Efflux and the steady state in alpha-methylglucoside transport in Escherichia coli

Efflux and the steady state in a group translocation system, the alpha-methylglucoside (alphaMG) transport system, were investigated. The maximum intracellular level of alpha-methylglucoside is a function of a steady state. There is no inhibition of alphaMG influx as the intracellular pool of alphaMG, and alpha-methylglucoside-6-phosphate (alphaMGP) rises. This steady state has three components: alphaMG influx, action of an alphaMGP phosphatase, and alphaMG efflux. The phosphatase is the rate-limiting step (half-time = 5.0 min); thus, the true efflux rate (half-time = 2.0 min) cannot be simply measured from the kinetics of alphaMG loss from the cell. Under our steady-state conditions the percentage of intracellular radioactivity present as alphaMGP was 71%. Under conditions of zero influx, after an efflux of 12 min the percentage present as alphaMGP fell to 55%. However, when fluoride was present during the efflux period, the percentage of the sugar as alphaMGP increased to about 85%. Fluoride greatly inhibits both influx and phosphatase activity (half-time = 50 min). The efflux of alphaMG from the cell is apparently also fluoride-sensitive but to a lesser extent (half-time = 4.1 min). These data are summarized in a model describing the three components of the steady-state and effect of fluoride.