Isolation of a homogeneous glucosidase II from pig kidney microsomes

Abstract

The processing of the oligosaccharide precursor chain, (GlcNAc)2(Man)9(Gle)3, of N‐glycosylated glyco‐proteins starts with the action of glucosidase I which excises the terminal (α1–2)‐linked glucose residue. Glucosidase II removes the two inner (α1–3)‐linked glucose residues. We have purified glucosidase II to homogeneity from pig kidney microsomes. The enzyme is a glycoprotein and contains a single type of subunit of molecular mass ∼ 100 kDa. The native enzyme is probably a tetramer. It cleaves glucosidic α1–3 and α1–4, but not α1–1, α1–2 or α1–6 bonds and lacks 2‐mannosidase and glucosidase I activity. The pH optimum is between 6.0 and 7.5. Specific antibodies against the native enzyme and the denatured subunit were prepared. By activity measurements and immune blotting, a similar enzyme was found in rat liver. In the fractionated rat liver, the enzyme was localized in the lumen of the endoplasmic reticulum, probably loosely bound to the inner face of the membrane. Purified Golgi fractions contained only low levels of the enzyme. Copyright © 1984, Wiley Blackwell. All rights reserved