Role of Glucosidase II beta subunit in the quality control of glycoprotein folding

IVAN STIGLIANO; CARLOS LABRIOLA; ARMANDO J. PARODI; CECILIA D'ALESSIO

Villa Carlos Paz, Córdoba, Argentina

Congreso; XLIV reunión de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB); 2008

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB)

Glucosidase II (GII) is a key player of the glycoprotein folding quality control in the lumen of the endoplasmic reticulum (ER). By this mechanism, misfolded glycoproteins are retained in the ER until proper folding is achieved or the protein is degraded. GII catalyzes the removal of the two innermost Glc residues of the Glc3Man9GlcNAc2 oligosaccharides transferred to proteins during N-glycosylation. GII is a soluble protein composed of subunits GII alpha and GII beta. GII alpha holds the catalytic activity site and it lacks any known ER retrieval sequence. This last sequence is present in GII beta, whose function is controversial. Microsomal fraction prepared from fission yeast Schizosaccharomyces pombe mutant cells GII alpha/GII beta minus expressing GII alpha with the ER retrieval signal VDEL hydrolyzes the substrate analogue p-NPG. Western blot analysis revealed that GIIalpha-VDEL is retained in the ER. This demonstrates that in S. pombe GII beta is responsible for GII alpha retention in the ER but not for its folding or maturation. In vivo oligosaccharide labeling in GII alpha/GII beta minus mutants expressing GIIalpha-VDEL and in vitro activity assays of microsomes from these cells using physiological substrates demonstrate that cells lacking GII beta are totally and partially impaired of removing Glc from Glc1Man9GlcNAc and Glc2Man9GlcNAc, respectively. This suggests that GII beta is responsible for oligosaccharide structure recognition by GII.