Elucidating microscopic events driven by GTP hydrolysis reaction in the Ras–GAP system with semi-reactive molecular dynamics simulations: the alternative role of a phosphate binding loop for mechanical energy storage

Abstract

ATPase and GTPase have been widely found as chemical energy–mechanical work transducers, whereas the physicochemical mechanisms are not satisfactorily understood. We addressed the problem by examining John Ross’ conjecture that repulsive Coulomb interaction between ADP/GDP and inorganic phosphate (P_i) does the mechanical work upon the system. We effectively simulated the consequence of a GTP hydrolysis reaction in a complex system of Rat sarcoma (Ras) and GTPase activation protein (GAP) in the framework of classical molecular dynamics by switching force field parameters between the reactant and product systems. We then observed a ca. 5 kcal mol⁻¹ increase of potential energy about the phosphate-binding loop (P-loop) in the Ras protein, indicating that the mechanical work generated via the GTP hydrolysis is converted into the local interaction energy and stored in the P-loop. Interestingly, this local energy storage in the P-loop depends on neither impulsive nor consecutive collisions of GDP and P_i with the P-loop. Instead, GTP–GDP conversion itself does work on the Ras system, elevating the potential energy. These observations encourage us to challenge a conjecture previously given by Ross. We assert that triphosphate nucleotide hydrolyses do mechanical work by producing emergent steric interaction accompanied by relaxation, namely, a shift of the biomolecular system to the non-equilibrium state on the reshaped potential energy landscape. Recalling the universality of the P-loop motif among GTPases and ATPases, the observations that we obtained through this study would progress the physicochemical understanding of the operating principles of GTP/ATP hydrolysis-driven biological nano-machines.