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Minor folding defects trigger local modification of glycoproteins by the ER folding sensor GT
Author(s) -
Ritter Christiane,
Quirin Katharina,
Kowarik Michael,
Helenius Ari
Publication year - 2005
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1038/sj.emboj.7600645
Subject(s) - biology , folding (dsp implementation) , protein folding , glycoprotein , minor (academic) , biophysics , posttranslational modification , microbiology and biotechnology , biochemistry , enzyme , engineering , political science , law , electrical engineering
UDP‐glucose:glycoprotein glucosyltransferase (GT) is a key component of the glycoprotein‐specific folding and quality control system in the endoplasmic reticulum. By exclusively reglucosylating incompletely folded and assembled glycoproteins, it serves as a folding sensor that prolongs the association of newly synthesized glycoproteins with the chaperone‐like lectins calnexin and calreticulin. Here, we address the mechanism by which GT recognizes and labels its substrates. Using an improved inhibitor assay based on soluble conformers of pancreatic ribonuclease in its glycosylated (RNase B) and unglycosylated (RNase A) forms, we found that the protein moiety of a misfolded conformer alone is sufficient for specific recognition by GT in vitro . To investigate the relationship between recognition and glucosylation, we tested a variety of glycosylation mutants of RNase S‐Protein and an RNase mutant with a local folding defect [RNase C65S, C72S], as well as a series of loop insertion mutants. The results indicated that local folding defects in an otherwise correctly folded domain could be recognized by GT. Only glycans attached to the polypeptide within the misfolded sites were glucosylated.