The protonmotive force and α-aminoisobutyric acid transport in an obligately alkalophilic bacterium

Measurements of the protonmotive force and studies of solute transport in B. alcalophilus were conducted to ascertain how cell processes are energized under conditions in which a pH gradient, interior alkaline, cannot be established across the membrane. B. alcalophilus grows on L-malate in a range of pH from 9.0 to 11.5, with an optimum at pH 10.5. The electrical potential across the membrane, Δ psi, increases progressively from -84 mV at pH 9.0 to -152 mV at pH 11.5. At pH 10.00 and above a ΔpH exists, interior acid, increasing from 36 mV at pH 10.0 to 151 mV at pH 11.5. At pH 10.5 and 11, at which growth is excellent, the total transmembrane protonmotive forces were only -80 mV and -15 mV, respectively. Uptake of α-aminoisobutyric acid (AIB), to concentrations 845-fold greater than the external concentration, occurs only at alkaline pH values, with an optimum at pH 10.5. Saturation kinetics, with a K(m) for AIB of 9.3 μM, were observed in the presence of sodium ions. Sodium ions are probably symported with AIB in an electrogenic transport that responds to the Δpsi: addition of sodium chloride and AIB to cells which are taking up triphenylmethylphosphonium⁺ results in a cessation of TPMP⁺ uptake; conversely, a variety of agents that abolish Δpsi inhibit sodium-dependent AIB uptake; and in cells in which the Δpsi has been dissipated (which occurs upon incubation in K₂CO₃ buffer containing ammonium sulfate), sodium chloride does not stimulate AIB uptake appreciably, but sodium thiocyanate does. The cellular levels of potassium are unusually low, and valinomycin inhibited both AIB and TPMP⁺ uptake even in the absence of external potassium. Similarly, carbonyl cyanide m-chlorophenylhydrazone caused an expected efflux of protons, but an abolition of Δpsi occurred concomitantly, rather than an increase in the Δpsi. The Δpsi might be established, at the highly alkaline pH of these studies, by a K⁺/H⁺ antiport such that agents which circumvent this system for transmembranal movements of either protons or potassium ions interfere with maintenance of the Δpsi.