Escherichia coli FocA/B-dependent H⁺ and K⁺ fluxes: Influence of exogenous versus endogenous formate

Escherichia coli translocates formate/formic acid bidirectionally across the cytoplasmic membrane by the FocA/FocB formate channels during fermentation. Depending on the pH and whether formate is supplied exogenously or generated internally, the mechanisms of translocation differ. This study elucidates the role of these channels in dependence on F_OF₁ ATPase activity in stationary phase cells after cultivation by mixed-carbon fermentation at pH 7.5. In cells cultivated with glucose plus glycerol, exogenously added formate increased the N,N'-dicyclohexylcarbodiimide (DCCD)-sensitive (F_OF₁ ATPase-dependent) proton flux in single or double foc mutants. Moreover, exogenously supplied formate also increased the DCCD-sensitive potassium flux, but only in mutants where focB was absent. In the cells grown on glucose, glycerol, and formate, addition of formate in the whole-cell assays increased F_OF₁ ATPase activity by ∼60% compared with cells grown on a mixture of only glucose and glycerol. In a focA mutant cultivated to the stationary phase on glucose, glycerol, and formate, F_OF₁ ATPase activity was double that compared with cells grown on only glucose and glycerol, while in a focA-focB double-null mutant F_OF₁ ATPase activity decreased by ∼50% in formate assays. These data suggest that the cell regulates the mechanism of formate translocation depending on whether formate is generated internally or added exogenously. Thus, F_OF₁-ATPase activity and the FocA/FocB channels together with formate hydrogenlyase activity combine to balance pH and ion gradients during fermentation in stationary phase cells in response to whether formate is generated metabolically or supplied in high concentration from the environment.