Dynamics and thermodynamics of the regulatory domain of human cardiac troponin C in the apo- and calcium-saturated states

The contraction of cardiac and skeletal muscles is triggered by the binding of Ca2+ to their respective troponin C (TnC) proteins, Recent structural d ata of both cardiac and skeletal TnC in both the apo and Ca2+ states have r evealed that the response to Ca2+ is fundamentally different for these two proteins. For skeletal TnC, binding of two Ca2+ to sites and 2 leads to lar ge changes in the structure, resulting in the exposure of a hydrophobic sur face. For cardiac TnC, Ca2+ binds site 2 only, as site is inactive, and the structures show that the Ca2+-induced changes are much smaller and do not result in the exposure of a large hydrophobic surface. To understand the di fferences between regulation of skeletal and cardiac muscle, we have invest igated the effect of Ca2+ binding on the dynamics and thermodynamics of the regulatory N-domain of cardiac TnC (cNTnC) using backbone N-15 nuclear mag netic resonance relaxation measurements for comparison to the skeletal syst em. Analysis of the relaxation data allows for the estimation of the contri bution of changes in picosecond to nanosecond time scale motions to the con formational entropy of the Ca2+-binding sites on a per residue basis, which can be related to the structural features of the sites. The results indica te that binding of Ca2+ to the functional site in cNTnC makes the site more rigid with respect to high-frequency motions; this corresponds to a decrea se in the conformational entropy (T Delta S) of the site by 2.2 kcal mol(-1 ). Although site 1 is defunct, binding to site 2 also decreases the conform ational entropy in the nonfunctional site by 0.5 kcal mol(-1). The results indicate that the Ca2+-binding sites in the regulatory domain are structura lly and energetically coupled despite the inability of site I to bind Ca2Comparison between the cardiac and skeletal isoforms in the apo state shows that there is a decrease in conformational entropy of 0.9 kcal mol(-1) fur site 1 of cNTnC and little difference for site 2.