Monday, August 11, 2008
P112

The ATP Limitation in a Pyruvate Formate Lyase mutant of Escherichia coli Increases Glycolytic Flux to D-Lactate

José Utrilla, Guillermo Gosset, and Alfredo Martinez. Instituto de Biotecnología, UNAM, Cuernavaca, 62210, Mexico

Derivatives of Escherichia coli MG1655 were constructed for D-Lactate production by deleting the pflB, adhE and frdA genes. Due to the inactivation of the main acetyl-CoA production pathway, pfl mutants grew one third slower than its parental strain in mineral media-glucose. The glycolytic flux was greatly improved in strain CL3 (MG1655: Δpfl, ΔadhE, ΔfrdA), and lactate yield was 95% of the theoretical. The cell mass was up to 1.2 g/L and the volumetric productivity of D-lactate with CL3 was 1.6 g/(L.h), one of the highest volumetric productivity reported for D-lactate producing E. coli strains from glucose. The ATP yield from glucose to lactate is very low in pflB mutans (2 ATP/Glucose), thus CL3 has to improve its glucose uptake rate in order to fulfill the ATP needs to grow. In contrast to E. coli B and W3110, MG1655 derivatives are able to grow in mineral media with glucose as the sole carbon source. It is suggested that the better performance, in terms of biomass formation and lactate production rate, of the MG1655 pfl mutants is due to a leaky expression of the pyruvate dehydrogenase complex, that channel some pyruvate flux to acetyl-CoA, and that this flux is able to fulfill the acetyl-CoA pool required to support CL3 growth.


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