Characterization of the Ca(2+)-triggered conformational transition in troponin C.
Troponin C is the Ca(2+)-binding subunit of troponin in vertebrate striated muscle. Binding of Ca2+ to troponin C is thought to induce a conformational change that triggers subsequent events in the initiation of muscle contraction. A molecular modeling study has proposed that, when Ca2+ binds to the N-terminal triggering sites, helices B and C separate from the helices D and A, thereby exposing a crucial interaction site for troponin I, the inhibitory subunit of troponin [Herzberg, O., Moult, J., and James, M. N. G. (1986) J. Biol. Chem. 261, 2638-2644]. In the present study the question of whether this separation actually occurs is addressed directly. A mutant rabbit skeletal troponin C containing a pair of cysteines at position 12 in helix A and position 49 in the polypeptide segment linking helices B and C was created by site-directed mutagenesis. Pyrene excimer fluorescence and resonance energy transfer studies on the labeled mutant troponin C reveal a Ca(2+)-induced increase in distance between the two cysteines. Under certain assumptions, the distance increase could be estimated from the extent of energy transfer to be approximately 13 A, in good agreement with the distance increase predicted by molecular modeling. Our results provide further experimental support for the model proposed by Herzberg et al. (above).
Wang
Z
Z
Department of Muscle Research, Boston Biomedical Research Institute, MA 02114.
Gergely
J
J
Tao
T
T
D000818:Animals; D001059:Apoproteins / Q000648:ultrastructure; D002118:Calcium / Q000502:physiology*; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D004735:Energy Transfer; D066298:In Vitro Techniques; D009119:Muscle Contraction; D009218:Myosins / Q000378:metabolism; D011487:Protein Conformation; D011817:Rabbits; D011994:Recombinant Proteins; D013050:Spectrometry, Fluorescence; D014336:Troponin / Q000648:ultrastructure*; D019209:Troponin C
D001059:Apoproteins; D011994:Recombinant Proteins; D014336:Troponin; D019209:Troponin C; D009218:Myosins; D000252:Calcium-Transporting ATPases; D002118:Calcium