Unveiling G-Protein-Coupled Receptor Conformational Dynamics via Metadynamics Simulations and Markov State Models

The dynamic character of G-protein-coupled receptors (GPCRs) is essential in their functionality as signal transducers. However, the molecular details of how ligands affect this conformational repertoire to steer intracellular signaling pathways remain elusive. To address this question, we present a generally applicable protocol that combines metadynamics simulations and Markov state modeling to compute the free energy landscape of the growth hormone secretagogue receptor (GHSR-1a), a prototypical class A GPCR, in its apo state and bound to pharmacologically distinct ligands. Consistent with the current multistate model of GPCR activity, we found that GHSR-1a populates multiple metastable states whose energies and transition probabilities change depending on the bound ligand. We identified intermediate states that have not yet been described by experimental structures and shed light on the molecular differences between basal and agonist-induced GHSR-1a activation. Our results are not only compatible with previously reported experimental data, but they capture the equilibria governing GHSR-1a activation in unprecedented detail. Due to its applicability to all class A GPCRs, our protocol is a valuable tool for the development of pharmaceuticals targeting this protein family.