Insight into the phosphodiesterase mechanism from combined QM/MM free energy simulations

Kin Yiu Wong, Jiali Gao

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18 Scopus citations


Molecular dynamics simulations employing a combined quantum mechanical and molecular mechanical potential have been carried out to elucidate the reaction mechanism of the hydrolysis of a cyclic nucleotide cAMP substrate by phosphodiesterase 4B (PDE4B). PDE4B is a member of the PDE superfamily of enzymes that play crucial roles in cellular signal transduction. We have determined a two-dimensional potential of mean force (PMF) for the coupled phosphoryl bond cleavage and proton transfer through a general acid catalysis mechanism in PDE4B. The results indicate that the ring-opening process takes place through an SN2 reaction mechanism, followed by a proton transfer to stabilize the leaving group. The computed free energy of activation for the PDE4B-catalyzed cAMP hydrolysis is about 13 kcal·mol-1 and an overall reaction free energy is about -17 kcal·mol-1, both in accord with experimental results. In comparison with the uncatalyzed reaction in water, the enzyme PDE4B provides a strong stabilization of the transition state, lowering the free energy barrier by 14 kcal·mol -1. We found that the proton transfer from the general acid residue His234 to the O3′ oxyanion of the ribosyl leaving group lags behind the nucleophilic attack, resulting in a shallow minimum on the free energy surface. A key contributing factor to transition state stabilization is the elongation of the distance between the divalent metal ions Zn2+ and Mg 2+ in the active site as the reaction proceeds from the Michaelis complex to the transition state.

Original languageEnglish (US)
Pages (from-to)2579-2595
Number of pages17
JournalFEBS Journal
Issue number14
StatePublished - Jul 2011


  • QM/MM on the fly
  • ensemble-average structure analysis
  • free-energy simulations
  • phosphate hydrolysis
  • phosphodiesterase

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