N‐glycosylation affects the proper folding, enzymatic characteristics and production of a fungal β‐glucosidase

ABSTRACT Heterologous expression of β‐glucosidase is one of the approaches to enhance the efficiency of fungal cellulase preparations. It has been reported that N ‐glycosylation affects the structure framework, function and stability of proteins. In this study, a β‐glucosidase from Aspergillus terreus (GenBank: XP_001216552, BglS) was heterologously expressed in Pichia pastoris and Trichoderma reesei . The four asparagine residues were all linked with high‐mannose‐type oligosaccharides in P. pastoris , whereas only N224 carried high‐mannose‐type glycan in T. reesei (the other three sites carried one N ‐acetylglucosamine). The long N ‐glycan chains on PpBglS weakened its substrate affinity, activity and thermostability. The moderate post‐translational and post‐secretory glycan modification in T. reesei makes it a suitable expression system for BglS. The N224 glycan played a critical role in BglS folding. The elucidation of the correlation between the different N ‐glycosylation patterns of BglS and their corresponding enzymatic characteristics is an important step towards improving the activity, thermostability and even production of heterologous β‐glucosidase by glycan engineering. Biotechnol. Bioeng. 2013;110: 3075–3084. © 2013 Wiley Periodicals, Inc.