Optimization of (S)-3-Hydroxybutyric Acid Biosynthesis from Glucose through the Reversed Fatty Acid β-Oxidation Pathway by Recombinant Escherichia coli Strains

The microaerobic synthesis of 3-hydroxybutyric acid by the Escherichia coli strain BOX3.1 increment 4 P-L-atoB P-L-tesB (MG1655 lacI(Q), increment ackA-pta, increment poxB, increment ldhA, increment adhE, increment fadE, P-L-SDphi10-atoB, Ptrc-ideal-4-SDphi10-fadB, P-L-SDphi10-tesB), which was previously directly engineered for the biosynthesis of the target compound from glucose through the reversed fatty acid beta-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 increment 4 P-L-tesB (MG1655 increment ackA-pta, increment poxB, increment ldhA, increment adhE, P-L-SDphi10-tesB) was engineered, and the genes encoding key enzymes of fatty acid beta-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 increment 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 increment 4 P-L-tesB increment 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.