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Calpain-1-dependent degradation of troponin I mutants found in familial hypertrophic cardiomyopathy

  • Judit Barta
  • Attila Tóth
  • Kornelia Jaquet
  • Alexander Redlich
  • István Édes
  • Zoltán Papp
Part of the Developments in Molecular and Cellular Biochemistry book series (DMCB, volume 43)

Abstract

The mechanism by which mutations of the cardiac troponin I (cTnI) gene evoke familial hypertrophic cardiomyopathy (fHCM) is unknown. In this investigation the potential effects of three fHCM-related cTnI mutations on Calpain-1-induced cTnI degradation were tested, and a study was made of whether additional conformational changes due to troponin complex formation and protein kinase A-induced phosphorylation affect the intensity of cTnI proteolysis. Purified recombinant wild-type cTnI and three of its fHCM-related missense mutants (R145G, G203S and K206Q), alone or in the troponin complex (i.e. together with troponin C and troponin T), in the non-phosphorylated or protein kinase A-bisphosphorylated forms were proteolyzed in vitro in the presence of Calpain-1 (0.05–2.5 U) at 30°C. Following incubation with Calpain-1 for 0.5, 30, 60 or 120 min, the extent of protein degradation was evaluated through the use of Western immunoblotting and densitometry. The results indicated that both the wild-type and the mutant cTnI molecules were susceptible to Calpain-1. However, the degradation of the cTnI molecules in the troponin complex was less intense than that of the non-complexed forms. Moreover, phosphorylation by protein kinase A conferred effective protection against cTnI proteolysis. The data suggested that mutations in the central inhibitory domain (R145G) and in the C-terminal region (G203S and K206Q) of cTnI do not affect its Calpain-1-mediated degradation, or the phosphorylation-induced protection against proteolysis.

Keywords

hypertrophic cardiomyopathy calpain troponin I mutation 

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Copyright information

© Springer Science+Business Media New York 2003

Authors and Affiliations

  • Judit Barta
    • 1
  • Attila Tóth
    • 1
  • Kornelia Jaquet
    • 2
  • Alexander Redlich
    • 2
  • István Édes
    • 1
  • Zoltán Papp
    • 1
    • 3
  1. 1.Department of CardiologyUniversity of Debrecen, MHSCDebrecenHungary
  2. 2.Institut für Physiologische ChemieRuhr-Universität BochumGermany
  3. 3.Department of CardiologyUniversity of Debrecen, Medical and Health Science CenterDebrecenHungary

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