| Identification of a functionally critical protein kinase C phosphorylation residue of cardiac troponin T. | |
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MedLine Citation:
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PMID: 12832403 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Cardiac Troponin T (cTnT) is one prominent substrate through which protein kinase C (PKC) exerts its effect on cardiomyocyte function. To determine the specific functional effects of the cTnT PKC-dependent phosphorylation sites (Thr197, Ser201, Thr206, and Thr287) we first mutated these residues to glutamate (E) or alanine (A). cTnT was selectively mutated to generate single, double, triple, and quadruple mutants. Bacterially expressed mutants were evaluated in detergent-treated mouse left ventricular papillary muscle fiber bundles where the endogenous troponin was replaced with a recombinant troponin complex containing either cTnT phosphorylated by PKC-alpha or a mutant cTnT. We simultaneously determined isometric tension development and actomyosin Mg-ATPase activity of the exchanged fiber bundles as a function of Ca2+ concentration. Our systematic analysis of the functional role of the multiple PKC phosphorylation sites on cTnT identified a localized region that controls maximum tension, ATPase activity, and Ca2+ sensitivity of the myofilaments. An important and novel finding of our study was that Thr206 is a functionally critical cTnT PKC phosphorylation residue. Its exclusive phosphorylation by PKC-alpha or replacement by Glu (mimicking phosphorylation) significantly decreased maximum tension, actomyosin Mg-ATPase activity, myofilament Ca2+ sensitivity, and cooperativity. On the other hand the charge modification of the other three residues together (T197/S201/T287-E) had no functional effect. Fibers bundles containing phosphorylated cTnT-wt (but not the T197/S201/T206/T287-E) exhibited a significant decrease of tension cost as compared with cTnT-wt. |
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Authors:
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Marius P Sumandea; W Glen Pyle; Tomoyoshi Kobayashi; Pieter P de Tombe; R John Solaro |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S. Date: 2003-06-28 |
Journal Detail:
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Title: The Journal of biological chemistry Volume: 278 ISSN: 0021-9258 ISO Abbreviation: J. Biol. Chem. Publication Date: 2003 Sep |
Date Detail:
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Created Date: 2003-09-08 Completed Date: 2003-11-12 Revised Date: 2007-11-15 |
Medline Journal Info:
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Nlm Unique ID: 2985121R Medline TA: J Biol Chem Country: United States |
Other Details:
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Languages: eng Pagination: 35135-44 Citation Subset: IM |
Affiliation:
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Department of Physiology and Biophysics, Program in Cardiovascular Sciences, College of Medicine, University of Illinois at Chicago, Chicago, Illinois 60612, USA. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Actomyosin
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metabolism Amino Acid Sequence Amino Acid Substitution Animals Ca(2+) Mg(2+)-ATPase / metabolism Calcium / pharmacology Cloning, Molecular Glutamic Acid Humans Mice Molecular Sequence Data Mutagenesis, Site-Directed Myocardial Contraction / physiology* Myocardium / metabolism Peptide Fragments / chemistry, metabolism Phosphorylation Phosphoserine / metabolism Phosphothreonine / metabolism Protein Kinase C / genetics, metabolism* Protein Structure, Secondary Recombinant Proteins / chemistry, metabolism Troponin T / chemistry, metabolism* Ventricular Function, Left / physiology* |
| Grant Support | |
ID/Acronym/Agency:
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F32 HL 10409/HL/NHLBI NIH HHS; P01-HL62426/HL/NHLBI NIH HHS; R37 HL2231/HL/NHLBI NIH HHS; T3207692//PHS HHS |
| Chemical | |
Reg. No./Substance:
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0/Peptide Fragments; 0/Recombinant Proteins; 0/Troponin T; 1114-81-4/Phosphothreonine; 17885-08-4/Phosphoserine; 56-86-0/Glutamic Acid; 7440-70-2/Calcium; 9013-26-7/Actomyosin; EC 2.7.11.13/Protein Kinase C; EC 3.6.1.-/Ca(2+) Mg(2+)-ATPase |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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