Biophysical Society Thematic Meeting | Stockholm 2022

Physical and Quantitative Approaches to Overcome Antibiotic Resistance

Monday Speaker Abstracts

MOLECULAR DYNAMICS SIMULATIONS OF MEXB, MEXF AND MEXY MULTIDRUG TRANSPORTERS OF PSEUDOMONAS AERUGINOSA Andrea Catte 1 ; Venkata K Ramaswamy 1 ; Attilio V Vargiu 1 ; Giuliano Malloci 1 ; Paolo Ruggerone 1 ; 1 University of Cagliari, Department of Physics, Monserrato (Cagliari), Italy The secondary multidrug transporters of the resistance-nodulation-cell division (RND) superfamily mediate multi-drug resistance in Gram-negative bacteria like Pseudomonas aeruginosa. This pathogen expresses four main polyspecific RND transporters, namely MexB, MexD, MexF and MexY, with partly overlapping specificities. However, only the structure of the former has been resolved experimentally to date. The lack of data about the structure and the dynamics of most transporters has limited a systematic investigation of the molecular determinants defining their activities. In a previous work [Ramaswamy et al, Front. Microbiol. 9:1144 (2018)], we employed computational methods to compare the main putative recognition sites (named access and deep binding pockets, AP and DP respectively) in MexB and MexY. In this work, we expand the comparison by performing extended molecular dynamics simulations of MexB, MexY and MexF embedded in a more realistic model of the inner phospholipid membrane of P. aeruginosa, using updated force-fields and newly developed protocols. Moreover, to elucidate how the structures and the dynamics of these transporters define their substrate specificity, we conduct a comparative dynamic fragment-based mapping on these three proteins. In addition to analyzing the binding of probes on access and distal/deep binding pockets, we investigated for the first time the accumulation of fragments at various entrance and exit channels of each protein. Our results highlight similarity and differences in the distribution of multi-functional sites in the AP and DP of the three transporters binding sites. Moreover, our findings pinpoint a peculiar behavior of MexF vs. MexB/Y regarding the features of the entrance gates of the periplasmic and transmembrane channels. Altogether, our results allow to rationalize the partial redundancy and the specificities of the substrate profiles of the three Mex transporters.

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