Understanding membrane proteins’ allosteric modulation with cryo-EM

Project Summary/Abstract Our laboratory is primarily focused on elucidating the molecular mechanisms integral to transmembrane receptor function and modulation of their signaling output. We employ cryo-electron microscopy (cryo-EM) in conjunction with advanced classification methodologies such as manifold embedding, modeling, and molecular dynamics to investigate how ligand binding influences conformational equilibria and signaling bias. Our research encompasses various systems, enabling us to examine the impact of different ligand types on ion channel gating and G protein-coupled receptor (GPCR) activation. Our longstanding research interest lies in receptors vital to heart and skeletal muscle function. We are investigating the mechanisms and modulation of ryanodine receptors (RyR) and beta-adrenergic receptors, which play a crucial role in cardiac and skeletal muscle functionality. Our objective is to elucidate how small molecules, protein regulators, and ions shape the conformational energy landscape of these receptors. This knowledge is critical for understanding calcium signaling regulation and for designing innovative therapeutics to address cardiac and muscle disorders. Another research interest is exploring lipid-triggered gating of mechanosensitive channels from the MscS family, a model system for membrane tension-sensing. This research can provide valuable insights into the fundamental principles governing ion channel gating and the role of lipid-protein interactions in this process. Our research approach combines cryo-EM, advanced image classification techniques, residue network analysis, and molecular dynamics simulations to mechanistically delineate allosteric pathways. We subsequently validate proposed models using biophysical techniques such as single-channel measurements, hydrogen-deuterium exchange mass spectrometry (HDX-MS), and mutagenesis. Our long-term goal is to significantly enhance our molecular understanding of receptor activation and allosteric modulation. Advancement in this field could potentially pave the way for the design of small molecule allosteric modulators with precise control over their effects on their targets, thus aiding in the development of drugs with minimized side effects. 1