Abstract

The possible contribution of collateral enzymes to the formation of the key precursor metabolite, 3-hydroxybutyryl-CoA, has been evaluated in a recombinant Escherichia coli strain engineered for 1,3-butanediol biosynthesis from glucose via the inverted fatty-acid beta-oxidation pathway. Inactivation of the 3-hydroxyadipyl-CoA dehydrogenase gene, paaH, did not prevent 1,3-butanol biosynthesis during anaerobic glucose utilization by a strain with an intact, essential gene, fabG. This gene encodes 3-ketoacyl-ACP reductase, which can catalyze the conversion of acetoacetyl-CoA to (R)-3-hydroxybutyryl-CoA. The subsequent inactivation in the strain of the fadB gene, which encodes (S)-stereospecific 3-hydroxyacyl-CoA dehydrogenase of the fatty-acid beta oxidation led to the cessation of 1,3-butanediol synthesis. The respective diol was also not found among the products secreted by the strain possessing the intact fabG and paaH genes upon the individual deletion of the fadB gene. It was established that the collateral enzymes did not participate in the formation of 3-hydroxybutyryl-CoA in the studied strains, and the respective CoA derivative was synthesized solely by the (S)-specific enzyme of the fatty-acid beta-oxidation pathway. The results indicate that reversal of the fatty-acid beta oxidation pathway can ensure the enantioselective biosynthesis of the (S)-stereoisomer of 1,3-butanediol in engineered E. coli strains.

中文翻译：

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重组大肠杆菌菌株从葡萄糖立体选择性生物合成（S）-1,3-丁二醇的脂酸β-氧化途径逆转潜力的研究

摘要

已在重组大肠杆菌菌株中评估了辅助酶对关键前体代谢物3-羟基丁酰辅酶A形成的可能贡献，该菌株经过工程改造，可通过葡萄糖通过反向脂肪酸β-氧化途径从葡萄糖中合成1,3-丁二醇。3-羟基己二酰辅酶A脱氢酶基因paaH的失活不会阻止厌氧葡萄糖利用具有完整必需基因fabG的菌株进行1,3-丁醇生物合成。该基因编码3-酮酰基-ACP还原酶，其可以催化乙酰乙酰基-CoA向（R）-3-羟基丁酰基-CoA的转化。在的应变后续的失活是fadb基因，其编码（小号β-脂肪酸的β-立体特异性3-羟酰基-CoA脱氢酶导致1,3-丁二醇的合成停止。相应二醇也未由应变具有完整分泌的产品中发现fabG和paaH在所述的单个缺失的基因是fadb基因。已经确定，在所研究的菌株中，辅助酶不参与3-羟基丁酰-CoA的形成，并且分别通过脂肪酸β-氧化途径的（S）-特异性酶来合成相应的CoA衍生物。结果表明，脂肪酸β氧化途径的逆转可以确保（S）对映体的生物合成。工程化大肠杆菌菌株中1,3-丁二醇的）-立体异构体。