Elimination of D-lactate synthesis increases poly(3-hydroxybutyrate) and ethanol synthesis from glycerol and affects cofactor distribution in recombinant Escherichia coli.
The effect of eliminating D-lactate synthesis in poly(3-hydroxybutyrate) (PHB)-accumulating recombinant Escherichia coli (K24K) was analyzed using glycerol as a substrate. K24KL, an ldhA derivative, produced more biomass and had altered carbon partitioning among the metabolic products, probably due to the increased availability of carbon precursors and reducing power. This resulted in a significant increase of PHB and ethanol synthesis and a decrease in acetate production. Cofactor measurements revealed that cultures of K24K and K24KL had a high intracellular NADPH content and that the NADPH/NADP(+) ratio was higher than the NADH/NAD(+) ratio. The ldhA mutation affected cofactor distribution, resulting in a more reduced intracellular state, mainly due to a further increase in NADPH/NADP(+). In 60-h fed-batch cultures, K24KL reached 41.9 g·liter⁻¹ biomass and accumulated PHB up to 63% ± 1% (wt/wt), with a PHB yield on glycerol of 0.41 ± 0.03 g·g⁻¹, the highest reported using this substrate.
Nikel
Pablo I
PI
Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Ciudad Universitaria-Pabellón 2, Av. Intendente Güiraldes 2160, C1428EGA, Buenos Aires, Argentina. jul@qb.fcen.uba.ar
Giordano
Andrea M
AM
de Almeida
Alejandra
A
Godoy
Manuel S
MS
Pettinari
M Julia
MJ
D018533:Biomass; D019149:Bioreactors; D002244:Carbon / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D000431:Ethanol / Q000378:metabolism*; D005990:Glycerol / Q000378:metabolism*; D006885:Hydroxybutyrates / Q000378:metabolism*; D045022:Lactate Dehydrogenases / Q000172:deficiency*; D019344:Lactic Acid / Q000378:metabolism*; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D009243:NAD / Q000032:analysis; D009249:NADP / Q000032:analysis; D011091:Polyesters / Q000378:metabolism*; D013997:Time Factors; D000094122:Polyhydroxybutyrates
D002244:Carbon; D000431:Ethanol; D005990:Glycerol; D006885:Hydroxybutyrates; D045022:Lactate Dehydrogenases; D019344:Lactic Acid; D009243:NAD; D009249:NADP; D011091:Polyesters; C003182:poly-beta-hydroxybutyrate; D000094122:Polyhydroxybutyrates