Subunit swapping in the Mex-extrusion pumps in Pseudomonas aeruginosa

Pseudomonas aeruginosa encodes three sets of antibiotic extrusion proteins designated as MexA,B-OprM, MexC,D-OprJ and MexE,F-OprN regulated by the nalB, nfxB and nfxC genes, respectively. MexB,D,F, OprM,J,N and MexA,C,E function as the inner membrane pumps, the outer membrane channels and the membrane fusion proteins, respectively. To investigate the possibility of subunit interchangeabilities, we constructed the following combinations of chimeric pumps: MexA,D-OprM/ΔMexB, MexC,B-OprM/ΔMexA, and MexA,B-OprJ/ΔOprM. The strains producing MexA,D-OprM/ΔMexB and MexC,B-OprM/ΔMexA failed to restore the antibiotic resistance shown in the strains producing the natural combinations of the subunit proteins. These results suggested that the inner membrane components cannot be interchanged. In contrast, the stains producing MexA,B-OprJ/ΔOprM exhibited higher resistance to several antibiotics than the mutant lacking OprM and lower resistance than the strain overexpressing OprM. This result suggests that OprJ may complement the OprM function partially. A spectrum of antibiotics, of which the minimum inhibitory concentrations were restored partially by the complementation, was the same as the spectrum to which the nalB type mutant shows resistance. We surmised from these results that the MexA/MexB unit sustains the substrate specificity of the MexA,B-OprM machinery.