Ca2+-induced conformational transition in the inhibitory and regulatory regions of cardiac troponin I.
Cardiac muscle activation is initiated by the binding of Ca(2+) to the single N-domain regulatory site of cardiac muscle troponin C (cTnC). Ca(2+) binding causes structural changes between cTnC and two critical regions of cardiac muscle troponin I (cTnI): the regulatory region (cTnI-R, residues 150-165) and the inhibitory region (cTnI-I, residues130-149). These changes are associated with a decreased cTnI affinity for actin and a heightened affinity for cTnC. Using Förster resonance energy transfer, we have measured three intra-cTnI distances in the deactivated (Mg(2+)-saturated) and Ca(2+)-activated (Ca(2+)-saturated) states in reconstituted binary (cTnC-cTnI) and ternary (cTnC-cTnI-cTnT) troponin complexes. Distance A (spanning cTnI-R) was unaltered by Ca(2+). Distances B (spanning both cTnI-R and cTnI-I) and C (from a residue flanking cTnI-I to a residue in the center of cTnI-R) exhibited Ca(2+)-induced increases of >8 A. These results compliment our previous determination of the distance between residues flanking cTnI-I alone. Together, the data suggest that Ca(2+) activation causes residues within cTnI-I to switch from a beta-turn/coil to an extended quasi-alpha-helical conformation as the actin-contacts are broken, whereas cTnI-R remains alpha-helical in both Mg(2+)- and Ca(2+)-saturated states. We have used the data to construct a structural model of the cTnI inhibitory and regulatory regions in the Mg(2+)- and Ca(2+)-saturated states.
Dong
Wen-Ji
WJ
Department of Biochemistry and Molecular Genetics, University of Alabama, Birmingham 35294-2041, USA.
Robinson
John M
JM
Stagg
Scott
S
Xing
Jun
J
Cheung
Herbert C
HC
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002645:Chickens; D004735:Energy Transfer; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism*; D009282:Naphthalenesulfonates; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D019210:Troponin I / Q000737:chemistry / Q000378:metabolism*
D009282:Naphthalenesulfonates; D011994:Recombinant Proteins; D019210:Troponin I; C048876:1,5-AEDANS; D002118:Calcium