Abstract
There are species differences between human histamine H1 receptor (hH1R) and guinea pig (gp) histamine H1 receptor (gpH1R) for phenylhistamines and histaprodifens. Several studies showed participation of the second extracellular loop (E2-loop) in ligand binding for some G protein-coupled receptors (GPCRs). Because there are large species differences in the amino acid sequence between hH1R and gpH1R for the N terminus and E2-loop, we generated chimeric hH1Rs with gp E2-loop (hgpE2H1R) and gp N terminus and gp E2-loop (hgpNgpE2H1R). hH1R, gpH1R, and chimeras were expressed in Sf9 insect cells. [3H]Mepyramine binding assays and steady-state GTPase assays were performed. In the series hH1R > hgpE2H1R > hgpNgpE2H1R, we observed a significant decrease in potency of histamine 1 in the GTPase assay. For phenoprodifen 5 and the chiral phenoprodifens 6R and 6S, a significant decrease in affinity and potency was found in the series hH1R > hgpE2H1R > hgpNgpE2H1R. In addition, we constructed new active-state H1R models based on the crystal structure of the human β2-adrenergic receptor (hβ2AR). Compared with the H1R active-state models based on the crystal structure of bovine rhodopsin, the E2-loop differs in its contact to the ligand bound in the binding pocket. In the bovine rhodopsin-based model, the backbone carbonyl of Lys187 (gpH1R) interacts with large histaprodifens in the binding pocket, but in the hβ2AR-based model, Lys187 (gpH1R) is located distantly from the binding pocket. In conclusion, the differences in N terminus and E2-loop between hH1R and gpH1R exert an influence on affinity and/or potency for histamine and phenoprodifens 5, 6R, and 6S.
Footnotes
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This work was supported by the Research Training Program (Graduiertenkolleg) GRK760 “Medicinal Chemistry: Molecular Recognition–Ligand-Receptor Interactions” of the Deutsche Forschungsgemeinschaft.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.108.140913.
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ABBREVIATIONS: GPCR, G protein-coupled receptor; H1R, histamine H1 receptor; gp, guinea pig; hH1R, human H1R; gpH1R, guinea pig H1R; E2-loop, second extracellular loop; hgpE2H1R, hH1R with gp E2-loop; hgpNgpE2H1R, hH1R with gp N terminus and gp E2-loop; RGS4, regulator of G protein signaling 4; h, human; TM, transmembrane domain; S, signal peptide; F, FLAG epitope; PCR, polymerase chain reaction; DMSO, dimethyl sulfoxide; hβ2AR, human β2-adrenergic receptor; MD, molecular dynamics; [3H]MEP, mepyramine; Cpd, compound; HA, histamine.
- Received May 7, 2008.
- Accepted June 23, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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