Binding of troponin I and the troponin I-troponin C complex to actin-tropomyosin. Dissociation by myosin subfragment 1.
Troponin I (TnI) is the component of the troponin complex, TnI, TnC, TnT, that is responsible for inhibition of actomyosin ATPase activity. Using the fluorescence of pyrene-labeled tropomyosin (Tm), we probed the interaction of TnI and TnIC with Tm on the reconstituted muscle thin filament. The results indicate that TnI and TnIC(-Ca(2+)) bind specifically and strongly to actin-Tm with a stoichiometry of 1 TnI or 1 TnIC/1 Tm/7 actin, in agreement with previous results. The binding of myosin heads (S1) to actin-Tm at low levels of saturation caused TnI and TnIC to dissociate from actin-Tm. These results are interpreted in terms of the S1-binding state allosteric-cooperative model of the actin-Tm thin filament, closed/open. Thus, TnI and TnIC(-Ca(2+)) bind to the closed state of actin-Tm and their binding is greatly weakened in the S1-induced open state, indicating that they act as allosteric inhibitors. The fluorescence change and the stoichiometry indicate that the TnI-binding site is composed of regions from both actin and Tm probably in the vicinity of Cys 190.
Zhou
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Muscle Research Group, Boston Biomedical Research Institute, 20 Staniford Street, Boston, Massachusetts 02114, USA.
Morris
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Lehrer
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D000199:Actins / Q000378:metabolism*; D000495:Allosteric Site; D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D005456:Fluorescent Dyes / Q000378:metabolism; D007460:Iodoacetamide / Q000031:analogs & derivatives / Q000378:metabolism; D018482:Muscle, Skeletal / Q000737:chemistry / Q000378:metabolism; D015879:Myosin Subfragments / Q000378:metabolism*; D011485:Protein Binding; D011817:Rabbits; D013050:Spectrometry, Fluorescence; D014335:Tropomyosin / Q000378:metabolism*; D019209:Troponin C / Q000378:metabolism*; D019210:Troponin I / Q000378:metabolism*; D014461:Ultracentrifugation
D000199:Actins; D005456:Fluorescent Dyes; D015879:Myosin Subfragments; D014335:Tropomyosin; D019209:Troponin C; D019210:Troponin I; C029358:N-(1-pyrenyl)iodoacetamide; D007460:Iodoacetamide