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Mechanism of beta-adrenergic receptor kinase activation by G proteins

1993; Elsevier BV; Volume: 268; Issue: 21 Linguagem: Inglês

10.1016/s0021-9258(18)82273-6

1083-351X

C.M. Kim, Sarah Dion, Jeffrey Benovic,

Neuropeptides and Animal Physiology

The t9-adrenergic receptor kinase (&ARK) specifically phosphorylates the activated form of various G protein-coupled receptors such as the &adrenergic receptor (Bz-AR).Recently, G protein j 3r subunits have been demonstrated to activate B-ARK-mediated receptor phosphorylation.To further elucidate B-ARKIG protein interactions, we have developed a direct binding assay.The direct binding of [3BS]methionine-labeled @-ARK to either brain GJG, or @r subunits was rapid and saturable with similar Kd values of -58 and -32 nM, respectively.Both heterotrimeric G proteins and fly subunits enhanced the initial rate of &AR and rhodopsin phosphorylation -10-fold.Kinetic studies demonstrate that @r enhances @-ARK-mediated &-AR phosphorylation both by decreasing the K,,, for the j3,-AR -4-fold and increasing the stoichiometry of phosphorylation from -4 to -11 mol/mol.An agonist-and ATP-dependent binding of &ARK to the reconstituted &-AR was also demonstrated.In addition, &ARK binding was enhanced in the presence of both the activated &AR and B-y subunits suggesting the formation of a transient ternary complex consisting of @-ARK, By, and &AR.Overall, these studies suggest that the specific association of @-ARK with heterotrimeric G proteins may play an important role in promoting receptorlkinase interaction and subsequent receptor phosphorylation.The &-adrenergic receptor (P2-AR)'-coupled adenylylcyclase system has provided an important model for characterizing the molecular mechanisms of signal transduction.In this system, agonist binding to the &AR promotes activation of the G protein G, leading to increased adenylylcyclase activity and intracellular cAMP levels.However, prolonged agonist stimulation of the &AR results in an attenuation of cAMP generation (Benovic et al., 1989b; Hausdorff et al.,