Engineering Corynebacterium glutamicum for the Efficient Production of 3-Hydroxypropionic Acid from a Mixture of Glucose and Acetate via the Malonyl-CoA Pathway
Engineering Corynebacterium glutamicum for the Efficient Production of 3-Hydroxypropionic Acid from a Mixture of Glucose and Acetate via the Malonyl-CoA Pathway
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改造谷氨酸棒杆菌，通过丙二酰辅酶 A 途径从葡萄糖和乙酸的混合物中高效生产 3-羟基丙酸
DOI：
10.3390/catal10020203
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发表时间：
2020-02-01
期刊：
影响因子：
3.9
通讯作者：
Chen, Tao
中科院分区：
文献类型：
--
作者：
Chang, Zhishuai;Dai, Wei;Chen, Tao
3 - 羟基丙酸（3 - HP）由于其众多潜在应用，已被视为最具增值潜力的基础化学品之一。在过去十年中，通过丙二酸单酰 - CoA途径生物合成3 - HP越来越受到青睐，因为它在ATP和还原当量方面是平衡的，不需要添加昂贵的辅酶，并且能够利用可再生的木质纤维素生物质。在本研究中，将来自橙色绿屈挠菌的编码丙二酸单酰 - CoA还原酶的基因mcr导入谷氨酸棒杆菌ATCC13032中构建了菌株Cgz1，该菌株积累了0.30 g/L的3 - HP。随后删除编码乳酸脱氢酶的基因ldhA以消除乳酸积累，但这降低了3 - HP的产量并大幅增加了乙酸积累。然后，过表达不同的乙酸利用基因以重新利用乙酸，表达内源基因pta的最佳候选菌株Cgz5能够有效减少乙酸积累并产生0.68 g/L的3 - HP。为了增加前体乙酰 - CoA的供应，使用乙酸作为辅助碳源来提高3 - HP的产量，从葡萄糖和乙酸的混合物中可产生1.33 g/L的3 - HP，产量比单独使用葡萄糖高2.06倍。最后，为了抑制主要的与3 - HP竞争的途径——脂肪酸合成，添加10 μM浅蓝菌素，菌株Cgz5在48小时内从15.47 g/L葡萄糖和4.68 g/L乙酸中产生3.77 g/L的3 - HP，底物得率为187 mg/g，比Cgz1高12.57倍。据我们所知，这是第一篇关于改造谷氨酸棒杆菌通过丙二酸单酰 - CoA途径生产3 - HP的报道。结果表明，无害的生物安全等级为I的微生物谷氨酸棒杆菌是一种潜在的工业化3 - HP生产菌株。
3-Hydroxypropionic acid (3-HP) has been recognized as one of the top value-added building block chemicals, due to its numerous potential applications. Over the past decade, biosynthesis of 3-HP via the malonyl-CoA pathway has been increasingly favored because it is balanced in terms of ATP and reducing equivalents, does not require the addition of costly coenzymes, and can utilize renewable lignocellulosic biomass. In this study, gene mcr encoding malonyl-CoA reductase from Chloroflexus aurantiacus was introduced into Corynebacterium glutamicum ATCC13032 to construct the strain Cgz1, which accumulated 0.30 g/L 3-HP. Gene ldhA encoding lactate dehydrogenase was subsequently deleted to eliminate lactate accumulation, but this decreased 3-HP production and greatly increased acetate accumulation. Then, different acetate utilization genes were overexpressed to reuse the acetate, and the best candidate Cgz5 expressing endogenous gene pta could effectively reduce the acetate accumulation and produced 0.68 g/L 3-HP. To enhance the supply of the precursor acetyl-CoA, acetate was used as an ancillary carbon source to improve the 3-HP production, and 1.33 g/L 3-HP could be produced from a mixture of glucose and acetate, with a 2.06-fold higher yield than from glucose alone. Finally, to inhibit the major 3-HP competing pathway-fatty acid synthesis, 10 μM cerulenin was added and strain Cgz5 produced 3.77 g/L 3-HP from 15.47 g/L glucose and 4.68 g/L acetate with a yield of 187 mg/g substrate in 48 h, which was 12.57-fold higher than that of Cgz1. To our best knowledge, this is the first report on engineering C. glutamicum to produce 3-HP via the malonyl-CoA pathway. The results indicate that the innocuous biosafety level I microorganism C. glutamicum is a potential industrial 3-HP producer.