Mechanism and Origin of Stereoselectivity of Halohydrin Dehalogenase-Catalyzed Ring-Expansion Reaction of Spiro-Epoxides.

The detailed mechanism of halohydrin dehalogenase (HHDH)-catalyzed ring-expansion reaction between spiro-epoxides and the nucleophile OCN- to form spiro-oxazolidinones was investigated using molecular docking, molecular dynamics (MD) simulations, and quantum mechanics/molecular mechanics (QM/MM) calculations. Molecular docking and molecular mechanics/Poisson-Boltzmann surface area (MM-PBSA) results revealed that the R-configurational substrate exhibited significantly superior binding affinity compared to the S-configurational one, validating the stable binding mode within the enzyme's active pocket. The fundamental pathway initiates with the nucleophilic attack of OCN- accompanied by proton transfer, which is followed by ring closure coupled with a proton transfer to form a five-membered ring. A total of four possible selective nucleophilic attack pathways were evaluated, and the most energetically favorable pathway associated with the R-configurational product has an energy barrier of 17.1 kcal/mol, which is close to the experimental value of 18.1 kcal/mol derived from kinetic parameters. This study provides valuable theoretical insights for the future engineering of HHDHs for chiral spiro-oxazolidinone synthesis.