The hydrolysis activity of adenosine triphosphate in myosin: a theoretical analysis of anomeric effects and the nature of the transition state.

Combined quantum mechanical/molecular mechanical (QM/MM) calculations with density functional theory are employed to analyze two issues related to the hydrolysis activity of adenosine triphosphate (ATP) in myosin. First, we compare the geometrical properties and electronic structure of ATP in the open (post-rigor) and closed (pre-powerstroke) active sites of the myosin motor domain. Compared to both solution and the open active site cases, the scissile P(gamma)-O(3beta) bond of ATP in the closed active site is shown to be substantially elongated. Natural bond orbital (NBO) analysis clearly shows that this structural feature is correlated with the stronger anomeric effects in the closed active site, which involve charge transfers from the lone pairs in the nonbridging oxygen in the gamma-phosphate to the antibonding orbital of the scissile bond. However, an energetic analysis finds that the ATP molecule is not significantly destabilized by the P(gamma)-O(3beta) bond elongation. Therefore, despite the notable perturbations in the geometry and electronic structure of ATP as its environment changes from solution to the hydrolysis-competent active site, ground-state destabilization is unlikely to play a major role in enhancing the hydrolysis activity in myosin. Second, two-dimensional potential energy maps are used to better characterize the energetic landscape near the hydrolysis transition state. The results indicate that the transition-state region is energetically flat and a range of structures representative of different mechanisms according to the classical nomenclature (e.g., "associative", "dissociative", and "concerted") are very close in energy. Therefore, at least in the case of ATP hydrolysis in myosin, the energetic distinction between different reaction mechanisms following the conventional nomenclature is likely small. This study highlights the importance of (i) explicitly evaluating the relevant energetic properties for determining whether a factor is essential to catalysis and (ii) broader explorations of the energy landscape beyond saddle points (even on free-energy surface) for characterizing the molecular mechanism of catalysis.