INSECT CELLS CONTAIN AN UNUSUAL, MEMBRANE-BOUND BETA-N-ACETYLGLUCOSAMINIDASE PROBABLY INVOLVED IN THE PROCESSING OF PROTEIN N-GLYCANS

The beta-N-acetylglucosaminidase activity in the lepidopteran insect c ell line Sf21 has been studied using pyridylaminated oligosaccharides and chromogenic synthetic glycosides as substrates. Ultracentrifugatio n experiments indicated that the insect cell beta-N-acetylglucosaminid ase exists in a soluble and a membrane-bound form. This latter form ac counted for two-thirds of the total activity and was associated with v esicles of the same density as those containing GlcNAc transferase I. Partial membrane association of the enzyme was observed with all subst rates tested, i.e. 4-nitrophenyl beta-N-acetylglucosaminide, tri-N-ace tylchitotriose, and an N-linked biantennary agalactooligosaccharide. I nhibition studies indicated a single enzyme to be responsible for the hydrolysis of all these substrates. With the biantennary substrate, th e beta-N-acetylglucosaminidase exclusively removed beta-N-acetylglucos amine from the alpha 1,3-antenna. GlcNAcMan(5)GlcNAc(2), the primary p roduct of GlcNAc-transferase I, was not perceptibly hydrolyzed. beta-N -Acetylglucosaminidases with the same branch specificity were also fou nd in the lepidopteran cell lines Bm-N and Mb-0503. In contrast, beta- N-acetylglucosaminidase activities from rat or frog (Xenopus laevis) l iver and from mung bean seedlings were not membrane-bound, and they di d not exhibit a strict branch specificity. An involvement of this unus ual beta-N-acetylglucosaminidase in the processing of asparagine-linke d oligosaccharides in insects is suggested.