Human OS-9, a lectin required for glycoprotein endoplasmic reticulum-associated degradation, recognizes mannose-trimmed N-glycans.
In the endoplasmic reticulum (ER), lectins and processing enzymes are involved in quality control of newly synthesized proteins for productive folding as well as in the ER-associated degradation (ERAD) of misfolded proteins. ER quality control requires the recognition and modification of the N-linked oligosaccharides attached to glycoproteins. Mannose trimming from ... the N-glycans plays an important role in targeting of misfolded glycoproteins for ERAD. Recently, two mammalian lectins, OS-9 and XTP3-B, which contain mannose 6-phosphate receptor homology domains, were reported to be involved in ER quality control. Here, we examined the requirement for human OS-9 (hOS-9) lectin activity in degradation of the glycosylated ERAD substrate NHK, a genetic variant of alpha1-antitrypsin. Using frontal affinity chromatography, we demonstrated that the recombinant hOS-9 mannose 6-phosphate receptor homology domain specifically binds N-glycans lacking the terminal mannose from the C branch in vitro. To examine the specificity of OS-9 recognition of N-glycans in vivo, we modified the oligosaccharide structures on NHK by overexpressing ER alpha1,2-mannosidase I or EDEM3 and examined the effect of these modifications on NHK degradation in combination with small interfering RNA-mediated knockdown of hOS-9. The ability of hOS-9 to enhance glycoprotein ERAD depended on the N-glycan structures on NHK, consistent with the frontal affinity chromatography results. Thus, we propose a model for mannose trimming and the requirement for hOS-9 lectin activity in glycoprotein ERAD in which N-glycans lacking the terminal mannose from the C branch are recognized by hOS-9 and targeted for degradation.
Mesh Terms:
Amino Acid Substitution, Base Sequence, Binding Sites, Carbohydrate Sequence, Cell Line, Endoplasmic Reticulum, Glycoproteins, Humans, Kinetics, Lectins, Mannose, Mannose-Binding Lectins, Molecular Sequence Data, Mutagenesis, Site-Directed, Neoplasm Proteins, Polysaccharides, Protein Folding, Protein Structure, Tertiary, Proteins, RNA, Small Interfering, Recombinant Proteins, alpha 1-Antitrypsin
Amino Acid Substitution, Base Sequence, Binding Sites, Carbohydrate Sequence, Cell Line, Endoplasmic Reticulum, Glycoproteins, Humans, Kinetics, Lectins, Mannose, Mannose-Binding Lectins, Molecular Sequence Data, Mutagenesis, Site-Directed, Neoplasm Proteins, Polysaccharides, Protein Folding, Protein Structure, Tertiary, Proteins, RNA, Small Interfering, Recombinant Proteins, alpha 1-Antitrypsin
J. Biol. Chem.
Date: Jun. 19, 2009
PubMed ID: 19346256
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