Analysis of troponin-tropomyosin binding to actin. Troponin does not promote interactions between tropomyosin molecules

• Published in: The Journal of biological chemistry 1992-08, Vol.267 (23), p.16106-16113
• Main Authors: HILL, L. E, MEHEGAN, J. P, BUTTERS, C. A, TOBACMAN, L. S
• Format: Article
• Language: English
• Subjects: actin; Actins - isolation & purification; Actins - metabolism; Analytical, structural and metabolic biochemistry; Animals; binding; Biological and medical sciences; Calcium - pharmacology; Contractile proteins; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; Holoproteins; Kinetics; Molecular Weight; Muscles - metabolism; Protein Binding; Proteins; Rabbits; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; specificity; tropomyosin; Tropomyosin - isolation & purification; Tropomyosin - metabolism; troponin; Troponin - isolation & purification; Troponin - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)41973-4

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.