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Pleckstrin homology domain-mediated membrane association and activation of the beta-adrenergic receptor kinase requires coordinate interaction with G beta gamma subunits and lipid.

Publication ,  Journal Article
Pitcher, JA; Touhara, K; Payne, ES; Lefkowitz, RJ
Published in: J Biol Chem
May 19, 1995

The pleckstrin homology (PH) domain is an approximately 100-amino-acid region of sequence homology present in numerous proteins of diverse functions, which forms a discrete structural module. Several ligands capable of binding to PH domain-containing proteins have been identified including phosphatidylinositol 4,5-bisphosphate (PIP2) and the G beta gamma subunits of heterotrimeric G proteins (G beta gamma), which bind to the amino and carboxyl termini of the PH domain, respectively. Here we report that the binding of G beta gamma and lipid to the PH domain of the beta-adrenergic receptor kinase (beta ARK) synergistically enhances agonist-dependent receptor phosphorylation and that both PH domain-binding ligands are required for membrane association of the kinase. PIP2 and to a lesser extent phosphatidylinositol 4-phosphate, phosphatidylinositol, and phosphatidic acid were the only lipids tested capable, in the presence of G beta gamma, of enhancing beta ARK activity. In contrast, the Km and Vmax for phosphorylation of a soluble beta ARK substrate (casein) was not altered in either the presence or absence of G beta gamma and/or PIP2. A fusion protein of the beta ARK containing an intact PH domain inhibits G beta gamma/PIP2-dependent beta ARK activity. In contrast, a mutant fusion protein in which a tryptophan residue, invariant in all PH domain sequences, is mutated to alanine shows no inhibitory activity. The requirement for the simultaneous presence of two PH domain binding ligands represents a previously unappreciated mechanism for effecting membrane localization of a protein and may have relevance to other PH domain-containing proteins.

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Published In

J Biol Chem

DOI

ISSN

0021-9258

Publication Date

May 19, 1995

Volume

270

Issue

20

Start / End Page

11707 / 11710

Location

United States

Related Subject Headings

  • beta-Adrenergic Receptor Kinases
  • Structure-Activity Relationship
  • Sequence Homology, Amino Acid
  • Recombinant Fusion Proteins
  • Protein Structure, Tertiary
  • Protein Processing, Post-Translational
  • Phosphorylation
  • Phosphoproteins
  • Phospholipids
  • Phosphatidylinositols
 

Citation

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Pitcher, J. A., Touhara, K., Payne, E. S., & Lefkowitz, R. J. (1995). Pleckstrin homology domain-mediated membrane association and activation of the beta-adrenergic receptor kinase requires coordinate interaction with G beta gamma subunits and lipid. J Biol Chem, 270(20), 11707–11710. https://doi.org/10.1074/jbc.270.20.11707
Pitcher, J. A., K. Touhara, E. S. Payne, and R. J. Lefkowitz. “Pleckstrin homology domain-mediated membrane association and activation of the beta-adrenergic receptor kinase requires coordinate interaction with G beta gamma subunits and lipid.J Biol Chem 270, no. 20 (May 19, 1995): 11707–10. https://doi.org/10.1074/jbc.270.20.11707.
Pitcher, J. A., et al. “Pleckstrin homology domain-mediated membrane association and activation of the beta-adrenergic receptor kinase requires coordinate interaction with G beta gamma subunits and lipid.J Biol Chem, vol. 270, no. 20, May 1995, pp. 11707–10. Pubmed, doi:10.1074/jbc.270.20.11707.

Published In

J Biol Chem

DOI

ISSN

0021-9258

Publication Date

May 19, 1995

Volume

270

Issue

20

Start / End Page

11707 / 11710

Location

United States

Related Subject Headings

  • beta-Adrenergic Receptor Kinases
  • Structure-Activity Relationship
  • Sequence Homology, Amino Acid
  • Recombinant Fusion Proteins
  • Protein Structure, Tertiary
  • Protein Processing, Post-Translational
  • Phosphorylation
  • Phosphoproteins
  • Phospholipids
  • Phosphatidylinositols