Influence of differential stability of G protein ßγ dimers containing the γ11 subunit on functional activity at the M1 muscarinic receptor, A1 adenosine receptor, and phospholipase C-ß.
Biochemistry
; 45(38): 11616-31, 2006 Sep 26.
Article
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| MEDLINE
| ID: mdl-16981721
ABSTRACT
Ggamma11 is an unusual guanine nucleotide-binding regulatory protein (G protein) subunit. To study the effect of different Gbeta-binding partners on gamma11 function, four recombinant betagamma dimers, beta1gamma2, beta4gamma2, beta1gamma11, and beta4gamma11, were characterized in a receptor reconstitution assay with the G(q)-linked M1 muscarinic and the G(i1)-linked A1 adenosine receptors. The beta4gamma11 dimer was up to 30-fold less efficient than beta4gamma2 at promoting agonist-dependent binding of [35S]GTPgammaS to either alpha(q) or alpha(i1). Using a competition assay to measure relative affinities of purified betagamma dimers for alpha, the beta4gamma11 dimer had a 15-fold lower affinity for G(i1) alpha than beta4gamma2. Chromatographic characterization of the beta4gamma11 dimer revealed that the betagamma is stable in a heterotrimeric complex with G(i1) alpha; however, upon activation of alpha with MgCl2 and GTPgammaS under nondenaturing conditions, the beta4 and gamma11 subunits dissociate. Activation of purified G(i1) alphabeta4gamma11 with Mg+2/GTPgammaS following reconstitution into lipid vesicles and incubation with phospholipase C (PLC)-beta resulted in stimulation of PLC-beta activity; however, when this activation preceded reconstitution into vesicles, PLC-beta activity was markedly diminished. In a membrane coupling assay designed to measure the ability of G protein to promote a high-affinity agonist-binding conformation of the A1 adenosine receptor, beta4gamma11 was as effective as beta4gamma2 when coexpressed with G(i1) alpha and receptor. However, G(i1) alphabeta4gamma11-induced high-affinity binding was up to 20-fold more sensitive to GTPgammaS than G(i1) alphabeta4gamma2-induced high-affinity binding. These results suggest that the stability of the beta4gamma11 dimer can modulate G protein activity at the receptor and effector.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Fosfolipasas de Tipo C
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Subunidades de Proteína
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Subunidades beta de la Proteína de Unión al GTP
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Subunidades gamma de la Proteína de Unión al GTP
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Receptor Muscarínico M1
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Receptor de Adenosina A1
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Isoenzimas
Límite:
Animals
Idioma:
En
Revista:
Biochemistry
Año:
2006
Tipo del documento:
Article
País de afiliación:
Estados Unidos