Combinatorial Metabolic Engineering Strategies for the Enhanced Production of Free Fatty Acids in Escherichia coli.

In this study, we evaluated the effects of several metabolic engineering strategies in a systematic and combinatorial manner to enhance the free fatty acid (FFA) production in Escherichia coli. The strategies included (i) overexpression of mutant thioesterase I ('TesAR64C) to efficiently release the FFAs from fatty acyl-ACP; (ii) coexpression of global regulatory protein FadR; (iii) heterologous expression of methylmalonyl-CoA carboxyltransferase and phosphoenolpyruvate carboxylase to synthesize fatty acid precursor molecule malonyl-CoA; and (iv) disruption of genes associated with membrane proteins (GusC, MdlA, and EnvR) to improve the cellular state and export the FFAs outside the cell. The synergistic effects of these genetic modifications in strain SBF50 yielded 7.2 ± 0.11 g/L FFAs at the shake flask level. In fed-batch cultivation under nitrogen-limiting conditions, strain SBF50 produced 33.6 ± 0.02 g/L FFAs with a productivity of 0.7 g/L/h from glucose, which is the maximum titer reported in E. coli to date. Combinatorial metabolic engineering approaches can prove to be highly useful for the large-scale production of FA-derived chemicals and fuels.

Repeated batch production of epothilone B by immobilized Sorangium cellulosum.

Production of extracellular epothilone B, one of the potent anticancer agents, by free and immobilized Sorangium cellulosum was studied using the repeated batch culture process. The concentration of alginate used in immobilization was directly related to the mass transfer rate of nutrients, mechanical stability, and the epothilone B production yield. With the optimized 3% (w/v) calcium alginate carrier, a prolonged repeated batch culture was investigated for the 5 repeated batches for 24 days. The maximum productivity of epothilone B obtained from the alginate-immobilized cells was 5.03 mg/l/day, which is 3 times higher than that of free cells (1.68 mg/l/day).