Regulation mechanisms in a Cation/Proton Antiporter (CPA)

We are studying a K+/H+ antiporter as a model system to understand the mechanism of regulation in cation/proton antiporter (CPA) superfamily. We have preciously performer a detailed biochemical characterization of this antiporter, which provided insights into how transport activity is inhibited or activated. Recently, we determined the structure of its inactive conformation, elucidating the molecular basis of transport inhibition. However, the mechanism underlying activation remains unclear. The goal of this proposal is to determine the active conformation(s) of the antiporter. Under the selected experimental conditions, we expect that most of the molecules will be in one or more conformations adopted during K⁺ transport, allowing us to reconstruct the full transport cycle. In order to describe this conformational landscape, large number of particles and high resolution data will be required, for which data collection in a Titan Krios will be essential.