[Construction of novel recombinant Escherichia coli capable of producing 1,3-propanediol].
[Construction of novel recombinant Escherichia coli capable of producing 1,3-propanediol].
复制标题
DOI：
--
复制
发表时间：
2005-09
期刊：
影响因子：
--
通讯作者：
Xiao-mei Zhang;Xueming Tang;B. Zhuge;W. Shen;Z. Rao;H. Fang;J. Zhuge
中科院分区：
文献类型：
--
作者：
Xiao-mei Zhang;Xueming Tang;B. Zhuge;W. Shen;Z. Rao;H. Fang;J. Zhuge
采用PCR扩增大肠杆菌1,3-丙二醇氧化还原酶同工酶编码基因（yqhD）。将yqhD插入pEtac中，得到重组表达载体pEtac-yqhD。pEtac-yqhD在大肠杆菌JM109中过表达。SDS-PAGE分析显示一个过表达的重组产物约为43 kD，与基因序列预测的分子量一致。与大肠杆菌JM109 （pEtac）相比，重组大肠杆菌（pEtac- yqhd）在1.0 mmol/L IPTG诱导下，37℃作用4小时，1,3-丙二醇氧化还原酶同工酶活性达到120 u/mg蛋白；在相同条件下，大肠杆菌JM109 （pEtac）的酶活性仅为0.5 u/mg蛋白。构建了重组大肠杆菌JM109 （pUCtac-dhaB, pEtac-yqhD）。重组菌株经1.0 mmol/L IPTG诱导后，在好氧条件下可将50 g/L甘油转化为38 g/L 1,3-丙二醇。本研究首次证明了1,3-丙二醇氧化还原酶同工酶在有氧条件下具有较高的活性。
The 1,3-propanediol oxidoreductase isoenzyme encoding gene (yqhD) from E. coli was amplified by PCR. yqhD was inserted in pEtac to yield the recombinant expression vector pEtac-yqhD. Over-expression of yqhD in E. coli JM109 was achieved with pEtac-yqhD. SDS-PAGE analysis showed an over-expressed recombinant product at about 43 kD, consistent with the molecular weight predicted from gene sequence. Compared with E. coli JM109 (pEtac), the 1,3-propanediol oxidoreductase isoenzyme activity of the recombinant E. coli (pEtac-yqhD) reached 120 u/mg protein under the induction of 1.0 mmol/L IPTG at 37 degrees C for 4 hours; at similar conditions, enzyme activity of E. coli JM109 (pEtac) was only 0.5 u/mg protein. The recombinant E. coli JM109 (pUCtac-dhaB, pEtac-yqhD) was constructed. After induction with 1.0 mmol/L IPTG, the recombinant strain could transform 50 g/L glycerol to 38 g/L 1,3-propanediol under aerobic conditions. This work demonstrated firstly that the 1,3-propanediol oxidoreductase isoenzyme could show high activity under aerobic conditions.