Inhibition of myosin ATPase by metal fluoride complexes

Magnesium (Mg2+) is the physiological divalent cation stabilizing nucleotid e or nucleotide analog in the active site of myosin subfragment 1 (S1). In the presence of fluoride, Mg(_)(2+)and MgADP form a complex that traps the active site of S1 and inhibits myosin ATPase. The ATPase inactivation rate of the magnesium trapped S1 is comparable but smaller than the other known gamma-phosphate analogs at 1.2 M-1 s(-1) with 1 mM MgCl2. The observed mola r ratio of Mg/S1 in this complex of 1.58 suggests that magnesium occupies t he gamma-phosphate position in the ATP binding site of S1 (S1-MgADP-MgFx). The stability of S1-MgADP-MgFx at 4 degrees C was studied by EDTA chase exp eriments but decomposition was not observed. However, removal of excess flu oride causes full recovery of the K+-EDTA ATPase activity indicating that f ree fluoride is necessary for maintaining a stable trap and suggesting that the magnesium fluoride complex is bonded to the bridging oxygen of beta-ph osphate more loosely than the other known phosphate analogs. The structure of S1 in S1-MgADP-MgFx, was studied with near ultraviolet circular dichrois m, total tryptophan fluorescence, and tryptophan residue 510 quenching meas urements. These data suggest that S1-MgADP-MgFx resembles the M**. ADP Pi s teady-state intermediate of myosin ATPase. Gallium fluoride was found to co mpete with MgFx, for the gamma-phosphate site in S1-MgADP-MgFx. The ionic r adius and coordination geometry of magnesium, gallium and other known gamma -phosphate analogs were compared and identified as important in determining which myosin ATPase intermediate the analog mimics. (C) 1999 Elsevier Scie nce B.V. All rights reserved.