Kinetic comparison of normal and thyrotoxic bovine cardiac myosin subfragment-1.

A comparison of the transient kinetics of cardiac ventricular normal and hyperthyroid modified myosin subfragment-1 reveals substantial similarities between the two proteins. The nucleotide-binding kinetics are nonexponential for both proteins, but the large tryptophan fluorescence changes, 34% for ATP binding and 12% for ADP binding which are comparable to those of rabbit skeletal myosin subfragment-1, permit the kinetic data to be resolved into a sum of two exponentials. Both the fast and slow forms of the proteins reach limiting rate constants at high nucleotide concentration. The fast forms of normal and thyrotoxic cardiac subfragment-1 are kinetically identical for nucleotide binding at 20 degrees C and pH 7 and the slow forms differ by less than a factor of 2. The kinetic data for ADP release and the single turnover of ATP could neither be fit by a single exponential nor resolved into two components, which indicates a difference in the rate constants by a factor of 2 or less. The largest difference found was in the steady state turnover of ATP for which thyrotoxic subfragment-1 had a 2.5 times faster turnover as compared to normal subfragment-1. The fractions of fast and slow forms of the two proteins are dependent on the nucleotide concentration and the fractions as well as the rate constants are a function of the protein concentration. This is consistent with the kinetic heterogeneity of cardiac myosin subfragment-1 resulting from aggregation. The differences in the rate constant for the steady state turnover of ATP and in aggregation properties between normal and hyperthyroid cardiac subfragment-1 are consistent with the induction of a myosin isozyme by thyroxine treatment. Moreover, the increase in the steady state turnover of ATP is consistent with the increase in contractility of the muscle in the hyperthyroid state.