Expression, purification, and characterization of highly active endo-α-N-acetylgalactosaminidases expressed by silkworm-baculovirus expression system

Abstract The O -glycosidase, endo-α- N -acetylgalactosaminidase from Enterococcus faecalis (endoEF) catalyzes the cleavage of core 1 and core 3 type O -linked disaccharides between GalNAc and serine or threonine residues from glycoproteins. The endoEF has broad substrate specificity and thus is extensively utilized for the structural and functional analysis of the O -linked glycans. In this study, we expressed and purified the recombinant endoEF (rEndoEF) by using the silkworm-baculovirus expression vector system (Silkworm-BEVS) and confirmed the deglycosylation activity of rEndoEF targeting reporter glycoproteins, which was equivalent to the commercial O -glycosidase. Thus, our study provides important clues to produce highly active rEndoEF O -glycosidases employing silkworm-BEVS as an alternative.