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美国卫生研究院文献>Frontiers in Bioengineering and Biotechnology
>Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida
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Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida
Sarcosine, an N-methylated amino acid, shows potential as antipsychotic, and serves as building block for peptide-based drugs, and acts as detergent when acetylated. N-methylated amino acids are mainly produced chemically or by biocatalysis, with either low yields or high costs for co-factor regeneration. Corynebacterium glutamicum, which is used for the industrial production of amino acids for decades, has recently been engineered for production of N-methyl-L-alanine and sarcosine. Heterologous expression of dpkA in a C. glutamicum strain engineered for glyoxylate overproduction enabled fermentative production of sarcosine from sugars and monomethylamine. Here, mutation of an amino acyl residue in the substrate binding site of DpkA (DpkAF117L) led to an increased specific activity for reductive alkylamination of glyoxylate using monomethylamine and monoethylamine as substrates. Introduction of DpkAF117L into the production strain accelerated the production of sarcosine and a volumetric productivity of 0.16 g L−1 h−1 could be attained. Using monoethylamine as substrate, we demonstrated N-ethylglycine production with a volumetric productivity of 0.11 g L−1 h−1, which to the best of our knowledge is the first report of its fermentative production. Subsequently, the feasibility of using rice straw hydrolysate as alternative carbon source was tested and production of N-ethylglycine to a titer of 1.6 g L−1 after 60 h of fed-batch bioreactor cultivation could be attained.
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机译：肌氨酸是一种N-甲基化氨基酸，具有抗精神病药的潜力，可作为肽基药物的组成部分，并在乙酰化时起清洁剂的作用。 N-甲基化氨基酸主要通过化学方法或通过生物催化生产，辅酶再生的收率低或成本高。谷氨酸棒杆菌用于氨基酸的工业生产已有数十年，最近经过工程改造，可用于生产N-甲基-L-丙氨酸和肌氨酸。 dpkA在为乙醛酸过量生产而工程化的谷氨酸棒杆菌菌株中的异源表达使得能够从糖和单甲胺发酵生产肌氨酸。在此，DpkA（DpkA F117L ）的底物结合位点的氨基酰基残基突变导致以单甲胺和单乙胺为底物的乙醛酸还原烷基胺化的比活增加。将DpkA F117L 引入生产菌株可加速肌氨酸的生产，可实现0.16 g L -1 h -1 的容积生产率。使用单乙胺作为底物，我们证明了N-乙基甘氨酸的生产，其容积生产率为0.11 g L -1 h -1 ，据我们所知，这是第一份报告其发酵生产。随后，测试了使用稻草水解物作为替代碳源的可行性，并在分批补料生物反应器培养60小时后，可生产N-乙基甘氨酸至滴定度为1.6 g L -1 。
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