Understanding the molecular mechanism of endothelin ET_A receptor selecting isopeptides endothelin-1 and -3

Endothelin-1 and -3 (ET1 and ET3) are potent vasoconstricting isopeptides that are involved in the pathophysiology of various diseases, such as cardiovascular and renal diseases. The two ETs exert their effects by binding to two G-protein-coupled receptors (GPCRs), the endothelin ET_A receptor (ET_AR) and the endothelin ET_B receptor (ET_BR). ET_AR and ET_BR reveal different preferences in the recognition of the two ETs: The binding affinity of ET_AR with ET1 is much higher than that with ET3, while the binding affinities of ET_BR with the two ETs are similar. Recently, the structures of ET_BR with ET1 and ET3 were determined. These crystal structures provide detailed descriptions of how ET_BR interacts with ET1 and ET3. However, the knowledge of how ET_AR recognizes the two isopeptides is still lacking. Based on the structure of ET_BR in complex with ET1, the structures of ET_AR with ET1 and ET3 were modeled. Then, molecular dynamics simulations were performed to study how ET_AR discriminates the two ETs. Simulation results demonstrate that ET1 has greater binding free energy with ET_AR than ET3, which is in agreement with the binding affinity data of ET_AR. By interaction energy analysis, the key residues of ET_AR that discriminate between ET1 and ET3 are identified. Structural and dynamical analyses indicate that, compared with ET3, ET1 is more stable in binding with ET_AR. Furthermore, the orientation change of W319 in the conserved CWxP motif that plays an important role in the signaling function of GPCRs was observed. Through the orientation change of this residue, the difference in the orthosteric pocket volume resulting from the binding of ET1 and ET3 on the extracellular side of ET_AR leads to a conformational difference of TM6 on the intracellular side of ET_AR. This study elucidates the molecular mechanism of how ET_AR selects the isopeptides ET1 and ET3.