Expression of the gene of glycerol dehydrogenase from Hansenula polymorpha Dl-1 in Escherichia coli for the production of chiral compounds
Expression of the gene of glycerol dehydrogenase from Hansenula polymorpha Dl-1 in Escherichia coli for the production of chiral compounds
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DOI：
10.1002/1521-3846(200207)22:3/4
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发表时间：
2002-01-01
期刊：
影响因子：
--
通讯作者：
Tani, Y
中科院分区：
文献类型：
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作者：
Yamada-Onodera, K;Yamamoto, H;Tani, Y
为了生产手性化合物，对一株大肠杆菌1113101进行了转化。利用手性色谱柱可以拆分2，3-丁二醇异构体和乙酰异构体，证明了该酶的立体专一性。该基因产物甘油脱氢酶催化依赖NAD+的2，3-丁二醇氧化制乙酰基以及相应的逆反应。含有该表达载体的重组大肠杆菌HB101产生了(3R)-乙酰基[99%产率，99.9%对映体过量(e.E.)]。从10 mM(2R，3R)-2，3-丁二醇和(3S)-乙酸乙酯(产率99%，99.9%e.E.)从110 mM的2，3-丁二醇孵育24小时后，对R构型的仲醇表现出特异性。由110 mM的2，3-丁二醇(摩尔比为17(2R，3R)：15(2S，3S)：78(Meso))，(2S，3S)2，3-丁二醇(15 mM，92%e.E.)在静息细胞反应中，不需要任何添加剂从NADH中再生NAD+。
For the production of chiral compounds, an Escherichia coli 1113 10 1 strain was transformed. A chiral column that could resolve the three 2,3-butanediol isomers and the two acetoin isomers was used to demonstrate the stereospecificity of the enzyme. The H.polymorpha DL-1 gene product, glycerol dehydrogenase, catalyses the NAD+-dependent oxidation of 2,3-butanediol to acetoin as well as the corresponding reverse reactions. The recombinant E. coli HB101 strain harbouring the expression plasmid produced (3R)-acetoin [99% yield, > 99.9% enantiomeric excess (e.e.)] from I 10 mM (2R, 3R)-2,3-butanediol and (3S)-acetoin (99% yield, > 99.9% e.e.) from 110 mM meso-2,3-butanediol after 24 h of incubation, showing specificity towards the secondary alcohol in R-configuration. From a racemate of 110 mM 2,3-butanediol (the molar ratio of 17 (2R, 3R) : 15 (2S, 3S) : 78 (Meso)), (2S, 3s)2,3-butanediol (15 mM, 92% e.e.) was obtained in the resting-cell reaction without any additive to regenerate NAD+ from NADH.