Attachment of oligosaccharides to peptide antigen profoundly affects binding to major histocompatibility complex class II molecules and peptide immunogenicity.
To investigate the immunogenicity of glycopeptides, a peptide fragment from hen egg lysozyme, HEL(81-96)-Y (here named 1) which is immunogenic in H-2k mice and known to bind to the murine major histocompatibility complex (MHC) class II molecule Ek, was synthesized in five different glycosylated forms. The N-terminal serine of HEL(81-96)-Y was derivatized with D-glucose (2), maltotriose (3), and a branched D-glucose pentasaccharide (4). Furthermore, 1 was prepared with a central serine or asparagine derivatized with the branched D-glucose pentasaccharide (5) and GlcNAc (6), respectively. The ability of the five glycopeptides and the non-glycosylated peptide, labeled with 125I, to bind to the two MHC class II molecules, Ak and Ek, was studied using a gel filtration assay. None of them could bind to Ak. Neither 5 nor 6 were able to bind to Ek. Surprisingly 2, 3 and 4 bound better to Ek than did the non-glycosylated peptide 1. The increased binding varied depending on the type of oligosaccharide attached to the N terminus of the peptide. The better binding to Ek of glycopeptide 4 was found to be due to an increased association rate. The binding of 1 as well as 4 was optimal at pH 5.0. Functional studies showed that 4 was able to elicit a heteroclitic proliferative response from T cells of mice immunized with the native non-glycosylated peptide. Circular dichroism studies of 1 and 4 indicated a more unordered structure of 4 and a predominant alpha-helical conformation of 1, suggesting that the MHC class II molecule may bind to peptides which are in a non-alpha-helical conformation. These results demonstrate that glycosylation has considerable influence on peptide immunogenicity for T lymphocytes.
Mouritsen
S
S
M & E ApS, Copenhagen, Denmark.
Meldal
M
M
Christiansen-Brams
I
I
Elsner
H
H
Werdelin
O
O
D000595:Amino Acid Sequence; D000818:Animals; D000941:Antigens / Q000737:chemistry*; D001667:Binding, Competitive / Q000502:physiology; D002240:Carbohydrate Sequence; D002942:Circular Dichroism; D006020:Glycopeptides / Q000276:immunology; D000949:Histocompatibility Antigens Class II / Q000378:metabolism*; D008213:Lymphocyte Activation; D051379:Mice; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000276:immunology*; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry / Q000276:immunology*; D011485:Protein Binding
D000941:Antigens; D006020:Glycopeptides; D000949:Histocompatibility Antigens Class II; D009844:Oligosaccharides; D010446:Peptide Fragments