The molecular mechanism by which Ca²⁺ binding and phosphorylation regulate muscle contraction through Troponin is not yet fully understood. Revealing the differences between the relaxed and active structure of cTn, as well as the conformational changes that follow phosphorylation has remained a challenge for structural biologists over the years. Here we review the current understanding of how Ca²⁺, phosphorylation and disease-causing mutations affect the structure and dynamics of troponin to regulate the thin filament based on electron microscopy, X-ray diffraction, NMR and molecular dynamics methodologies.