CaATP as a substrate to investigate the myosin lever arm hypothesis of force generation

In an effort to test the lever arm model of force generation, the effects o f replacing magnesium with calcium as the ATP-chelated divalent cation were determined for several myosin and actomyosin reactions, The isometric forc e produced by glycerinated muscle fibers when CaATP is the substrate is 20% of the value obtained with MgATP, For myosin subfragment 1 (S1), the degre e of lever arm rotation, determined using transient electric birefringence to measure rates of rotational Brownian motion in solution, is not signific antly changed when calcium replaces magnesium in an S1-ADP-vanadate complex . Actin activates S1 CaATPase activity, although less than it does MgATPase activity, The increase in actin affinity when S1 . CaADP . P-1 is converte d to S1 . CaADP is somewhat greater than it is for the magnesium case. The ionic strength dependence of actin binding indicates that the change in app arent electrostatic charge at the acto-S1 interface for the S1 . CaADP . P- i to S1 . CaADP step is similar to the change when magnesium is bound. In g eneral, CaATP is an inferior substrate compared to MgATP, but all the data are consistent with force production by a lever arm mechanism for both subs trates, Possible reasons for the reduced magnitude of force when CaATP is t he substrate are discussed.