Molecular Effects of Cardiac Troponin DCM Mutations on Calcium Sensitivity and Myofilament Activation - an Integrated Multi-Scale Modeling Study

Abstract

Mutations in cTnC (D75Y, E59D and G159D), a key regulatory protein of myofilament contraction, have been associated with dilated cardiomyopathy (DCM). Despite reports of altered myofilament function in these mutants, the underlying molecular alterations caused by these mutations remain elusive. Here we investigate in silico the intra-molecular mechanisms (both structure and dynamics) by which these mutations affect myofilament contraction (i.e. function). Based on the location of cTnC mutations, we tested the hypothesis that intra-molecular effects can explain the altered myofilament calcium sensitivity of force development for D75Y and E59D cTnC, whereas altered cTnC-cTnI interaction contributes to the reported contractile effects of the G159D mutation. We employed a multi-scale approach combining molecular dynamics (MD) and Brownian dynamics (BD) simulations to estimate cTnC calcium association and hydrophobic patch opening. We then integrated these parameters into a Markov model of myofilament activation to compute the steady-state force-pCa relationship. The analysis showed that myofilament calcium sensitivity with D75Y and E59D can be explained by changes in calcium binding affinity of cTnC and the rate of hydrophobic patch opening, if a partial cTnC interhelical opening angle (110°) is sufficient for cTnI switch peptide association to cTnC. In contrast, interactions between cTnC and cTnI within the cardiac troponin complex must also be accounted for to explain contractile alterations due to G159D. In conclusion, this is the first multi-scale in silico study to elucidate how direct molecular effects of genetic mutations in cTnC translate to altered myofilament contractile function.