Komatsu_2017_Biochem.Biophys.Res.Commun_490_849

2,3-Butandione 2-monoxime (BDM) is a widely used myosin inhibitor with an unclear mode of action. In this report, we investigated the mechanism of BDM oxime group nucleophilic reactivity on the phosphoester bond of ATP. BDM increased the ATPase activity of skeletal myosin subfragment 1 (S1) under conditions in which ATP cleavage is the rate-limiting step (K(+), EDTA-ATPase activity of native S1 and Mg(2+)-ATPase activity of trinitrophenylated S1 and partially unfolded S1). Furthermore, the effect of BDM on the S1-bound adenosine 5'-(beta,gamma-imido) triphosphate (AMPPNP) (31)P NMR spectrum suggests that BDM changes the microenvironment around the phosphorus atoms of myosin-bound nucleotide. A computational search for the BDM-binding site in the adenosine 5'-[gamma-thio] triphosphate (myosin-ATPgammaS) complex predicted that BDM is located adjacent to the nucleotide on myosin. Therefore, we propose that the BDM oxime group catalytically assists in ATP cleavage, thereby enhancing the ATPase activity of myosin in a manner analogous to pralidoxime-mediated reactivation of organophosphate-inactivated acetylcholinesterase. This is the first study suggesting that oxime provides catalytic assistance for ATP cleavage by an ATP-hydrolyzing enzyme.