Epitope mapping of the N-terminal portion of tissue transglutaminase protein antigen to identify linear epitopes in celiac disease.
Celiac disease (CD) is an autoimmune mediated disease with complex and multifactorial etiology. Gluten intake triggers a composite immune response involving T-cells and B-cells and leading to the secretion of autoantibodies if a genetic predisposition is present. Untreated CD patients show high levels of circulating autoantibodies directed to different auto-antigens present in the intestinal mucosa. The most important auto-antigen is the endomysial enzyme tissue transglutaminase (tTG). Both IgA and IgG antibody isotypes to tTG are known, but only the IgA antibodies demonstrate the highest disease specificity and thus are considered disease biomarkers. Because the pathogenicity and exact tTG binding properties of these autoantibodies are still unclear, the characterization of tTG antigenic domains is a crucial step in understanding CD onset and the autoimmune pathogenesis. Overlapping peptide libraries can be used for epitope mapping of selected protein portions to determine antigenic fragments contributing to the immunological activity and possibly develop innovative peptide-based tools with high specificity and sensitivity for CD. We performed an epitope mapping study to characterize putative linear auto-antigenic epitopes present in the tTG N-terminal portion (1-230). A library of 23 overlapping peptides spanning tTG(1-230) was generated by Fmoc/tBu solid-phase peptide synthesis and screened by immunoenzymatic assays employing patients' sera. The results indicate that four synthetic peptides, that is, Ac-tTG(1-15)-NH2 , Ac-tTG(41-55)-NH2 , Ac-tTG(51-65)-NH2 , and Ac-tTG(151-165)-NH2 , are recognized by IgA autoantibodies circulating in CD patients' sera. These results offer important insight on the nature of the antigen-antibody interaction.
Laboratory of Peptide and Protein Chemistry and Biology, University of Florence, I-50019, Sesto Fiorentino, Italy; Department of Chemistry 'Ugo Schiff', University of Florence, I-50019, Sesto Fiorentino, Italy.
Margherita
M
Di Pisa
Patrick
P
Buccato
Giuseppina
G
Sabatino
Feliciana
F
Real Fernández
Brunilde
B
Berti
Francesco
F
Cocola
Anna Maria
AM
Papini
Paolo
P
Rovero
D001324:Autoantigens / Q000737:chemistry / Q000276:immunology*; D001327:Autoimmune Diseases / Q000276:immunology*; D002446:Celiac Disease / Q000276:immunology*; D018604:Epitope Mapping; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D005260:Female; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D007070:Immunoglobulin A / Q000737:chemistry / Q000276:immunology*; D008297:Male; D019151:Peptide Library; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases / Q000737:chemistry / Q000276:immunology*
D001324:Autoantigens; D000939:Epitopes; D007070:Immunoglobulin A; D019151:Peptide Library; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D019204:GTP-Binding Proteins
antigenic peptides; celiac disease; diagnostic peptides; epitope mapping; tissue transglutaminase