Distinct interaction of human and guinea pig histamine H-2-receptor with guanidine-type agonists

It is unknown why the potencies and efficacies of long-chained guanidine-ty pe histamine H-2-receptor (H2R) agonists are lower at the H2R of human neut rophils than at the H2R of the guinea pig atrium. To elucidate these differ ences, we analyzed fusion proteins of the human H2R (hH(2)R) and guinea pig H2R (gpH(2)R), respectively, and the short splice variant of G(s alpha) (G (s alphaS)) expressed in Sf9 cells, The potencies and efficacies of small H 2R agonists in the GTPase assay and the potencies of antagonists at inhibit ing histamine-stimulated GTP hydrolysis by hH(2)R-G(s alphaS) and gpH(2)R-G (s alphaS) were similar. In contrast, the potencies and efficacies of guani dines were lower at hH(2)R-G(s alphaS) than at gpH(2)R-Gs alphaS. Guanidine s bound to hH(2)R-G(s alphaS) with lower affinity than to gpH(2)R-G(s alpha S), and high-affinity binding of guanidines at gpH(2)R-G(s alphaS) was more resistant to disruption by GTP-gammaS than binding at hH(2)R-Gs alphaS. Mo lecular modeling suggested that the nonconserved Asp-271 in transmembrane d omain 7 of gpH(2)R (Ala-271 in hH(2)R) confers high potency to guanidines. This hypothesis was confirmed by Ala-271 --> Asp-271 mutation in hH(2)R-G(s alphaS). Intriguingly, the efficacies of guanidines at the Ala-271 --> Asp -271 mutant and at hH(2)R/gpH(2)R chimeras were lower than at gpH(2)R. Our model suggests that a Tyr-17/Asp-271 H-bond, present only in gpH(2)R-G(s al phaS) but not the other constructs studied, stabilizes the active guanidine -H2R state. Collectively, our data show 1) distinct interaction of H2R spec ies isoforms with guanidines, 2) that a single amino acid in transmembrane domain 7 critically determines guanidine potency, and 3) that an interactio n between transmembrane domains 1 and 7 is important for guanidine efficacy .