Molecular Determinantsof Ligand Binding Modes inthe Histamine H4Receptor: Linking Ligand-Based Three-DimensionalQuantitative Structure–Activity Relationship (3D-QSAR) Modelsto in Silico Guided Receptor Mutagenesis Studies.

The histamine H4receptor (H4R)is a G protein-coupledreceptor (GPCR) that plays an important role in inflammation. Similarto the homologous histamine H3receptor (H3R),two acidic residues in the H4R binding pocket, D3.32and E5.46, act as essential hydrogen bond acceptors ofpositively ionizable hydrogen bond donors in H4R ligands.Given the symmetric distribution of these complementary pharmacophorefeatures in H4R and its ligands, different alternativeligand binding mode hypotheses have been proposed. The current studyfocuses on the elucidation of the molecular determinants of H4R–ligand binding modes by combining (3D) quantitativestructure–activity relationship (QSAR), protein homology modeling,molecular dynamics simulations, and site-directed mutagenesis studies.We have designed and synthesized a series of clobenpropit (N-(4-chlorobenzyl)-S-[3-(4(5)-imidazolyl)propyl]isothiourea)derivatives to investigate H4R–ligand interactionsand ligand binding orientations. Interestingly, our studies indicatethat clobenpropit (2) itself can bind to H4R in two distinct binding modes, while the addition of a cyclohexylgroup to the clobenpropit isothiourea moiety allows VUF5228 (5) to adopt only one specific binding mode in the H4R binding pocket. Our ligand-steered, experimentally supported proteinmodeling method gives new insights into ligand recognition by H4R and can be used as a general approach to elucidate the structureof protein–ligand complexes. [ABSTRACT FROM AUTHOR]