Abstract

Anaerobic production of pyruvic acid from glucose by recombinant Escherichia coli strains with impaired fermentation ability during respiration with nitrate as an external terminal electron acceptor was studied. During nitrate respiration in a minimal salt medium lacking ammonium ions, the core E. coli strain MG1655 ∆ackA-pta, ∆poxB, ∆ldhA, ∆adhE, ∆ptsG, P_Lglk, P_tacgalP, ∆frdAB, ∆pflB, ∆sdhAB, ∆aceEF converted glucose into pyruvic acid with a yield of 1.72 mol/mol, secreting lactic acid as the only detected byproduct. The deletion of the lldD and dld genes blocked the secretion of this byproduct. The corresponding strain lacking the respiratory L- and D-lactate dehydrogenases LldD and Dld synthesized pyruvic acid from glucose with a yield of 1.76 mol/mol, consuming the available carbohydrate substrate incompletely. Enforced ATP hydrolysis due to the action of the pyruvic acid–oxaloacetic acid–malic acid–pyruvic acid or pyruvic acid–phosphoenolpyruvate–pyruvic acid futile cycles led to a drastic increase in glucose consumption by recombinants while maintaining the levels of substrate to the target product conversion. As a result, during anaerobic nitrate respiration and enforced ATP hydrolysis pyruvic acid was produced from glucose with a yield of 1.77–1.78 mol/mol with almost exhaustive consumption of the substrate by recombinants and no or minimal byproduct formation.

中文翻译：

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发酵能力受损的重组大肠杆菌菌株通过强制 ATP 水解优化葡萄糖厌氧生产丙酮酸

摘要

研究了在呼吸过程中发酵能力受损的重组大肠杆菌菌株从葡萄糖厌氧生产丙酮酸，硝酸盐作为外部终端电子受体。期间以极限盐培养基缺乏铵离子硝酸盐呼吸，核心大肠杆菌菌株MG1655Δ ackA-PTA，Δ POXB，Δ的ldhA，Δ的adhE，Δ PTSG，P_大号GLK，P _tac启动的galP，Δ frdAB，Δ PFLB， ∆ sdhAB , ∆ aceEF将葡萄糖转化为丙酮酸，产率为 1.72 mol/mol，分泌乳酸作为唯一检测到的副产物。lldD和dld的删除基因阻止了这种副产物的分泌。缺乏呼吸性 L- 和 D-乳酸脱氢酶 LldD 和 Dld 的相应菌株从葡萄糖合成丙酮酸，产量为 1.76 mol/mol，不完全消耗可用的碳水化合物底物。由于丙酮酸 - 草酰乙酸 - 苹果酸 - 丙酮酸或丙酮酸 - 磷酸烯醇丙酮酸 - 丙酮酸无效循环的作用，强制 ATP 水解导致重组体的葡萄糖消耗急剧增加，同时保持目标产物的底物水平转换。结果，在厌氧硝酸盐呼吸和强制 ATP 水解期间，丙酮酸由葡萄糖产生，产量为 1.77-1.78 mol/mol，重组体几乎耗尽底物，没有或几乎没有副产物形成。