N-linked oligosaccharide processing enzyme glucosidase II produces 1,5-anhydrofructose as a side product.
alpha-1,4-Glucan lyase cleaves alpha-1,4-linkages of nonreducing termini of alpha-1,4-glucans to produce 1,5-anhydrofructose (1,5-AnFru). The enzymes isolated from fungi and algae show high homology with glycoside hydrolase family 31. Purification of alpha-1,4-glucan lyase from rat liver using DEAE Cellulose chromatography resulted in separation of two enzymatic active fractions, one was bound to the column and the other was in the flow-through. Partial amino acid sequence determined from the lyase, retained on the anion exchange column, were identical with that of the N:-linked oligosaccharide processing enzyme glucosidase II. The lyase showed similar enzymatic properties as the microsomal glucosidase such as inhibition by 1-deoxynojirimycin and castanospermine. On the other hand, glucosidase II purified from rat liver microsomes produced not only glucose but also a small amount of 1,5-AnFru using maltose as substrate. Furthermore, CHO cells overexpressing pig liver glucosidase II showed a 1.5- to 2-fold higher lyase activity compared to the nontransfected CHO cells. Conversely, no lyase activity was detectable either in PHAR2.7, the glucosidase II-deficient mutant from a mouse lymphoma cell line, or in Saccharomyces cerevisiae strain YG427 having the glucosidase II gene disrupted. These data demonstrate that glucosidase II possesses an additional enzymatic activity of releasing 1,5-AnFru from maltose.
Hirano
K
K
Department of Life Sciences (Chemistry), Graduate School of Arts and Sciences, The University of Tokyo, Komaba, Meguro-ku, Tokyo 153-8902, Japan.
Ziak
M
M
Kamoshita
K
K
Sukenaga
Y
Y
Kametani
S
S
Shiga
Y
Y
Roth
J
J
Akanuma
H
H
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D005632:Fructose / Q000031:analogs & derivatives / Q000096:biosynthesis*; D008862:Microsomes, Liver / Q000201:enzymology; D009844:Oligosaccharides / Q000378:metabolism*; D011133:Polysaccharide-Lyases / Q000737:chemistry / Q000378:metabolism; D000520:alpha-Glucosidases / Q000737:chemistry / Q000378:metabolism*
D009844:Oligosaccharides; D005632:Fructose; C048927:1,5-anhydrofructose; C034251:4-nitrophenyl-alpha-glucosidase; D000520:alpha-Glucosidases; D011133:Polysaccharide-Lyases; C080474:alpha-1,4-glucan lyase