Biocatalysis of azidolysis of epoxides: Computational evidences on the role of halohydrin dehalogenase (HheC)

Abstract

Biocatalytic azidolysis of 9 unsymmetrical epoxides by halohydrin dehalogenase enzyme (HheC) in gas phase and uncatalysed azidolysis of the same epoxides in gas phase and in aqueous solution have been modelled at DFT level. Aliphatic epoxides (1–6) and aromatic epoxides (9) undergo β cleavage while styrene oxide (7) and p-nitro styrene (8) oxide prefer α cleavage in the gas phase. Inclusion of aqueous solvation effect via Polarizable Continuum Model (PCM) increases the activation barrier and makes the reaction endothermic due to extensive solvation of azide anion and oxido anionic products, but does not alter the regioselectivity. Halohydrin dehalogenase from Agrobacterium radiobactor AD1 catalyses (E1–E9) ring opening of all these epoxides by azide ion with β selectivity and the reversal of selectivity in epoxide 7 and 8 is notable. These reactions follow, in both enzymatic and non-enzymatic environment, S N 2 mechanism. Calculations while agreeing totally with experimental results offer better insights on the factors determining the regioselectivity and particularly the role of enzyme. Active site model and crystal structure data reveal that the Tyr145 and Ser132 form weak hydrogen bonds with epoxide oxygen lone pair and form reactant enzyme complex (REC). The enzyme complex activates the epoxide ring towards azidolysis. The NBO deletion and second order perturbation analyses clearly bring out the role of catalytic duo Tyr145 and Ser132 and particularly shed light on the dominant contribution of Tyr145 in selectively activating C β –O bond. The present results indicate that Arg149 or other residues in the pocket do not seem to have any significant effect on the reaction. The bio-catalysed and uncatalysed ring opening of various epoxides by azide anion have been modelled at B3LYP/6-31+g(2d, p) level. Nine epoxides that include six aliphatic (1–6) and three aromatic (7–9) epoxides have been used for the study and experimental reports where available have been compared.