Heterologous expression and kinetic characterisation of Neurospora crassa β-xylosidase in Pichia pastoris

•We report the secretion of Neurospora crassa β-xylosidase from Pichia pastoris.•Recombinant β-xylosidase has high catalytic capacity against natural substrates.•The reaction product xylose is a non-competitive inhibitor of the β-xylosidase.•Transxylosilation activity of the enzyme has been identified.

To degrade plant hemicelluloses fungi employ β-xylosidases to hydrolyse xylooligosaccharides, released by endo-xylanases, into xylose. We have expressed the β-xylosidase from Neurospora crassa in Pichia pastoris under the control of alcohol oxidase 1 (AOX1) promoter. The recombinant enzyme is optimally active at 50 °C and pH 5.0 with Km and Vmax values of 8.9 mM and 1052 μmol min−1 mg−1 respectively against 4-nitrophenyl β-xylopyranoside. Xylose is a non-competitive inhibitor with a Ki of 1.72 mM. The enzyme is characterised to be an exo-cutting enzyme releasing xylose from the non-reducing ends of β-1,4 linked xylooligosaccharides (X2, X3 and X4) but also capable of transxylosilation. Catalytic conversion of X2, X3 and X4 decreases (Vmax and kcat) with increasing chain length.