Members of the G protein-coupled receptor kinase family that phosphorylate the beta2-adrenergic receptor facilitate sequestration

Biochemistry. 1996 Apr 2;35(13):4155-60. doi: 10.1021/bi952961+.


We recently reported that a beta2-adrenergic receptor (beta2AR) mutant, Y326A, defective in its ability to sequester in response to agonist stimulation was a poor substrate for G protein-coupled receptor kinase (GRK)-mediated phosphorylation; however, its ability to be phosphorylated and sequestered could be restored by overexpressing GRK2 [Ferguson et al. (1995) J. Biol. Chem. 270, 24782]. In the present report, we tested the ability of each of the known GRKs (GRK1-6) to phosphorylate and rescue the sequestration of the Y326A mutant in HEK-293 cells. We demonstrate that in addition to GRK2, GRK3-6 can phosphorylate the Y326A mutant and rescue its sequestration; however, GRK1 was totally ineffective in rescuing either the phosphorylation or the sequestration of the mutant receptor. We found that the agonist-dependent rescue of Y326A mutant phosphorylation by GRK2, -3, and -5 was associated with the agonist-dependent rescue of sequestration. In contrast, overexpression of GRK4 and -6 led mainly to agonist-independent phosphorylation of the Y326A mutant accompanied by increased basal receptor sequestration. Our results demonstrate that phosphorylation per se, but not the interaction with a specific GRK, is required to facilitate beta2AR sequestration.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adrenergic beta-Antagonists / pharmacology
  • Cell Line
  • Cloning, Molecular
  • GTP-Binding Proteins / metabolism*
  • Humans
  • Immunoblotting
  • Iodine Radioisotopes
  • Kidney
  • Phosphates / metabolism
  • Phosphorus Radioisotopes
  • Phosphorylation
  • Pindolol / metabolism
  • Point Mutation
  • Propanolamines / pharmacology
  • Receptor Protein-Tyrosine Kinases / isolation & purification
  • Receptor Protein-Tyrosine Kinases / metabolism*
  • Receptors, Adrenergic, beta-2 / isolation & purification
  • Receptors, Adrenergic, beta-2 / metabolism*
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Sequence Tagged Sites
  • Substrate Specificity
  • Transfection


  • Adrenergic beta-Antagonists
  • Iodine Radioisotopes
  • Phosphates
  • Phosphorus Radioisotopes
  • Propanolamines
  • Receptors, Adrenergic, beta-2
  • Recombinant Proteins
  • Pindolol
  • Receptor Protein-Tyrosine Kinases
  • GTP-Binding Proteins
  • CGP 12177