Purification and characterization of hen oviduct alpha 1,2-mannosidase.
An alpha-mannosidase capable of hydrolyzing three Man alpha 1,2-residues from pyridylamine-(PA-) labeled Man9GlcNAc2 was purified from hen oviduct. The purity of the preparation was analyzed by PAGE; its molecular weight was 42,000 by SDS-PAGE or 50,000 by gel filtration. The pH optimum was 6.5. The enzyme was inactivated with EDTA; enzyme activity was restored by the addition of Ca2+. The enzyme activity was inhibited by 1-deoxymannojirimycin, but not by swainsonine. The substrate specificity of the purified enzyme was analyzed using PA-oligomannose-type sugar chains. When Man9GlcNAc2-PA was digested, Man alpha 1-6(Man alpha 1-2Man alpha 1-3)Man alpha 1-6(Man alpha 1-3)Man beta 1-4GlcNAc beta 1-4Glc-NAc-PA was obtained as an end product, and the enzyme was incapable of hydrolyzing p-nitrophenyl alpha-D-mannoside and Man alpha 1,3- or Man alpha 1,6-residues. Judging from these characteristics, the enzyme was classified as a Man9-mannosidase or Golgi mannosidase I and speculated to participate in the processing or catabolism of glycoproteins.
Hamagashira
N
N
Department of Chemistry, Osaka University College of Science.
Oku
H
H
Mega
T
T
Hase
S
S
D017485:1-Deoxynojirimycin / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002240:Carbohydrate Sequence; D002645:Chickens; D004492:Edetic Acid / Q000494:pharmacology; D005260:Female; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008361:Mannosidases / Q000302:isolation & purification* / Q000378:metabolism; D008362:Mannosides / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism*; D010057:Oviducts / Q000201:enzymology*; D013379:Substrate Specificity
D008362:Mannosides; D009844:Oligosaccharides; D017485:1-Deoxynojirimycin; D004492:Edetic Acid; D008361:Mannosidases; C046090:mannosyl-oligosaccharide 1,2-alpha-mannosidase; D002118:Calcium