Unusual uniformity of the N-linked oligosaccharides of HLA-A, -B, and -C glycoproteins.

Barber, Linda D.; Patel, T.P.; Percival, L; Gumperz, Jenny E.; Lanier, Lewis L.; Phillips, Joseph H.; Bigge, J C; Wormwald, M R; Parekh, Raj B.; Parham, Peter

doi:10.4049/jimmunol.156.9.3275

Cited by 43 publications

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“…Heavy chains that carry a single N ‐acetylglucosamine are therefore not a substrate for mammalian N ‐glycanase. Treatment of class I heavy chains with with jack bean mannosidase yielded a preparation with truncated oligosaccharides, predicted to be a Manβ1–4GlcNAc–GlcNAc core trisaccharide (Figure 8, lanes 6 and 13) (Barber et al ., 1996). No shift in isoelectric point was seen after mannosidase treatment.…”

Section: Resultsmentioning

confidence: 99%

“…Cytosolic PNG1 is therefore unlikely to encounter glycoproteins bearing complex type N ‐glycans. The complex type oligosaccharide of properly assembled human MHC class I chain is α1,6 fucosylated in the Golgi (Barber et al ., 1996), which may sterically hinder N ‐glycanase from attacking complex oligosaccharides of this type. While some of the experiments shown were carried out using partially purified mammalian PNG1, we also observed deglycosylation of class I MHC molecules and TCRα using the purified yeast PNG1 enzyme.…”

Section: Discussionmentioning

confidence: 99%

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A role for N-glycanase in the cytosolic turnover of glycoproteins

Hirsch

Blom

Ploegh

2003

EMBO J

202

190

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Successful maturation determines the intracellular fate of secretory and membrane proteins in the endoplasmic reticulum (ER). Failure of proteins to fold or assemble properly can lead to their retention in the ER and redirects them to the cytosol for degradation by the proteasome. Proteasome inhibitors can yield deglycosylated cytoplasmic intermediates that are the result of an N-glycanase activity, believed to act prior to destruction of these substrates by the proteasome. A gene encoding a yeast peptide:N-glycanase, PNG1, has been cloned, but this N-glycanase and its mammalian homolog were reported to be incapable of deglycosylating full-length glycoproteins. We show that both the yeast PNG1 enzyme and its mammalian homolog display N-glycanase activity towards intact glycoproteins. As substrates, cytosolic PNGase activity prefers proteins containing high-mannose over those bearing complex type oligosaccharides. Importantly, PNG1 discriminates between non-native and folded glycoproteins, consistent with a role for N-glycanase in cytoplasmic turnover of glycoproteins.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

A role for N-glycanase in the cytosolic turnover of glycoproteins

Hirsch

Blom

Ploegh

2003

EMBO J

202

190

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Effect of glycosylation on structure and dynamics of MHC class I glycoprotein: A molecular dynamics study

Mandal

Mukhopadhyay

2001

Biopolymers

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Complex carbohydrates linked to glycoproteins are recently being implicated to play a variety of biological roles. The lack of well‐resolved crystallographic coordinates of the carbohydrates makes it difficult to assess the contributions of the glycan chain on protein structure and dynamics. We have modeled two different oligosaccharides NeuNAc2Gal3Man3GlcNAc5Fuc and Man3GlcNAc4 to generate two glycosylation variants of major histocompatibility complex (MHC) class I glycoprotein. Molecular dynamics simulations of the isolated fourteen‐ and seven‐residue oligosaccharides have been done in vacuo and in solution. The dynamics of the two glycoforms of MHC class I protein have been simulated in solution in the free as well as in the peptide‐bound form. Good agreement between the calculated solution conformations of the oligosaccharides in isolated and conjugated forms and the average conformations obtained from x‐ray or NMR data was observed for most of the glycosidic linkages. These molecular dynamics simulations of the isolated glycan chains and the glycoconjugates reveal the details of the conformational flexibility of the glycan chains; they also provide atomic level details of protein–carbohydrate interactions and the effect of the ligand binding on the carbohydrate structure and dynamics. It was found that though there is some flexibility in some of the glycosidic linkages in the isolated oligosaccharides, in the protein‐conjugated form the linkages adopt more restricted conformations. The glycan chains protrude out into the solvent and might hinder the lateral association of the proteins. The presence of the bulky glycan chains does not affect the average backbone fold of the protein but induces local changes in protein structure and dynamics. It has been noted that the extent of the changes depends upon the nature of the attached glycan chain. The glycan chains do not appear to influence the peptide binding property of the protein directly, but may stabilize the protein residues that are involved in ligand binding. © 2001 John Wiley & Sons, Inc. Biopolymers 59: 11–23, 2001

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Disparate folding and stability of the ankylosing spondylitis–associated HLA–B1403 and B2705 proteins

Merino¹,

Galocha²,

Vázquez³

et al. 2008

Arthritis & Rheumatism

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Objective. To investigate the folding, assembly, maturation, and stability of HLA-B*1402 and B*1403, which differ by 1 amino acid change and are differentially associated with ankylosing spondylitis (AS), and to compare these features with those of B*2705.Methods. Stable transfectants expressing B*1402, B*1403, and B*2705 were used. Folding rates were estimated from the ratio of unfolded heavy chains to folded heavy chains that had been immunoprecipitated with specific antibodies in pulse-chase experiments. Heavy chain misfolding was measured as the half-life of endoglycosidase H (Endo H)-sensitive ␤ 2 -microglobulinfree heavy chains. Maturation/export rates were measured by acquisition of Endo H resistance. Association with calnexin or tapasin was analyzed by coprecipitation with chaperone-specific antibodies, and surface expression was estimated by flow cytometry. Thermostability of HLA-peptide complexes was assessed by immunoprecipitation after incubation at various temperatures. Heavy chain expression was quantified by Western blotting.Results. The folding rates of B*1402 and B*1403 were similar, and both were faster and more efficient than B*2705, but some unfolded heavy chains from both B14 subtypes remained in the endoplasmic reticulum (ER) with a long half-life. The export rates of B*1402 and B*1403 were slow, and the heterodimers partially dissociated after exiting the ER, as revealed by significant amounts of Endo H-resistant and surface-expressed free heavy chains. Both interaction with tapasin and thermostability were higher for B*2705 than for B*1402 and higher for B*1402 than for B*1403, suggesting that the repertoires of the B*1402-bound peptide and especially the B*1403-bound peptide were less optimized than that of B*2705.Conclusion. Our results indicate that the folding, maturation, and stability of B*1403 differ more from B*2705 than from B*1402. Thus, these features cannot account for the fact that only the 2 former allotypes are associated with AS.

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Unusual uniformity of the N-linked oligosaccharides of HLA-A, -B, and -C glycoproteins.

Cited by 43 publications

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A role for N-glycanase in the cytosolic turnover of glycoproteins

A role for N-glycanase in the cytosolic turnover of glycoproteins

Effect of glycosylation on structure and dynamics of MHC class I glycoprotein: A molecular dynamics study

Disparate folding and stability of the ankylosing spondylitis–associated HLA–B1403 and B2705 proteins

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Unusual uniformity of the N-linked oligosaccharides of HLA-A, -B, and -C glycoproteins.

Cited by 43 publications

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A role for N-glycanase in the cytosolic turnover of glycoproteins

A role for N-glycanase in the cytosolic turnover of glycoproteins

Effect of glycosylation on structure and dynamics of MHC class I glycoprotein: A molecular dynamics study

Disparate folding and stability of the ankylosing spondylitis–associated HLA–B*1403 and B*2705 proteins

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Disparate folding and stability of the ankylosing spondylitis–associated HLA–B1403 and B2705 proteins