Abstract
We employed QM/MM molecular dynamics (MD) simulations to characterize the rate-limiting step of the glycosylation reaction of pancreatic α-amylase with combined DFT/molecular dynamics methods (PBE/def2-SVP : AMBER). Upon careful choice of four starting active site conformations based on thorough reactivity criteria, Gibbs energy profiles were calculated with umbrella sampling simulations within a statistical convergence of 1–2 kcal ⋅ mol−1. Nevertheless, Gibbs activation barriers and reaction energies still varied from 11.0 to 16.8 kcal ⋅ mol−1 and −6.3 to +3.8 kcal ⋅ mol−1 depending on the starting conformations, showing that despite significant state-of-the-art QM/MM MD sampling (0.5 ns/profile) the result still depends on the starting structure. The results supported the one step dissociative mechanism of Asp197 glycosylation preceded by an acid-base reaction by the Glu233, which are qualitatively similar to those from multi-PES QM/MM studies, and thus support the use of the latter to determine enzyme reaction mechanisms.
| Original language | English |
|---|---|
| Article number | e202200159 |
| Journal | ChemPhysChem |
| Volume | 23 |
| Issue number | 13 |
| DOIs | |
| State | Published - Jul 5 2022 |
| Externally published | Yes |
Funding
This work was supported by UIBD/50006/2020 with funding from FCT/MCTES through national funds. We thank the Oak Ridge National Laboratory – Leadership Computing Facility, which is supported by the Office of Science of the U.S. Department of Energy under Contract No. DE‐AC05‐00OR22725, for the resources granted under the Director's Discretion call project CHM150. We also acknowledge the funding from project PTDC/QUI‐QFI/28714/2017 from FCT/MCTES.
Keywords
- computational chemistry
- conformational space
- density functional theory
- enzyme catalysis
- umbrella sampling simulations