title: Synthesis of a C-glycoside analogue of beta-D-galactosyl hydroxynorvaline and its use in immunological studies.
abstract: A C-linked isostere of beta-D-galactosylated hydroxynorvaline has been prepared in eight steps from per-O-benzylated galactopyranolactone. Addition of a homoallylic Grignard reagent to the lactone, reduction of the resulting hemiacetal with triethylsilane, and a Wittig reaction with Garner's aldehyde were key steps in this synthesis. The C-linked building block was then incorporated at position 264 into the fragment CII(256--270) from typeII collagen by solid-phase synthesis using a combination of the tert-butoxycarbonyl (Boc) and 9-fluorenylmethoxycarbonyl (Fmoc) protective group strategies. Deprotection of the benzylated C-linked galactosyl moiety was achieved simultaneously with cleavage of the glycopeptide from the solid phase by using triethylsilyl trifluoromethanesulfonate in TFA. Helper T-cell hybridomas obtained in a mouse model for rheumatoid arthritis responded to the C-linked glycopeptide when presented by classII MHC molecules. However, 10- to 20-fold higher concentrations were required as compared to when O-linked beta-D-galactosylated hydroxynorvaline or hydroxylysine (Hyl) were present at position 264 of CII(256--270). Thus, replacement of a single oxygen atom by a methylene group in the carbohydrate moiety of a glycopeptide antigen had a substantial influence on the T-cell response. This reveals that T cells are able to recognize the carbohydrate moiety of glycopeptide antigens with high specificity. Finally, the results suggest that structural modifications of beta-D-Gal-Hyl(264) in CII(256--270) may give altered peptide ligands that can be used for induction of tolerance in autoimmune rheumatoid arthritis.
authors.affiliation: Organic Chemistry Department of Chemistry Umeå University, Umeå, Sweden.
authors.forename: E
authors.initials: E
authors.lastname: Wellner
authors.forename: T
authors.initials: T
authors.lastname: Gustafsson
authors.forename: J
authors.initials: J
authors.lastname: Bäcklund
authors.forename: R
authors.initials: R
authors.lastname: Holmdahl
authors.forename: J
authors.initials: J
authors.lastname: Kihlberg
mesh_terms: D000818:Animals; D001169:Arthritis, Experimental / Q000276:immunology; D001172:Arthritis, Rheumatoid / Q000276:immunology; D002236:Carbohydrate Conformation; D024043:Collagen Type II / Q000737:chemistry / Q000276:immunology*; D005690:Galactose / Q000031:analogs & derivatives / Q000737:chemistry; D006020:Glycopeptides / Q000138:chemical synthesis* / Q000378:metabolism; D006031:Glycosylation; D006680:HLA Antigens / Q000276:immunology; D006825:Hybridomas / Q000276:immunology; D016056:Immunodominant Epitopes / Q000737:chemistry / Q000378:metabolism; D007376:Interleukin-2 / Q000378:metabolism; D008024:Ligands; D051379:Mice; D008809:Mice, Inbred C3H; D008811:Mice, Inbred DBA; D015394:Molecular Structure; D051381:Rats; D013601:T-Lymphocytes / Q000276:immunology*; D014633:Valine / Q000031:analogs & derivatives / Q000737:chemistry
chemical_list: D024043:Collagen Type II; D006020:Glycopeptides; D006680:HLA Antigens; D016056:Immunodominant Epitopes; D007376:Interleukin-2; D008024:Ligands; D014633:Valine; D005690:Galactose