Differential permeability for lipophilic compounds in uncoupler-resistant cells of Escherichia coli

The acrA strain AS-1 of Escherichia coli is more sensitive than its parent W3110 to growth inhibition by Methylene blue, sodium dodecyl sulfate and novobiocin. UR-3 is an uncoupler-resistant strain isolated from AS-1 which is resistant to growth inhibition by carbonylcyanide m-chlorophenylhydrazone (CCCP), 3,3',4',5-tetrachlorsalicylanilide (TCS) and tributyltin chloride, while remaining sensitive to the first group of compounds. A revertant of AS-1 acquired resistance to Methylene blue and sodium dodecyl sulfate but remained sensitive to uncouplers. In contrast to AS-1, proline uptake in UR-3 was resistant to uncouplers. Strain UR-3 grown in the presence of uncoupler incorporated elongation factor Tu to high levels in the outer membrane of the cell. A role for the outer membrane in the acquisition of uncoupler-resistance by UR-3 is suggested by the behaviour of the mutant to the fluorescence probe N-phenyl-1-naphthylamine. The fluorescence intensity of this probe was quenched by membrane energization in the wild-type strain W3110 but not in AS-1. UR-3 behaved like W3110, suggesting that an outer membrane barrier to neutral lipophilic compounds like N-phenyl-1-naphthylamine (NPN) and uncouplers had been restored in UR-3. By contrast, AS-1 and UR-3 both allowed energized uptake of the fluorescent lipophilic cation 2-(dimethylaminostyryl)-1-ethylpyridinium (DMP+). It is concluded that lipophilic materials must permeate the outer membrane of E. coli by at least two different routes. However, uncoupler-resistance in UR-3 appears to be more complex than the provision of an outer membrane barrier to uncouplers. Thus, uncouplers readily discharged a pH gradient established in both AS-1 and UR-3 by addition of HCl to cell suspensions.