Effect of glycosylation of β-glucuronidase on its catalytic properties in ionic liquids

The purification of glycosylated recombinant β-glucuronidase from Pichia pastoris GS115 (PGUS-P) was investigated by a novel two-step process: ammonium sulfate precipitation and molecular sieve chromatography. The highest purification fold obtained was 66.79. The catalytic properties of glycosylated PGUS-P in hydrophobic ionic liquids (ILs)/buffer biphasic system were investigated. A 2.2-fold enhancement in the catalytic efficiency was observed using 50% (v/v) 1-butyl-3-methylimidazolium hexafluorophosphate, in comparison with the acetate buffer medium. When compared with the glycosylated PGUS-P, the deglycosylated enzyme at T₅₅ and T₆₅ exhibited low activity and low thermal stability in both ILs and acetate buffer. It was also observed that ILs had effect on the pH profile of deglycosylated PGUS-P - the optimum pH was extended from 5.0 (acetate buffer) to 5.0-7.0 (ILs). Therefore, this study indicates that the glycosylation of PGUS-P plays an important role in both catalytic activity and stability in hydrophobic ionic liquids (ILs)/buffer biphasic system.