Many mutations causing the inherited diseases hypertrophic cardiomyopathy (HCM) and dilated cardiomyopathy(DCM) act by affecting Ca2+-regulation of contraction by troponin. HCM Mutations can increase Ca2+-sensitivity 2-3 fold and most mutations uncouple the Ca2+-sensitivity change from TnI phosphorylation by PKA. Small molecules that bind to troponin and reverse these abnormalities are of therapeutic interest. We have found that Epigallocatechin 3-Gallate (EGCG) is a Ca2+-desensitiser and also re-couples uncoupled mutant troponin. Silybin exists as stereoisomers: we found that Silybin B is a pure recoupler (no effect on Ca2+-sensitivity) whilst Silybin A has no activity. We propose that these compounds act by differentially binding to troponin in the phosphorylated, unphosphorylated, wild type and mutant states. In this work, we attempt to address the structural and dynamical effects of these compounds with atomistic detail building on the results obtained from our existing computational model of phosphorylated and unphosphorylated cTn in its Ca2+ activated state, totalling 419 amino acid residues. The objective is to study the binding modes of EGCG and Silybins A and B using extensive Molecular Dynamics simulations. Based on the same starting structure as previously, both Autodock and MD simulations indicate two preferred binding sites for EGCG, both are intercalated between the stable TnC N-terminal domain and the disordered N-terminal phosphorylatable peptide of cTnI forming hydrogen bonds with both peptides. According to Autodock calculations, Silybin A and B preferentially bind at the same interface as EGCG but they present a less favourable energy of interaction with cTn as the simulation proceeds, and in some simulations, they only remain bound for a fraction of the simulation time. Simulations of ligand binding to phosphorylated troponin and to troponin containing HCM and DCM-causing mutations are in progress and should yield valuable data on differential binding that may explain how these potentially therapeutic small molecules function.

EGCG and Sylibins as treatment for inherited cardiomyopathies: Binding simulations to cardiac troponin

253rd National Meeting (2017)

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