Recombinant expression of the Z. mobilis homoethanol pathway has been the cornerstone of E. coli ethanologenesis. However, recent progress has en­abled ethanol production by a mutant E. coli strain lacking foreign genes [46]. Due to the inability to regenerate NAD+ and maintain redox balance, wild — type E. coli is unable to grow anaerobically in the absence of both IdhA and pflB [47]. Chemical mutagenesis was used to isolate AldhA/ApflB deriva­tives capable of anaerobic growth. The resulting strain SE2378 fermented glucose and xylose to ethanol with 82% yield. Further analysis of SE2378 revealed an essential mutation within the pyruvate dehydrogenase (PDH) operon. In native strains, pyruvate formate-lyase is primarily responsible for production of acetyl-CoA during anaerobic growth; PDH is reportedly inactive [48] or weakly active [49] under these conditions. The essential mu­tation in the pdh operon restored function during anaerobic growth and produced an additional NADH for each pyruvate. This additional NADH al­lowed the balanced production of 2 moles of ethanol per mole of glucose by a novel pathway not previously known in nature. The anaerobic spe­cific growth rate of SE2378 was reduced approximately 50% relative to the parental strain in rich media and no growth was observed in glucose mini­mal media without acetate, glutamate, or corn steep liquor supplementation. Despite growth challenges, the maximum specific productivity of SE2378, 2.24 g ethanol h-1 gcells-1, is comparable to KO11 and ethanol was pro­duced from 50 g L-1 glucose and xylose at greater than 80% of the theoretical yield.

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