Analysis of troponin-tropomyosin binding to actin. Troponin does not promote interactions between tropomyosin molecules.
The binding of tropomyosin to actin and troponin-tropomyosin to actin was analyzed according to a linear lattice model which quantifies two parameters: Ko, the affinity of the ligand for an isolated site on the actin filament, and gamma, the fold increase in affinity when binding is contiguous to an occupied site (cooperativity). Tropomyosin-actin binding is very cooperative (gamma = 90-137). Troponin strengthens tropomyosin-actin binding greatly but, surprisingly, does so solely by an 80-130-fold increase in Ko, while cooperativity actually decreases. Additionally, troponin complexes containing TnT subunits with deletions of either amino acids 1-69 (troponin70-259) or 1-158 (troponin159-259) were examined. Deletion of amino acids 1-69 had only small effects on Ko and y, despite this peptide's location spanning the joint between adjacent tropomyosins. Ca2+ reduced Ko by half for both troponin and troponin70-159 and had no detectable effect on cooperativity. Troponin159-259 had much weaker effects on tropomyosin-actin binding than did troponin70-259 and had no effect at all in the presence of Ca2+. This suggests the importance of Ca(2+)-insensitive interactions between tropomyosin and troponin T residues 70-159. Cooperativity was slightly lower for troponin159-259 than tropomyosin alone, suggesting that the globular head region of troponin affects tropomyosin-tropomyosin interactions along the thin filament.
Hill
L E
LE
Department of Internal Medicine, College of Medicine, University of Iowa, Iowa City 52242.
Mehegan
J P
JP
Butters
C A
CA
Tobacman
L S
LS
D000199:Actins / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008970:Molecular Weight; D009132:Muscles / Q000378:metabolism*; D011485:Protein Binding; D011817:Rabbits; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D014335:Tropomyosin / Q000302:isolation & purification / Q000378:metabolism*; D014336:Troponin / Q000302:isolation & purification / Q000378:metabolism*
D000199:Actins; D011994:Recombinant Proteins; D014335:Tropomyosin; D014336:Troponin; D002118:Calcium