Separation and characterization of the two functional regions of troponin involved in muscle thin filament regulation.
Mild proteolytic cleavage of the troponin complex yields TnT1, the N-terminal fragment of troponin T, and TnT2IC, a complex of the C-terminal fragment of troponin T (TnT2) with troponin I (TnI) and troponin C (TnC) [Morris, E. P., & Lehrer, S. S. (1984) Biochemistry 23, 2214-2220]. Both TnT1 and TnT2IC bind tightly to the tropomyosin.actin (Tm.actin) thin filament and influence the interaction of myosin subfragment 1 (S1) with Tm.actin. TnT1 does not affect the rate of S1 binding to Tm.actin but does increase the cooperativity with which S1 "turns on" Tm.actin, monitored by the excimer fluorescence of a pyrene label attached to Cys 190 of Tm [Geeves, M.A., & Lehrer, S. S. (1994) Biophys. J. 67, 273-282]. The apparent cooperative unit size of Tm.actin is increased from 6 to 9 by TnT1 and to 12 by whole troponin. In contrast, TnT2IC has no effect on the cooperativity of Tm.actin but does make the apparent S1-binding rate constant, kapp, Ca(2+)-sensitive; i.e., in the absence of Ca2+, kapp is reduced 2-3-fold by both TnT2IC and whole troponin. Thus, the N- and C-terminal regions of TnT appear to act independently in modulating effects of S1 binding to the Tm.actin thin filament that are important in regulation.
Schaertl
S
S
Max Planck Institute für Molekulare Physiologie, Dortmund, Germany.
Lehrer
S S
SS
Geeves
M A
MA
D000199:Actins / Q000302:isolation & purification / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D007700:Kinetics; D018482:Muscle, Skeletal / Q000378:metabolism*; D015879:Myosin Subfragments / Q000302:isolation & purification / Q000378:metabolism; D010446:Peptide Fragments; D011485:Protein Binding; D011817:Rabbits; D013997:Time Factors; D014336:Troponin / Q000302:isolation & purification / Q000378:metabolism*; D020107:Troponin T
D000199:Actins; D015879:Myosin Subfragments; D010446:Peptide Fragments; D014336:Troponin; D020107:Troponin T; D000255:Adenosine Triphosphate; D002118:Calcium