Source:http://linkedlifedata.com/resource/pubmed/id/10722752
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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
12
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| pubmed:dateCreated |
2000-4-27
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| pubmed:abstractText |
Gbetagamma binds directly to the third intracellular (i3) loop subdomain of the M(3)-muscarinic receptor (MR). In this report, we identified the Gbetagamma binding motif and G-protein-coupled receptor kinase (GRK2) phosphorylation sites in the M(3)-MR i3 loop via a strategy of deletional and site-directed mutagenesis. The Gbetagamma binding domain was localized to Cys(289)-His(330) within the M(3)-MR-Arg(252)-Gln(490) i3 loop, and the binding properties (affinity, influence of ionic strength) of the M(3)-MR-Cys(289)-His(330) i3 loop subdomain were similar to those observed for the entire i3 loop. Site-directed mutagenesis of the M(3)-MR-Cys(289)-His(330) i3 loop subdomain indicated that Phe(312), Phe(314), and a negatively charged region (Glu(324)-Asp(329)) were required for interaction with Gbetagamma. Generation of the full-length M(3)-MR-Arg(252)-Gln(490) i3 peptides containing the F312A mutation were also deficient in Gbetagamma binding and exhibited a reduced capacity for phosphorylation by GRK2. A similar, parallel strategy resulted in identification of major residues ((331)SSS(333) and (348)SASS(351)) phosphorylated by GRK2, which were just downstream of the Gbetagamma binding motif. Full-length M(3)-MR constructs lacking the 42-amino acid Gbetagamma binding domain (Cys(289)-His(330)) or containing the F312A mutation exhibited ligand recognition properties similar to wild type receptor and also effectively mediated agonist-induced increases in intracellular calcium following receptor expression in Chinese hamster ovary and/or COS 7 cells. However, the M(3)-MRDeltaCys(289)-His(330) and M(3)-MR(F312A) constructs were deficient in agonist-induced sequestration, indicating a key role for the Gbetagamma-M(3)-MR i3 loop interaction in receptor regulation and signal processing.
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| pubmed:grant | |
| pubmed:language |
eng
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| pubmed:journal | |
| pubmed:citationSubset |
IM
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| pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/Cyclic AMP-Dependent Protein Kinases,
http://linkedlifedata.com/resource/pubmed/chemical/G-protein Beta gamma,
http://linkedlifedata.com/resource/pubmed/chemical/GTP-Binding Protein beta Subunits,
http://linkedlifedata.com/resource/pubmed/chemical/GTP-Binding Protein gamma Subunits,
http://linkedlifedata.com/resource/pubmed/chemical/GTP-Binding Proteins,
http://linkedlifedata.com/resource/pubmed/chemical/Heterotrimeric GTP-Binding Proteins,
http://linkedlifedata.com/resource/pubmed/chemical/Peptide Fragments,
http://linkedlifedata.com/resource/pubmed/chemical/Receptor, Muscarinic M3,
http://linkedlifedata.com/resource/pubmed/chemical/Receptors, Muscarinic,
http://linkedlifedata.com/resource/pubmed/chemical/Recombinant Proteins,
http://linkedlifedata.com/resource/pubmed/chemical/Serine,
http://linkedlifedata.com/resource/pubmed/chemical/beta-Adrenergic Receptor Kinases
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| pubmed:status |
MEDLINE
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| pubmed:month |
Mar
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| pubmed:issn |
0021-9258
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| pubmed:author | |
| pubmed:issnType |
Print
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| pubmed:day |
24
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| pubmed:volume |
275
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| pubmed:owner |
NLM
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| pubmed:authorsComplete |
Y
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| pubmed:pagination |
9026-34
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| pubmed:dateRevised |
2009-11-19
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| pubmed:meshHeading |
pubmed-meshheading:10722752-Amino Acid Sequence,
pubmed-meshheading:10722752-Binding Sites,
pubmed-meshheading:10722752-Biological Transport,
pubmed-meshheading:10722752-Cyclic AMP-Dependent Protein Kinases,
pubmed-meshheading:10722752-GTP-Binding Protein beta Subunits,
pubmed-meshheading:10722752-GTP-Binding Protein gamma Subunits,
pubmed-meshheading:10722752-GTP-Binding Proteins,
pubmed-meshheading:10722752-Heterotrimeric GTP-Binding Proteins,
pubmed-meshheading:10722752-Molecular Sequence Data,
pubmed-meshheading:10722752-Mutagenesis, Site-Directed,
pubmed-meshheading:10722752-Peptide Fragments,
pubmed-meshheading:10722752-Phosphorylation,
pubmed-meshheading:10722752-Receptor, Muscarinic M3,
pubmed-meshheading:10722752-Receptors, Muscarinic,
pubmed-meshheading:10722752-Recombinant Proteins,
pubmed-meshheading:10722752-Serine,
pubmed-meshheading:10722752-Signal Transduction,
pubmed-meshheading:10722752-beta-Adrenergic Receptor Kinases
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| pubmed:year |
2000
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| pubmed:articleTitle |
Identification of Gbetagamma binding sites in the third intracellular loop of the M(3)-muscarinic receptor and their role in receptor regulation.
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| pubmed:affiliation |
Department of Pharmacology, Medical University of South Carolina, Charleston, South Carolina 29425, USA.
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| pubmed:publicationType |
Journal Article,
Research Support, U.S. Gov't, P.H.S.
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