Factors affecting proton translocation by facultative methylotrophs

The general properties of respiration-driven proton translocation by the two facultative methylotrophs, Pseudomonas AM1 and Pseudomonas extorquens, were similar to those of other bacteria. The stoichiometry of H⁺ extruded/O atom consumed ($\text{H}\text{←}{}^{\mathrm{+}}\text{O}$) during respiration with a particular substrate depended, however, on the concentration of the permeant anion SCN^? used to abolish the membrane potential and on the physiological state of the organism. This variability makes the use of proton translocation data of dubious value in the elucidation of electron-transport pathways, at least in these species, unless the physiological condition of the organisms can be accurately described and reproduced. Methanol oxidation was inhibited by SCN^? but substitution of valinomycin for most of the SCN^? during pulse oxidant experiments allowed measurement of proton translocation when methanol was the substrate. Starved organisms were used to eliminate ambiquity as to whether added test substrates or endogenous reserve materials were being oxidised. Viability remained high during starvation and endogenous O₂ uptake remained detectable long after endogenously driven proton translocation was undetectable. In the absence of endogenously driven proton translocation, measured $\text{H}\text{←}{}^{\mathrm{+}}\text{O}$ stoichiometries differed substantially from those when it was present, suggesting that the physiological state of the organisms is an essential parameter in assessing proton translocation data.