Proton Tunneling Accelerates ATP Hydrolysis in Eg5 Kinesin

Abstract

In ATP hydrolysis, a proton from the water nucleophile must be abstracted and transferred in order to create a hydroxide capable of attacking the substrate. Herein, solvent kinetic isotope experiments with Eg5 kinesin show unanticipated accelerated proton transfer involving an active-site water cluster. The positive kinetic isotope effect (KIE) confirms proton abstraction from water commits kinesin to catalysis and its pH-dependence verifies that switch salt-bridge residues direct chemotransduction. Additionally, a classical description for this proton transfer is refuted by the KIE magnitude, temperature-independent Arrhenius pre-exponential factor ratios, and activation energy differences. Taken together, we conclude that the first step in Eg5 catalysis has a tunneling component, a quantum mechanical event by which a particle transfers through a reaction barrier. This first detection of tunneling in an ATPase is of consequence for two reasons. First, proton tunneling is likely widespread in biomolecules, rather than solely a characteristic of metalloenzymes. Second, energy barrier penetration by proton tunneling is an alternate explanation to classical transition-state stabilization theory for the fast reactivities of motor proteins.