Crystal structure of a constitutive active mutant of adenosine A2A receptor

Previously we reported a common activation pathway of the class A G protein-coupled receptors (GPCRs) in which a series of conserved residues/motifs undergo conformational change during extracellular agonist binding and finally induce the coupling of intracellular G protein, and successfully predicted several novel constitutive active or inactive mutations for A2A adenosine receptor (A2AAR) through this mechanism (Zhou et al., 2019). Here we determined the crystal structure of a typical A2AAR constitutive active mutant I92N in complex with agonist UK-432097 to reveal the molecular mechanism of mutation-induced constitutive activity. The mutated I92N forms a hydrophilic interaction network with nearby residues including W6.48 of the CWxP motif, which is absent in the wild type A2AAR. Although the mutant structure is overall similar to the previously determined intermediate state A2AAR structure (PDB ID: 3QAK), the molecular dynamics simulations suggested that the I92N mutant stabilizes the metastable intermediate state through the hydrophilic interaction network and favors the receptor conformational transition towards the active state. This research provides a structural template toward the special pharmacological outcome triggered by conformational mutation and sheds light on future structural or pharmacological studies among class A GPCRs.