Theoretical insights into the catalytic mechanism of β-hexosaminidase

The enzyme (β-Hex) has a specific role in the degradation of a particular type of glycolipid, the GM2 ganglioside. This specificity is also manifested by its unusual mechanism of substrate-assisted catalysis, which is considered to be an alternative pathway for the breakage of glycosidic bonds. We have studied its catalytic mechanism using a small model of the enzyme: substrate Michaelis complex with DFT and MP2/MP3 methods. The results reflect the intrinsic chemical reactivity of the active site, decoupled from the long-range enzyme electrostatic field. The mechanism supports, and adds further atomic detail, on the earlier mechanistic suggestions based on experimental data. Moreover, we also have compared the geometry and full potential energy surface obtained with nine different exchange–correlation functionals. It was surprising that the B3LYP activation energies were lower than the ones from some of the hybrid meta functionals (known by their excellent performance on kinetics) by as much as 10 kcal/mol and lower than the MP2 energies by more than 12 kcal/mol. It is known that B3LYP underestimates barriers but underestimations of this extent are unusual and surprising. The effect of the theoretical method on the geometry, usually supposed to be less significant, had in some cases a relevant influence in the mechanism. Therefore, a more thoughtful choice should be made when choosing a methodology for geometry optimization.