Similar apparent constitutive activity of human histamine H-2-receptor fused to long and short splice variants of G(s alpha)

Wenzel-Seifert, Katharina and Kelley, Melissa T. and Buschauer, Armin and Seifert, Roland (2001) Similar apparent constitutive activity of human histamine H-2-receptor fused to long and short splice variants of G(s alpha). JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS, 299 (3). pp. 1013-1020. ISSN 0022-3565

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Abstract

Fusion proteins allow for the analysis of receptor/G protein coupling underdefined conditions. The beta (2)-adrenoceptor (beta (2)AR) fused to the long splice variant of G(s alpha) (G(s alphaL)) exhibits a higher apparent constitutive activity than the beta (2)-adrenoceptor fused to the short splice variant of G(s alpha) (G(s alphaS)). Experimentally, this results in higher efficacy and potency of partial agonists and in higher efficacy of inverse agonists at the beta (2)AR fused to G(s alphaL) relative to the beta (2)AR fused to G(s alphaS), indicating that the agonist-free beta (2)AR and the beta (2)AR occupied by partial agonists promote GDP dissociation from G(s alphaL) more efficiently than from G(s alphaS). In fact, the GDP affinity of G(s alphaS) fused to the beta (2)AR is higher than the GDP affinity of G(s alphaL) fused to the beta (2)AR. We asked the question whether the histamine H-2-receptor (H2R) exhibits similar coupling to G(s alpha) splice variants as the beta (2)AR. To address this question, we studied H2R-G(s alpha) fusion proteins expressed in Sf9 cells. In contrast to beta (2)AR-G(s alpha) fusion proteins, the potencies and efficacies of partial agonists and the efficacies of inverse agonists were similar at the H2R fused to G(s alphaL) and G(s alphaS) as assessed by guanosine-5'-O-(3-thio)triphosphate binding and/or steady-state GTPase activity. However, the time course analysis of guanosine-5'-O-(3-thio)triphosphate binding indicated that G(s alphaS) fused to the H2R possesses a higher GDP-affinity than G(s alphaL) fused to the H2R. Our data show that the H2R fused to G(s alphaL) and G(s alphaS) possesses similar constitutive activity and is insensitive to differences in GDP affinity of G(s alpha) splice variants. Thus, GDP affinity of G proteins does not generally determine constitutive activity of receptors.

Item Type: Article
Uncontrolled Keywords: MULTIPLE RECEPTOR CONFORMATIONS; PROTEIN-COUPLED RECEPTORS; H-2 RECEPTORS; REGULATORY PROTEINS; MOLECULAR ANALYSIS; HUMAN-NEUTROPHILS; FUSION PROTEINS; GS-ALPHA; CELLS; PHARMACOLOGY;
Subjects: 600 Technology > 615 Pharmacy
Divisions: Chemistry and Pharmacy > Institute of Pharmacy > Alumni or Retired Professors > Pharmaceutical/Medicinal Chemistry II (Prof. Buschauer)
Depositing User: Dr. Gernot Deinzer
Date Deposited: 24 Nov 2021 07:55
Last Modified: 24 Nov 2021 07:56
URI: https://pred.uni-regensburg.de/id/eprint/40908

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