Standard free energy changes for formation of various intermediates in the reaction of H-meromyosin ATPase.
Two reaction intermediates of H-meromyosin (HMM) ATPase [EC 3.6.1.3], E2AT32P, and (see article), were formed by mixing excess HMM with AT32P. Then a large excess of unlabelled ATP was added, and the amount of AT32P liberated from E2AT32P was measured as the difference between the total amount of AT32P in the reaction mixture and the amount of AT32P bound to HMM, obtained by filtering the mixture after adding charcoal to adsorb nucleotides (charcoal-filtration method). The amount of free AT32P was also measured as the amount of glucose-6-32P formed within 15 sec after adding large excesses of hexokinase [EC 2.7.1.1] and glucose to the reaction mixture. The rate constant, k-2, for the step E2ATP yields E plus ATP was calculated at various KCl concentrations from the time-course of liberation of AT32P. The intermediate, (see article), was formed by mixing HMM with AT32P in a molar ratio of 1:2, and the rate constant, k-6, for the step (see article) was also determined by the same procedures used for k-2. In 0.5 M KCl and 2 mM MgCl2 at pH 7.8 and 0 degrees, k-2 and k-6 were 0.002 sec-1 and 0.1 sec-1 or more, respectively. From the rate constants determined in this work and the rate and equilibrium constants which we reported previously, the standard free energy changes (kcal/mole) for formation of various reaction intermediates in the reaction of HMM ATPase in 0.5 M KCl and 2 mM MgCl2 at pH 7.8 and 0 degrees were calculated to be as follows: (see article).
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D006593:Hexokinase / Q000378:metabolism; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism; D008433:Mathematics; D015879:Myosin Subfragments; D010761:Phosphorus Radioisotopes; D011189:Potassium Chloride / Q000378:metabolism
D015879:Myosin Subfragments; D010761:Phosphorus Radioisotopes; D011189:Potassium Chloride; D000255:Adenosine Triphosphate; D006593:Hexokinase; D000251:Adenosine Triphosphatases; D008274:Magnesium