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A Novel Mutant Cardiac Troponin C Disrupts Molecular Motions Critical for Calcium Binding Affinity and Cardiomyocyte Contractility | Author(s) | Chee Chew Lim, Haijun Yang, Mingfeng Yang, Chien-Kao Wang, Jianru Shi, Eric A. Berg, David R. Pimentel, Judith K. Gwathmey, Roger J. Hajjar, Michiel Helmes, Catherine E. Costello, Shuanghong Huo, and Ronglih Liao | | Journal | Biophys. J., May 2008; 94: 3577 - 3589. | Method Extract | | ...Cardiovascular Research Center, Charlestown, Massachusetts; IonOptix Europe, Wageningen, the Netherlands; and Cardiovascular...real-time using specialized acquisition and analysis software (IonOptix Corp., Milton, MA). A single myocyte was selected and exclusively... | Abstract | Troponin C (TnC) belongs to the superfamily of EF-hand (helix–loop–helix) Ca2+-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 Ca2+-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 TnCD75Y was able to recapitulate the TnC(E59D,D75Y) phenotype, whereas TnCE59D 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 Ca2+ sensitivity and Ca2+ binding affinity, respectively. Furthermore, computational analysis of TnC showed the Ca2+-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 Ca2+-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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| Keywords | calcium, ionwizard | Our Ref: 1632
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