Stochastic Switching Into Hydrolytically Active Conformations In A Homodimeric ABC Exporter

Abstract

ATP binding cassette (ABC) transporters are a large family of membrane proteins with high clinical relevance in, for example, bacterial multidrug resistance, tumor resistance, cystic fibrosis, or insulin secretion. Bacterial ABC exporters are homodimers in which each identical half contributes a transmembrane domain (TMD) and a nucleotide binding domain (NBD). Many mammalian ABC transporters, instead, consist of asymmetric halves. ABC transporters are thought to hydrolyze MgATP only at one of their two nucleotide binding sites at a time. In homodimeric ABC exporters, the process of switching one of the binding sites into a hydrolytic conformation ought to be stochastic. Recent evidence suggests that the asymmetry in the binding sites of various mammalian exporters induces a directional preference in their nucleotide hydrolysis that may improve the choreography of complex transport processes. Currently, it is poorly understood how exactly the switching of only one binding site into a hydrolytically favorable conformation occurs. Furthermore, it is mostly unknown how this conformational change is reflected at the NBD-TMD interface. In this study, we apply molecular dynamics simulations to probe the switching of the MgATP-bound bacterial multidrug exporter Sav1866 into pre-hydrolytic states. The simulations are performed of the full-length structure embedded in a phospholipid bilayer. Our simulations show that the switching in Sav1866 is of stochastic nature. We identify specific changes at the binding sites that characterize a pre-hydrolytic conformation, and show that the switching event causes pronounced changes in NBD-TMD interactions. We also extend our findings to asymmetric transporters and suggest mechanisms of directionality in the nucleotide handling of some mammalian ABC transporters.