Inhibition of actomyosin ATPase by vanadate.

Actin-myosin subfragment-1 (SF-1) or actin-heavy meromyosin is dissociated by the binding of ADP and vanadate (Vi) under conditions such that ADP alone does not dissociate the complex. The association constant of the stable complex M.ADP.Vi, in which M indicates myosin [Goodno, C. C. (1979) Proc. Natl. Acad. Sci. USA 76, 2620-2624] with actin is smaller than the average association constant of the intermediate states of the actin-SF-1 ATPase cycle. Actin-SF-1 ATPase activity is 90% inhibited by ADP plus vanadate. The reaction of actin with M.ADP.Vi produces a slow release of ADP and vanadate and quantitative recovery of ATPase activity. The rate of dissociation of ligands was almost linear in actin concentration; consequently, the rate constant of dissociation could only be roughly estimated as 0.5-1 sec-1. The rate of dissociation of ADP and vanadate is thus increased by a factor of 10(5) compared to M.ADP.Vi. The rate of release of ligands by regulated actin (actin-tropomyosin-troponin) was reduced to 1/10th to 1/20th by removal of calcium ion. Therefore the M.ADP.Vi complex has the properties of a more stable analogue of the myosin-ADP-phosphate complex that is generated in the normal ATPase cycle. The activation of ligand release (ratio of rate of dissociation of ADP and vanadate from actomyosin relative to myosin) is much larger than the activation of myosin ATPase by actin, whereas the actual rates of the reactions are much slower.