The α- and β-subunits are required for expression of catalytic activity in the hetero-dimeric glucosidase II complex from human liver

The alpha- and beta-subunits of the hetero-dimeric glucosidase II complex from human liver were cloned and expressed in COS-1 cells. The 4106 bp full-length cDNA for the alpha-subunit contained a 2835 bp ORF encoding a 107 kDa polypeptide. The 2095 bp cDNA for the beta-subunit encodes a similar to 60 kDa protein in a continuous 1605 bp ORF, The a- and beta-subunits each contain two potential Asn-Xaa-Thr/Ser acceptor sites, with only one site in the oc-subunit (Asn97) being glycosglated. Additional lambda-clones were isolated for each subunit containing in-frame insertions/deletions within the coding region, indicating alternative splicing. Analysis of different human tissues revealed similar to 4.4 kb and similar to 2.4 kb transcripts for alpha- and beta-subunit, respectively, consistent with their full-length cDNA. Coexpression of the alpha- and beta-subunits in COS-1 cells resulted in >4-fold increase of glucosidase II activity, An inactive protein was obtained, however, after transfection with the alpha-subunit alone, showing that both subunits are essential for expression of active glucosidase II, The observation that the enzyme, previously purified from pig liver and lacking the beta-subunit, was catalytically active indicates that the beta-subunit is involved in alpha-subunit maturation rather than being required for enzymatic activity once the alpha-subunit has acquired its mature form. The alpha-subunit is expressed in COS-1 cells as an ER-located protein, whether inactive or part of a catalytically active complex, This suggests that ER-localization of the alpha-subunit, when associated with the dimeric enzyme complex, is mediated by the C-terminal HDEL-signal in the beta-subunit, whereas the apparently incompletely folded form of the inactive alpha-subunit could be retained in the ER by the putative "glycoprotein-specific quality control machinery".