Mg2+ Dependent Modulation of Striated Muscle Myosin ATPase by Thin Filament Components

While it's been reported that non-muscle myosin such as myosin V can be regulated by Mg2+, striated muscle myosin modulation by Mg2+-concentration dependency, especially in relation to thin filament components, has not been well studied up to now. It has been suggested that ADP release (product release) from myosin V is affected by Mg2+ concentration. Therefore, Mg2+-dependent modulation of striated muscle myosin ATPase was explored in the presence of reconstituted muscle thin filament components, i.e. actin, tropomyosin and troponin. ATPase activity assays and Nuclear Magnetic Resonance (NMR) spectroscopy were employed to understand the mechanisms of ATP hydrolysis by striated muscle myosin and the role of Mg2+ in the enzymatic activity. An NMR technique called Water Ligand-Observed Gradient Spectroscopy (WaterLOGSY) was utilized to probe if the interaction of myosin with ADP (one of the products resulted from ATP hydrolysis) is different depending on Mg2+ concentration as well as the presence of muscle thin filament components. Our WaterLOGSY data demonstrated that this technique is a promising method to monitor ADP binding to myosin head S1 even in the presence of actin filaments and that there was a change in the appearance of peaks between with and without tropomyosin at a low Mg2+ concentration. This set of results indicates that the way by which myosin binds ADP is different depending on the states of thin filaments. Moreover, actomyosin S1 ATPase rate decreased as Mg2+ concentration increased. When tropomyosin was reconstituted in the thin filaments, the ATPase rate decreased more rapidly compared with actomyosin alone. Thus we conclude that Mg2+ plays an important role in striated muscle myosin ATPase and that regulatory components such as tropomyosin modulate Mg2+ dependency of myosin ATPase in the striated muscle.