Analysis of the Molecular Pathogenesis of Cardiomyopathy-Causing cTnT Mutants 179N, ΔE96, and ΔK210

Three troponin T (TnT) mutants that cause hypertrophic, restrictive, and dilated cardiomyopathy (I79N, AE96, and K210, respectively), were examined using the thin-filament extraction/reconstitution technique. Effects of Ca2+, ATP, phosphate, and ADP concentrations on force and its transients were studied at 25 degrees C. Maximal Ca2+ tension (T-HC) and Ca2+-activatable tension (T-act), resectively, were similar among I79N, Delta E96, and WT, whereas Delta K210 led to a significantly lower T-HC (similar to 20% less) and T-act (similar to 25% less) than did WT. In pCa solution containing 8 mM Pi and ionic strength adjusted to 200 mM, the Ca2+ sensitivity (pCa(50)) of I79N (5.63 +/- 0.02) and Delta E96 (5.60 +/- 0.03) was significantly greater than that of WT (5.45 +/- 0.04), but the pCa(50) of Delta K210 (5.54 +/- 0.04) remained similar to that of WT. Five equilibrium constants were deduced using sinusoidal analysis. All three mutants showed significantly lower K-0 (ADP association constant) and larger K-4 (equilibrium constant of force generation step) relative to the corresponding values for WT. I79N and Delta K210 were associated with a K-2 (equilibrium constant of cross-bridge detachment step) significantly lower than that of Delta E96 and WT. These results demonstrated that at pCa 4.66, the force/cross-bridge is similar to 18% less in I79N and similar to 41% less in Delta K210 than that in WT. These results indicate that the molecular pathogenesis of the cardiac TnT mutation-related cardiomyopathies is different for each mutation.