A Highly Conserved NA+ Binding Site in Prokaryotic Multi-Drug Mate Transporters

Abstract

Multi-drug and Toxic-compound Extrusion (MATE) proteins are a recently recognized and largely uncharacterized family of secondary-active transporters. Present in both eukaryotes and prokaryotes, MATE transporters protect the cell by catalyzing the efflux of a broad range of cytotoxic compounds, including man-made antibiotics and anti-cancer drugs. These transporters are therefore potential pharmacological targets against drug-resistant pathogenic bacteria and tumor cells. The efflux activity of MATE transporters is powered by a transmembrane electrochemical ion gradient, but their molecular mechanism and ion-specificity are not well understood, in part due to the scarcity of high-quality structural information. Here, we use computational methods to study PfMATE, from the archaeon Pyrococcus furiosus, whose structure is the best resolved among all available. Seemingly at odds with the notion that this transporter is solely coupled to H+, our analysis of the original crystallographic data and additional molecular dynamics simulations unequivocally point to the existence of Na+-binding site in the N-lobe of this transporter. Moreover, we find this site to be broadly conserved among prokaryotic MATEs, including members of the family known to be coupled to Na+ gradients, such as NorM and ClbM, for which a Na+-binding site had not been conclusively identified. We further posit this site in the N-lobe may mediate H+ coupling instead at sufficiently low Na+ concentrations, as has been noted for other Na+-dependent transport systems. In summary, our study provides new insights into the structural basis for the complex ion dependency of MATE transporters.