Pyruvate oxidase contributes to the aerobic growth efficiency of Escherichia coli

The metabolic importance of pyruvate oxidase (PoxB), which converts pyruvat e directly to acetate and CO2, was assessed using an isogenic set of geneti cally engineered strains of Escherichia coli, In a strain lacking the pyruv ate dehydrogenase complex (PDHC), PoxB supported acetate-independent aerobi c growth when the poxB gene was expressed constitutively or from the IPTG-i nducible tac promoter. Using aerobic glucose-limited chemostat cultures of PDH-null strains, it was found that steady-states could be maintained at a low dilution rate (0.05 h(-1)) when PoxB is expressed from its natural prom oter, but not at higher dilution rates (up to at least 0.25 h(-1)) unless e xpressed constitutively or from the tac promoter. The poor complementation of PDH-deficient strains by poxB plasmids was attributed to several factors including the stationary-phase-dependent regulation of the natural poxB pr omoter and deleterious effects of the multicopy plasmids, As a consequence of replacing the PDH complex by PoxB, the growth rate (mu (max)), growth yi eld (Y-max) and the carbon conversion efficiency (flux to biomass) were low ered by 33 %, 9-25% and 29-39 % (respectively), indicating that more carbon has to be oxidized to CO2 for energy generation. Extra energy is needed to convert PoxB-derived acetate to acetyl-CoA for further metabolism and enzy me analysis indicated that acetyl-CoA synthetase is induced for this purpos e. In similar experiments with a PoxB-null strain it was shown that PoxB no rmally makes a significant contribution to the aerobic growth efficiency of E, coli, In glucose minimal medium, the respective growth rates (mu (max)) . growth yields (Y-max) and carbon conversion efficiencies were 16%. 14% an d 24% lower than the parental values, and correspondingly more carbon was f luxed to CO2 for energy generation. It was concluded that PoxB is used pref erentially at low growth rates and that E, coli benefits from being able to convert pyruvate to acetyl-CoA by a seemingly wasteful route via acetate.