Regulation of binding of myosin subfragments with regulated actin by calcium ions in the presence of magnesium ATP.
Previously, several workers reported that at very low ionic strength and in the presence of ATP, the extent of binding of S-1 with the F-actin-tropomyosin-troponin complex or regulated actin (FA-TM-TN) is unaffected by removal of Ca2+. However, in this study we found that during the ATPase reaction at physiological ionic strength, the extent of binding of HMM with FA-TM-TN decreased markedly upon removal of CA2+. Therefore, the effects of Ca2+ were studied on the intermediate steps in the acto-HMM or acto-S-1 ATPase reaction. 1. The nucleotide-induced dissociation of acto-s-1 was studied using AMPPNP as substrate. The extent of binding of S-1 with regulated actin in the presence of Mg2+-AMPPNP increased in a sigmoidal manner as the S-1 concentration increased. When the molar ratio of actin monomer to S-1 was higher than 5-10, the removal of Ca2+ shifted the equilibrium of the dissociation reaction, FA-S-1-AMPPNP in equilibrium FA + S-1-AMPPNP, to the right. 2. The recombination rate of HMMPADP or S-1PADP with regulated actin in the absence of free Mg2+-ATP was estimated by measuring the time course of recovery in light-scattering intensity after addition of ATP. The rate decreased upon removal of Ca2+, when the molar ratio of actin monomer to S-1 was higher than 5-10. 3. The decomposition rate of HMMPADP was measured in the presence of Mg2+-ATP. In the absence of Ca2+, regulated actin did not affect this rate, whereas in its presence, regulated actin markedly accelerated the rate. These findings clearly indicated that at physiological ionic strength, removal of Ca2+ affects various elementary steps in the ATPase reaction to promote the dissociation of myosin heads from FA-TM-TN.
D000199:Actins / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D002118:Calcium / Q000502:physiology*; D006868:Hydrolysis; D007700:Kinetics; D009218:Myosins / Q000378:metabolism*; D011485:Protein Binding
D000199:Actins; D000255:Adenosine Triphosphate; D009218:Myosins; D002118:Calcium