Recovery of recombinant β-glucosidase by expanded bed adsorption from Pichia pastoris high-cell-density culture broth

Abstract

Methanol limited fed-batch cultivation was applied for production of a plant derived β-glucosidase by Pichia pastoris. The β-glucosidase was recovered by expanded bed adsorption chromatography applied to the whole culture broth. The new Streamline Direct HST1 adsorbent was compared with Streamline SP. Higher bead density made it possible to operate at two times higher feedstock concentration and at two times higher flow velocity. The higher binding capacity in the conductivity range 0–48 mS cm −1 of Streamline Direct HST1 might be caused by the more complex interaction of multi-modal ligand in Streamline Direct HST1 compared to the single sulphonyl group in Streamline SP. Harsher elution condition had to be applied for dissociation of β-glucosidase from Streamline Direct HST1 due to stronger binding interaction. The 5% dynamic binding capacity was 160 times higher for Streamline Direct HST1 compared to Streamline SP. The yield of β-glucosidase on Streamline Direct HST1 (74%) was significantly higher than on Streamline SP (48%). Furthermore, β-glucosidase was purified with a factor of 4.1 and concentrated with a factor of 17 on Streamline Direct HST1 while corresponding parameters were half of these values for Streamline SP. Thus, for all investigated parameters Streamline Direct HST1 was a more suitable adsorbent for recovery of recombinant β-glucosidase from unclarified P. pastoris high-cell-density cultivation broth.