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 cell line Sf21 has been studied using pyridylaminated oligosaccharides and chromogenic synthetic glycosides as substrates. Ultracentrifugation experiments indicated that the insect cell beta-N-acetylglucosminidase exists in a soluble and a membrane-bound form. This latter form accounted for two-thirds of the total activity and was associated with vesicles of the same density as those containing GlcNAc-transferase I. Partial membrane association of the enzyme was observed with all substrates tested, i.e. 4-nitrophenyl beta-N-acetylglucosaminide, tri-N-acetylchitotriose, and an N-linked biantennary agalactooligosaccharide. Inhibition studies indicted a single enzyme to be responsible for the hydrolysis of all these substrates. With the biantennary substrate, the beta-N-acetylglucosaminidase exclusively removed beta-N-acetylglucosamine from the alpha 1,3-antenna. GlcNAcMan5GlcNAc2, the primary product of GlcNAc-transferase I, was not perceptibly hydrolyzed. beta-N-Acetylglucosaminidases with the same branch specificity were also found in the lepidopteran cell lines Bm-N and Mb-0503. In contrast, beta-N-acetylglucosaminidase activities from rat or frog (Xenopus laevis) liver and from mung bean seedlings were not membrane-bound, and they did not exhibit a strict branch specificity. An involvement of this unusual beta-N-acetylglucosaminidase in the processing of asparagine-linked oligosaccharides in insects is suggested.
Altmann
F
F
Institut für Chemie, Universität für Bodenkultur, Wien, Austria.
Schwihla
H
H
Staudacher
E
E
Glössl
J
J
März
L
L
D000118:Acetylglucosaminidase / Q000502:physiology*; D000818:Animals; D002240:Carbohydrate Sequence; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D018411:Spodoptera / Q000201:enzymology*
D011134:Polysaccharides; D011506:Proteins; D000118:Acetylglucosaminidase