| 18212018 |
Lim CC, Yang H, Yang M, Wang CK, Shi J, Berg EA, Pimentel DR, Gwathmey JK, Hajjar RJ, Helmes M, Costello CE, Huo S, Liao R: A novel mutant cardiac troponin C disrupts molecular motions critical for calcium binding affinity and cardiomyocyte contractility. Biophys J. 2008 May 1;94(9):3577-89. Epub 2008 Jan 22.
Troponin C (TnC) belongs to the superfamily of EF-hand (helix-loop-helix) Ca (2+)-binding proteins and is an essential component of the regulatory thin filament complex. In a patient diagnosed with idiopathic dilated cardiomyopathy, we identified two novel missense mutations localized in the regulatory Ca (2+)-binding Site II of TnC, TnC ((E59D,D75Y)). Expression of recombinant TnC ((E59D,D75Y)) in isolated rat cardiomyocytes induced a marked decrease in contractility despite normal intracellular calcium homeostasis in intact cardiomyocytes and resulted in impaired myofilament calcium responsiveness in Triton-permeabilized cardiomyocytes. Expression of the individual mutants in cardiomyocytes showed that TnC (D75Y) was able to recapitulate the TnC ((E59D,D75Y)) phenotype, whereas TnC (E59D) was functionally benign. Force-pCa relationships in TnC ((E59D,D75Y)) reconstituted rabbit psoas fibers and fluorescence spectroscopy of TnC ((E59D,D75Y)) labeled with 2-[(4'-iodoacetamide)-aniline] naphthalene-6-sulfonic acid showed a decrease in myofilament Ca (2+) sensitivity and Ca (2+) binding affinity, respectively. Furthermore, computational analysis of TnC showed the Ca (2+)-binding pocket as an active region of concerted motions, which are decreased markedly by mutation D75Y. We conclude that D75Y interferes with proper concerted motions within the regulatory Ca (2+)-binding pocket of TnC that hinders the relay of the thin filament calcium signal, thereby providing a primary stimulus for impaired cardiomyocyte contractility. This in turn may trigger pathways leading to aberrant ventricular remodeling and ultimately a dilated cardiomyopathy phenotype. |
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