Calcium binding to the regulatory domain of skeletal muscle troponin C induces a highly constrained open conformation.
We have used fluorescence resonance energy transfer to investigate the conformation of the apo and calcium-loaded states of the regulatory N-terminal domain of full-length troponin C mutants from skeletal muscle. The mutants studied each contained a single tryptophan residue (position 22 or 90) and a single cysteine residue (position 52 or 101). The intrinsic fluorophore in each mutant served as an energy donor and the cysteine was conjugated to the acceptor probe 5-(iodoacetamidoethyl)amino-naphthalene-1-sulfonic acid. The distributions of two intersite distances (between residues 22 and 52, and residues 90 and 52) were broad in the apo state, indicative of considerable structural dynamics. These distributions were shifted to longer distances and considerably sharpened in the calcium-loaded state. The shifts to longer distances by 8 to 11 A indicate a calcium-induced opening of the N-terminal domain conformation. The transition of the troponin C structure from a closed conformation to an open conformation is accompanied by a substantial reduction of structural fluctuations that dominate in the apo structure as evidenced from the large decrease of the widths of the distributions. This highly constrained open conformation is required as part of the structural basis to facilitate productive interaction between troponin C and troponin I to trigger contraction in skeletal muscle.
Department of Physics, University of Alabama at Birmingham, Birmingham, AL, 35294, USA.
M
M
She
J
J
Xing
W J
WJ
Dong
P K
PK
Umeda
H C
HC
Cheung
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D004735:Energy Transfer; D005454:Fluorescence Polarization; D008958:Models, Molecular; D018482:Muscle, Skeletal / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D019209:Troponin C / Q000737:chemistry* / Q000378:metabolism; D014364:Tryptophan / Q000737:chemistry; D014422:Turkeys
D019209:Troponin C; D014364:Tryptophan; D002118:Calcium