title: Intramolecular interactions in the N-domain of cardiac troponin C are important determinants of calcium sensitivity of force development.
abstract: Myocardial contraction is initiated when Ca2+ binds to site II of cardiac troponin C. This 12-residue EF-hand loop (NH2-DEDGSGTVDFDE-COOH) contains six residues (bold) that coordinate Ca2+ binding and six residues that do not appear to influence Ca2+ binding directly. We have introduced six single-cysteine substitutions (italics) within site II of cTnC to investigate whether these residues are essential for Ca2+ binding affinity in isolation and Ca2+ sensitivity of force development in single muscle fibers. Ca2+ binding properties of mutant proteins were examined in solution and after substitution into rat skinned soleus fibers. Except for the serine mutation, cysteine substitution had no effect on Ca2+ binding on cTnC in solution. However, as part of the myofilament, the threonine mutation reduced Ca2+ sensitivity while the phenylalanine mutation increased Ca2+ sensitivity. Analysis of the available crystal and NMR structures reveals specific structural mechanisms for these effects.
authors.affiliation: Department of Physiology, UniVersity of Wisconsin School of Medicine and Public Health, 123 Service Memorial Institute, 1300 University Avenue, Madison, Wisconsin 53706, USA.
authors.forename: Karen L
authors.initials: KL
authors.lastname: Reece
authors.forename: Richard L
authors.initials: RL
authors.lastname: Moss
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002645:Chickens; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009206:Myocardium / Q000737:chemistry* / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D051381:Rats; D012680:Sensitivity and Specificity; D019209:Troponin C / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D019209:Troponin C; D003545:Cysteine; D002118:Calcium