Abstract

The microaerobic synthesis of 3-hydroxybutyric acid by the Escherichia coli strain BOX3.1 ∆4 P_L-atoB P_L-tesB (MG1655 lacI^Q, ∆ackA-pta, ∆poxB, ∆ldhA, ∆adhE, ∆fadE, P_L-SD_phi10-atoB, P_trc-ideal-4-SD_phi10-fadB, P_L-SD_phi10-tesB), which was previously directly engineered for the biosynthesis of the target compound from glucose through the reversed fatty acid β-oxidation pathway, was studied. A target product yield of 0.12 mol/mol was achieved. Inactivation of the nonspecific YciA thioesterase gene in the strain led to an increase in the yield of 3-hydroxybutyric acid to 0.15 mol/mol. For the optimization of biosynthesis of target product the strain MG∆4 P_L-tesB (MG1655 ∆ackA-pta, ∆poxB, ∆ldhA, ∆adhE, P_L-SD_phi10-tesB) was engineered, and the genes encoding key enzymes of fatty acid β-oxidation were overexpressed in the strain from the plasmid pMW118m-atoB-fadB. The level of microaerobic synthesis of 3-hydroxybutyric acid by the strain MG∆4 P_L-tesB (pMW118m-atoB-fadB) achieved in primary evaluation conditions reached 0.35 mol/mol. Inactivation in the strain of the gene of nonspecific thioesterase YciA led to only minor decrease in acetate byproduction. Further inactivation in the strain of gene encoding nonspecific thioesterase YdiI had virtually no effect on the level of synthesis of side products. Cultivation of the constructed strain MG∆4 P_L-tesB ∆yciA (pMW118m-atoB-fadB) in bioreactor under the controlled conditions ensured achievement of a yield of 3‑hydroxybutyric acid amounting to 0.75 mol/mol.

中文翻译：

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重组大肠杆菌通过逆脂肪酸β-氧化途径优化葡萄糖（S）-3-羟基丁酸的生物合成

摘要

的3-羟基丁酸由所述微需氧合成大肠杆菌菌株BOX3.1Δ4P_大号-的atoB P_大号- TESB（MG1655的lacI ^Q，Δ ackA-PTA，Δ POXB，Δ的ldhA，Δ的adhE，Δ褪色，P_大号-SD _phi10 - atoB，P _trc-理想-4 -SD _phi10 - fadB，P _L -SD _phi10 - tesB），这是以前通过反向脂肪酸β-氧化途径从葡萄糖生物合成目标化合物而直接设计的。目标产物的产率为0.12mol / mol。菌株中非特异性YciA硫酯酶基因的失活导致3-羟基丁酸的产率增加至0.15mol / mol。为了优化目标产物的生物合成，菌株MG∆4 P _L - tesB（MG1655 ∆ ackA-pta，∆ poxB，∆ ldhA，∆ adhE，P _L -SD _phi10 - tesB）进行了工程改造，从质粒pMW118m- atoB - fadB的菌株中过表达了编码脂肪酸β-氧化关键酶的基因。在初步评估条件下，菌株MG∆4 P _L - tesB（pMW118m- atoB - fadB）产生的3-羟基丁酸微好氧合成水平达到0.35 mol / mol。非特异性硫酯酶YciA基因菌株的失活仅导致乙酸的少量减少。编码非特异性硫酯酶YdiI的基因菌株中的进一步失活实际上对副产物的合成水平没有影响。菌株MG∆4 P _L - tesB的培养在受控条件下，在生物反应器中的∆ yciA（pMW118m- atoB - fadB）确保获得3-羟基丁酸的产率为0.75 mol / mol。