1887

Abstract

Efflux pumps play a major role in multidrug resistance of pathogenic bacteria. The TolC homologue HI1462 was identified as the single channel-tunnel in required to form a functional multidrug efflux pump. The outer-membrane protein was expressed in , purified and reconstituted in black lipid membranes. It exhibited a comparatively small single-channel conductance of 43 pS in 1 M KCl and is the first known TolC homologue which is anion-selective. The HI1462 structure was modelled and an arginine residue lining the tunnel entrance was identified. The channel-tunnel of a mutant with the arginine substituted by an alanine residue was cation-selective and had a sevenfold higher single-channel conductance compared to wild-type. These results confirm that the arginine is responsible for anion selectivity and forms a salt bridge with a glutamate residue of the adjacent monomer, establishing a circular network, which keeps the tunnel entrance in a tightly closed conformation. In experiments, both the wild-type HI1462 and the mutant were able to substitute for TolC in the haemolysin secretion system, but not in the AcrAB/TolC multidrug efflux pump. The structure–function relationship of HI1462 is discussed in the context of the well-studied TolC channel-tunnel of .

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2006-06-01
2019-11-19
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