The transmission of mutation effects in a multiprotein machine: A comprehensive metadynamics study of the cardiac thin filament.

The binding of Ca ions within the troponin core of the cardiac thin filament (CTF) regulates normal contraction and relaxation. Mutations within the troponin complexes are known to alter normal functions and result in the eventual development of cardiomyopathy. However, despite the importance of the problem, detailed microscopic knowledge of the mechanism of pathogenic effect of point mutations and their effects on the conformational free energy surface of CTF remains elusive. Mutations are known to transmit their effects hundreds of angstroms along this protein complex and between different component proteins. To explore the impact of point mutations on the conformational free energy barrier between the closed and blocked state of CTF, and to understand the transmission of mutation, we have carried out metadynamics simulations for the wild-type (WT) and two mutants (cardiac troponin T Arg92Trp (R92W) and Arg92Leu (R92L)). Specifically, we have investigated the conformational modification of the tropomyosin (Tm) and the troponin (Tn) complex during the closed-to-blocked state transition for both the WT and two hypertrophic cardiomyopathy causing mutations. Our calculations demonstrated that mutations within the cardiac troponin T (cTnT) protein alter conformational properties of the Tm and the other proteins of the Tn complex as well as the Ca binding affinity of the cTnC protein through the indirect mediation of cardiac troponin I (cTnI). Importantly, the data revealed a significant influence of the mutations on the conformational transition free energy barriers for both the Tm and cTnC proteins. Furthermore, we found both mutations independently alter the free energy barrier of transitions of cTnT. Such alteration in the free energy upon mutation of one protein in a complex, allosterically affects the others through structural and dynamical changes, leading to a pathogenic effect on the function of the thin filament.