Hydrogen-deuterium exchange coupled to mass spectrometry: A multifaceted tool to decipher the molecular mechanism of transporters.

Transporters regulate trafficking through the biological membrane of living cells and organelles. Therefore, these proteins play an important role in key cellular processes. Obtaining a molecular-level description of the mechanism of transporters is highly desirable to understand and modulate such processes. Different challenges currently complicate this effort, mostly due to transporters' intrinsic properties. They are dynamic and often averse to in vitro characterization. The crossing of the membrane via a transporter depends on both global and local structural changes that will enable substrate binding from one side of the membrane and release on the other. Dedicated approaches are required to monitor these dynamic changes, ideally within the complex membrane environment. Hydrogen-deuterium exchange coupled to mass spectrometry (HDX-MS) has recently emerged as a powerful biophysical tool to understand transporters' mechanism. This mini-review aims to offer to the reader an overview of the field of HDX-MS applied to transporters. It first summarizes the current workflow for HDX-MS measurements on transporters. It then provides illustrative examples on the molecular insights that are accessible thanks to the technique; following conformational transitions between different states, observing structural changes upon ligand binding and finally understanding the role of lipid-protein interactions.