Directed evolution and expression tuning of geraniol synthase for efficient geraniol production in Escherichia coli.
Directed evolution and expression tuning of geraniol synthase for efficient geraniol production in Escherichia coli.
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DOI：
10.2323/jgam.2017.01.006
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发表时间：
2017-09
期刊：
影响因子：
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通讯作者：
M. Tashiro;A. Fujii;S. Kawai-Noma;Kyoichi Saito;D. Umeno
中科院分区：
文献类型：
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作者：
M. Tashiro;A. Fujii;S. Kawai-Noma;Kyoichi Saito;D. Umeno
为了实现在大肠杆菌中高效生产香叶醇及其衍生物，我们旨在通过单轮诱变和筛选更高的底物消耗来提高香叶醇合酶（GES）的活性。我们分离出在香叶醇生产中表现优于其亲本的GES变体。对GES变异体的分析表明，GES的表达水平是香叶醇合成的瓶颈。具有设计用于高翻译效率的5 '-非翻译序列的突变体GESM 53的过表达，沿着甲羟戊酸途径酶、异戊烯基焦磷酸异构酶和香叶基焦磷酸合酶的额外表达，在摇瓶中产生300 mg/L/12 h香叶醇及其衍生物（总计>1000 mg/L/42 h）。
To achieve an efficient production of geraniol and its derivatives in Escherichia coli, we aimed to improve the activity of geraniol synthase (GES) through a single round of mutagenesis and screening for higher substrate consumption. We isolated GES variants that outperform their parent in geraniol production. The analysis of GES variants indicated that the expression level of GES was the bottleneck for geraniol synthesis. Over-expression of the mutant GESM53 with a 5'-untranslated sequence designed for high translational efficiency, along with the additional expression of mevalonate pathway enzymes, isopentenyl pyrophosphate isomerase, and geranyl pyrophosphate synthase, yielded 300 mg/L/12 h geraniol and its derivatives (>1000 mg/L/42 h in total) in a shaking flask.