Suppression of Hypersensitivity of Escherichia coli acrB Mutant to Organic Solvents by Integrational Activation of the acrEF Operon with the IS 1 or IS 2 Element

Abstract

ABSTRACT The AcrAB-TolC efflux pump plays an intrinsic role in resistance to hydrophobic solvents in Escherichia coli. E. coli OST5500 is hypersensitive to solvents due to inactivation of the acrB gene by insertion of IS 30 . Suppressor mutants showing high solvent resistance were isolated from OST5500. These mutants produced high levels of AcrE and AcrF proteins, which were not produced in OST5500, and in each mutant an insertion sequence (IS 1 or IS 2 ) was found integrated upstream of the acrEF operon, coding for the two proteins. The suppressor mutants lost solvent resistance on inactivation of the acrEF operon. The solvent hypersensitivity of OST5500 was suppressed by introduction of the acrEF operon with IS 1 or IS 2 integrated upstream but not by introduction of the operon lacking the integrated IS. It was concluded that IS integration activated acrEF , resulting in functional complementation of the acrB mutation. The acrB mutation was also complemented by a plasmid containing acrF or acrEF under the control of P lac . The wild-type tolC gene was found to be essential for complementation of the acrB mutation by acrEF . Thus, it is concluded that in these cells a combination of the proteins AcrA, AcrF, and TolC or the proteins AcrE, AcrF, and TolC is functional in solvent efflux instead of the AcrAB-TolC efflux pump.