Studying Calcium Ion-Dependent Effect on the Inter-subunit Interaction Between the cTnC N-terminal Domain and cTnI C-terminal Switch Peptide of Human Cardiac Troponin via Chou’s 5-Steps Rule

Abstract

Human cardiac troponin (cTn) is a calcium ion (Ca2+)-sensitive hetero-trimer complex that consists of three subunits: cTnC, cTnI and cTnT. The protein is a primary biomarker and potential target for the diagnosis and therapy of myocardial necrosis in acute coronary syndrome. Previously, a minimal region (mSwt peptide) of cTnI C-terminal switch peptide is identified as a functionally required segment for mediating cTnI interaction with the cTnC N-terminal domain to which Ca2+ binds. Here, we attempt to investigate the Ca2+-dependent effect on the inter-subunit interaction between cTnC and cTnI at molecular level. The structure of Ca2+-bound and Ca2+-free cTnC N-terminal domains as well as their complexes with mSwt peptide are stripped from the crystal structure of full-length cTn complex, which are then subjected to 600-ns molecular dynamics simulations for conformational equilibrium and post energetics analysis. It is revealed that Ca2+ can effectively stabilize the native conformation of cTnC N-terminal domain and the tight binding mode of mSwt peptide to the domain; lack of Ca2+ would cause a considerable unfolding phenomenon in the domain, particularly in helices H1 and H5, and largely shift the interaction manner between the domain and peptide. In addition, there is also a significant difference between the binding curves of mSwt peptide to Ca2+-bound and Ca2+-free domains along the dynamics trajectory; the former monotonically increases over the whole simulations, whereas the latter exhibits a unimodal profile. Binding analysis also observes a significant concentration-dependent effect of Ca2+ on the domain–peptide affinity, that is, just a little of Ca2+ can improve the affinity remarkably. Overall, it is demonstrated that Ca2+ plays a crucial role in domain–peptide interaction and Ca2+ should bind very efficiently to the domain.