Proton shuttles and phosphatase activity in soluble epoxide hydrolase

J Am Chem Soc. 2007 Jan 17;129(2):387-94. doi: 10.1021/ja066150c.


Recently, a novel metal Mg2+-dependent phosphatase activity has been discovered in the N-terminal domain of the soluble epoxide hydrolase (sEH), opening a new branch of fatty acid metabolism and providing an additional site for drug targeting. Importantly, the sEH N-terminal fold belongs to the haloacid dehalogenase (HAD) superfamily, which comprises a vast majority of phosphotransferases. Herein, we present the results of a computational study of the sEH phosphatase activity, which includes classical molecular dynamics (MD) simulations and mixed quantum mechanical/molecular mechanics (QM/MM) calculations. On the basis of experimental results, a two-step mechanism has been proposed and herein investigated: (1) phosphoenzyme intermediate formation and (2) phosphoenzyme intermediate hydrolysis. Building on our earlier work, we now provide a detailed description of the reaction mechanism for the whole catalytic cycle along with its free energy profile. The present computations suggest metaphosphate-like transition states for these phosphoryl transfers. They also reveal that the enzyme promotes water deprotonation and facilitates shuttling of protons via a metal-ligand connecting water bridge (WB). These WB-mediated proton shuttles are crucial for the activation of the solvent nucleophile and for the stabilization of the leaving group. Moreover, due to the conservation of structural features in the N-terminal catalytic site of sEH and other members of the HAD superfamily, we suggest a generalization of our findings to these other metal-dependent phosphatases.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Catalysis
  • Computer Simulation
  • Enzyme Activation
  • Epoxide Hydrolases / chemistry*
  • Hydrolysis
  • Models, Molecular
  • Molecular Conformation
  • Phosphoric Monoester Hydrolases / chemistry*
  • Protein Structure, Secondary
  • Protons
  • Quantum Theory
  • Solubility


  • Protons
  • Phosphoric Monoester Hydrolases
  • Epoxide Hydrolases