Dextranase (α‐1,6 glucan‐6‐glucanohydrolase) from Penicillium minioluteum expressed in Pichia pastoris: two host cells with minor differences in N‐glycosylation

Differences in glycosylation between the natural α-1,6 glucan-6-glucanohydrolase from Penicillium minioluteum and the heterologous protein expressed in the yeast Pichia pastoris were analyzed. Glycosylation profiling was carried out using fluorophore-assisted carbohydrate electrophoresis and amine absorption high-performance liquid chromatography (NH2-HPLC) in combination with matrix-assisted laser desorption-time of flight-mass spectrometry. Both microorganisms produce only oligomannosidic type structures, but the oligosaccharide population differs in both enzymes. The native enzyme has mainly short oligosaccharide chains ranging from Man5GlcNAc2 to Man9GlcNAc2, of which Man8GlcNAc2 was the most represented oligosaccharide. The oligosaccharides linked to the protein produced in P. pastoris range from Man7GlcNAc2 up to Man14GlcNAc2, with Man8GlcNAc2 and Man9GlcNAc2 being the most abundant structures. In both enzymes the first glycosylation site (Asn5) is always glycosylated. However, Asn537 and Asn540 are only partially glycosylated in an alternate manner.