Linked Life Data

14507916

Source:http://linkedlifedata.com/resource/pubmed/id/14507916

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
48
pubmed:dateCreated
2003-11-24
pubmed:abstractText
G protein-coupled receptor kinases (GRKs) specifically bind and phosphorylate the agonist-occupied form of G protein-coupled receptors. To further characterize the mechanism of GRK/receptor interaction, we developed a yeast-based bioassay using strains engineered to functionally express the somatostatin receptor subtype 2 and exhibit agonist-dependent growth. Here, we demonstrate that agonist-promoted growth was effectively inhibited by co-expression with either wild type GRK2 or GRK5, whereas catalytically inactive forms of these kinases were without effect. In an effort to identify residues involved in receptor interaction, we generated a pool of GRK5 mutants and then utilized the bioassay to identify mutants selectively deficient in inhibiting agonist-promoted growth. This resulted in the identification of a large number of mutants, several of which were expressed, purified, and characterized in more detail. Two of the mutants, GRK5-L3Q/K113R and GRK5-T10P, were defective in receptor phosphorylation and also exhibited a partial defect in phospholipid binding and phospholipid-stimulated autophosphorylation of the kinase. In contrast, these mutants had wild type activity in phosphorylating the non-receptor substrate tubulin. To further characterize the function of the NH2-terminal region of GRK5, we generated a deletion mutant lacking residues 2-14 and found that this mutant was also severely impaired in receptor phosphorylation and phospholipid-promoted autophosphorylation. In addition, an NH2-terminal 14-amino acid peptide from GRK5 selectively inhibited receptor phosphorylation by GRK5 but had minimal effect on GRK2 activity. Based on these findings, we propose a model whereby the extreme NH2 terminus of GRK5 mediates phospholipid binding and is required for optimal receptor phosphorylation.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Nov
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
28
pubmed:volume
278
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
47466-76
pubmed:dateRevised
2007-11-15
pubmed:meshHeading
pubmed-meshheading:14507916-Animals, pubmed-meshheading:14507916-Biological Assay, pubmed-meshheading:14507916-COS Cells, pubmed-meshheading:14507916-Catalysis, pubmed-meshheading:14507916-Cell Division, pubmed-meshheading:14507916-Cell Line, pubmed-meshheading:14507916-Cyclic AMP-Dependent Protein Kinases, pubmed-meshheading:14507916-Dose-Response Relationship, Drug, pubmed-meshheading:14507916-G-Protein-Coupled Receptor Kinase 5, pubmed-meshheading:14507916-Genetic Vectors, pubmed-meshheading:14507916-Humans, pubmed-meshheading:14507916-Insects, pubmed-meshheading:14507916-Kinetics, pubmed-meshheading:14507916-Lipids, pubmed-meshheading:14507916-Mutagenesis, pubmed-meshheading:14507916-Mutation, pubmed-meshheading:14507916-Peptides, pubmed-meshheading:14507916-Phospholipids, pubmed-meshheading:14507916-Phosphorylation, pubmed-meshheading:14507916-Plasmids, pubmed-meshheading:14507916-Protein Binding, pubmed-meshheading:14507916-Protein Structure, Tertiary, pubmed-meshheading:14507916-Protein-Serine-Threonine Kinases, pubmed-meshheading:14507916-Receptors, Adrenergic, beta-2, pubmed-meshheading:14507916-Saccharomyces cerevisiae, pubmed-meshheading:14507916-Time Factors, pubmed-meshheading:14507916-beta-Adrenergic Receptor Kinases
pubmed:year
2003
pubmed:articleTitle
Development of a yeast bioassay to characterize G protein-coupled receptor kinases. Identification of an NH2-terminal region essential for receptor phosphorylation.
pubmed:affiliation
Department of Microbiology and Immunology, Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.