Structural basis of ligand recognition and design of antihistamines targeting histamine H(4) receptor.

The histamine H(4) receptor (H(4)R) plays key role in immune cell function and is a highly valued target for treating allergic and inflammatory diseases. However, structural information of H(4)R remains elusive. Here, we report four cryo-EM structures of H(4)R/G(i) complexes, with either histamine or synthetic agonists clobenpropit, VUF6884 and clozapine bound. Combined with mutagenesis, ligand binding and functional assays, the structural data reveal a distinct ligand binding mode where D94(3.32) and a π-π network determine the orientation of the positively charged group of ligands, while E182(5.46), located at the opposite end of the ligand binding pocket, plays a key role in regulating receptor activity. The structural insight into H(4)R ligand binding allows us to identify mutants at E182(5.46) for which the agonist clobenpropit acts as an inverse agonist and to correctly predict inverse agonism of a closely related analog with nanomolar potency. Together with the findings regarding receptor activation and G(i) engagement, we establish a framework for understanding H(4)R signaling and provide a rational basis for designing novel antihistamines targeting H(4)R.