Giovanni Bonaiuti
Effects of Point Mutations on the Activity of AcrB
RUGGERONE, PAOLO;VARGIU, ATTILIO VITTORIO;
2012-01-01
Abstract
The tripartite efflux pump AcrAB-TolC is responsible for the intrinsic and acquired multidrug resistance in Escherichia coli. Its active part, the homotrimeric transporter AcrB, is in charge of the selective binding of substrates and energy transduction. The presence of several phenilalanine residues in the only binding pocket identified by X-ray has inspired an experimental work where the effects of single point Phe->Ala mutations on the MIC and on the efflux of several antibiotics have been determined [1,2]. Concerning the MICs, interestingly, the mutation F610A has been shown to significantly reduce the minimum inhibitory concentration of doxorubicin and many other substrates, although F610 does not appear to interact strongly with them or to be involved in the squeezing mechanism of the binding pocket suggested as preliminary step of the extrusion process. In this work, we assess the impact of the experimental mutations on the functionality of AcrB by means of computational techniques, using doxorubicin and minocycline as substrates. We found that for F610A the compounds slide deeply inside the binding pocket after mutation, increasing the strength of the interaction. During subsequent conformational alterations of the transporter, the substrates were either not extruded from the binding site or displaced along a direction other than the one associated with extrusion. The other mutations are not able to modify the binding affinity of the substrates, which are kept in positions similar to the ones assumed in the wild type protein. Our study indicates how subtle interactions determine the functionality of multidrug transporters, since decreased transport might not be simplistically correlated to decreased substrate binding affinity. [1] Bohnert, J. A., et al. J. Bacteriol. 2008, 190, 8225-9. [2] Bohnert, J. A.; Schuster, S.; Szymaniak-Vits, M.; Kern, W. V. PlosOne 2011, 6, e21196.Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.