工大学 1生命科学与技术学院；
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： | 国家重点基础研究发展计划(973计划)
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国家自然科学基金
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国家自然科学基金
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北京市自然科学基金
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北京市科技计划 |
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| 摘 要： | 甘油脱水酶(GDHt)和1,3-丙二醇氧化还原酶(PDOR)是甘油歧化为1,3-丙二醇(1,3-PD)的两个关键酶。采用多顺反子重组和质粒共存两种策略，对来自克雷伯肺炎杆菌(Klebsiela pneumoniae)的两个关键酶进行共表达。构建表达载体pET-28a-dhaB1B2B3-dhaT将两酶基因dhaB1B2B3和dhaT用SD序列相隔，在E.coli BL21(DE3)高水平共表达了GDHt 3个亚基和PDOR，表达蛋白分别约占菌体总蛋白的18%、9%、7%和9%。质粒pET-28a-dhaB1B2B3和pET-22b-dhaT共转化E.coli BL21(DE3)得到稳定的双质粒系统，48h后84%的细胞能同时含有两种质粒，GDHt 3亚基和PDOR分别约占菌体总蛋白的16%、8%、6%和14%。两种酶在两种表达方法下均显示高于原始菌株的酶活力。
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| 关 键 词： | 甘油脱水酶 1 3-丙二醇氧化还原酶 共表达 多顺反子重组 质粒共存 甘油脱水酶 1 3-丙二醇氧化还原酶 共表达 多顺反子重组 质粒共存 |
| 修稿时间： | 2007-01-27 |
Two strategies for coexpression of the key enzymes involved in 1,3-propanediol production |
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| Qu HuiJin,WANG FengHuan,TIAN PingFang,TAN TianWei.Two strategies for coexpression of the key enzymes involved in 1,3-propanediol production[J].Journal of Beijing University of Chemical Technology,2007,34(4):421-424. |
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| Authors: | Qu HuiJin WANG FengHuan TIAN PingFang TAN TianWei |
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| Affiliation: | 1College of Life Science and Technology; 2Beijing Key Laboratory of Bioprocess, Beijing University of Chemical Technology, Beijing 100029, China |
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| Abstract: | Glycerol dehydratase(GDHt) and 1,3-propanediol oxidoreductase(PDOR) are the two key enzymes that catalyse the conversion of glycerol to 1,3-propanediol(1,3-PD).In this study,PDOR and GDHt from Klebsiella pneumoniae were coexpressed by polycistronic expression and coexistence of incompatible plasmids.The recombinant plasmid pET-28a-dhaB1B2B3-dhaT harboring the genes encoding GDHt and PDOR with a Shine-Dalgarno(SD) sequence was transformed into E.coli BL21(DE3).After induction with isopropyl-beta-D-thiogalactoside(IPTG),the three subunits of GDHt as well as PDOR were highly coexpressed,accounting for 18%,9%,7% and 9% respectively of the total protein in the host.A stable two plasmid system was obtained via cotransformation of pET-28a-dhaB1B2B3 and pET-22b-dhaT into E.coli BL21(DE3).Approximately 84% of the daughter cells maintained the two plasmids after 48 fermentation hours and the proportion of expressed enzymes was 16%,8%,6% and 14%,respectively. The activities of the two enzymes are also discussed in this paper. |
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| Keywords: | glycerol dehydratase 1 3-propanediol dehydrogenase coexpression polycistronic expression plas mid coexistence |
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