Lectin purified human class I MHC-derived peptides: evidence for presentation of glycopeptides in vivo.
Previously, using synthetic glycopeptides carrying a natural cytosolic type of monosaccharide O-beta-linked N-acetylglucosamine (GlcNAc) glycosylation of serine residues, we have shown that glycopeptides act as suitable substrates for TAP-mediated transport into the endoplasmic reticulum (ER), and that they bind efficiently to class I major histocompatibility complex (MHC) molecules and can elicit glycopeptide-specific cytotoxic T-lymphocyte (CTL) responses in mice. Recently, we have reported that peptides presented by human class I MHC molecules in vivo encompass a small but significant amount of peptides which seem to be carrying O-beta-linked monosaccharide GlcNAc. In the present report we provide further evidence that glycosylated peptides are indeed presented by class I MHC molecules in vivo. Thus, peptides derived from HLA-A*0201 were purified by wheat germ agglutinin (WGA) lectin affinity chromatography as previously described. Subsequently, the peptides contained in the WGA-eluate were subjected to sequence analysis by Edman degradation. It was found that the peptides derived from HLA-A*0201 which had been retained by the O-GlcNAc-binding lectin WGA did indeed carry a HLA-A*0201 binding motif. Furthermore, using an enzymatic labeling procedure we present evidence that the HLA-A*0201-derived peptides which bind to the WGA lectin are glycosylated with terminal GlcNAc residues. Together, these data provide further evidence for the natural presentation by human class I MHC of glycopeptides carrying terminal O-GlcNAc residues in vivo.
Kastrup
I B
IB
Institute of Cancer Biology, Danish Cancer Society, Copenhagen.
Stevanovic
S
S
Arsequell
G
G
Valencia
G
G
Zeuthen
J
J
Rammensee
H G
HG
Elliott
T
T
Haurum
J S
JS
D000117:Acetylglucosamine / Q000276:immunology; D017951:Antigen Presentation / Q000276:immunology*; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D005700:Galactosyltransferases; D006020:Glycopeptides / Q000276:immunology* / Q000378:metabolism; D006031:Glycosylation; D015395:Histocompatibility Antigens Class I / Q000276:immunology* / Q000302:isolation & purification*; D006801:Humans; D011485:Protein Binding / Q000276:immunology; D011499:Protein Processing, Post-Translational / Q000276:immunology; D020539:Sequence Analysis, Protein; D014909:Wheat Germ Agglutinins
D006020:Glycopeptides; D015395:Histocompatibility Antigens Class I; D014909:Wheat Germ Agglutinins; D005700:Galactosyltransferases; D000117:Acetylglucosamine