title: Physical Characteristics of a Citrullinated Pro-Filaggrin Epitope Recognized by Anti-Citrullinated Protein Antibodies in Rheumatoid Arthritis Sera.
abstract: Rheumatoid arthritis (RA) is an autoimmune disease of complex etiology. A characteristic feature of a subset of RA is the presence of anti-citrullinated protein antibodies (ACPA), which correlate with a progressive disease course. In this study, we employed streptavidin capture enzyme-linked immunosorbent assay to analyze ACPA reactivity. Using the pro-filaggrin peptide HQCHQEST-Cit-GRSRGRCGRSGS, as template, we analyzed the reactivity of RA sera and healthy donor sera to various peptides in order to determine the physical characteristics of the citrullinated pro-filaggrin epitope and to examine whether biotin labelling influence antibody recognition. The full-length cyclic pro-filaggrin peptide and a linear form with a N-terminal biotin, was recognized to the same level, whereas, a notable difference in ACPA reactivity to the linear peptides with a C-terminal biotin was found, probably due to steric hindrance. Screening of linear and cyclic truncated peptides, revealed that small cyclic peptides containing 10-12 amino acids are favored over the linear. Moreover, the charged amino acids C-terminal to citrulline were found to be essential for antibody reactivity, most important was the charged amino acid in position 4 C-terminal to citrulline. Collectively, peptide structure, length, the presence of charged amino acids and biotin labelling markedly influence antibody reactivity. In relation to the clinical diagnostics of ACPA, these findings may reflect the differences in diagnostic assays used for detection of ACPA, which relates to differences in sensitivity and specificity dependent on the assay applied.
authors.affiliation: Department of Autoimmunology and Biomarkers, Statens Serum Institut, Artillerivej, Copenhagen S, Denmark.
authors.forename: Nicole Hartwig
authors.initials: NH
authors.lastname: Trier
authors.affiliation: Department of Autoimmunology and Biomarkers, Statens Serum Institut, Artillerivej, Copenhagen S, Denmark.
authors.forename: Bettina Eide
authors.initials: BE
authors.lastname: Holm
authors.affiliation: Department of Rheumatology, Glostrup Hospital, Nordre Ringvej, Glostrup, Denmark.
authors.forename: Ole
authors.initials: O
authors.lastname: Slot
authors.affiliation: Department of Rheumatology, Frederiksberg Hospital, Nordre Fasanvej, Frederiksberg, Denmark.
authors.forename: Henning
authors.initials: H
authors.lastname: Locht
authors.affiliation: Department of Rheumatology, Odense University Hospital, Søndre Boulevard, Odense C, Denmark.
authors.forename: Hanne
authors.initials: H
authors.lastname: Lindegaard
authors.affiliation: Department of Epidemiology, Biostatistics and Bio-demography, Institute of Public Health, University of Southern Denmark, Campusvej, Odense M, Denmark.
authors.forename: Anders
authors.initials: A
authors.lastname: Svendsen
authors.affiliation: Department of Autoimmunology and Biomarkers, Statens Serum Institut, Artillerivej, Copenhagen S, Denmark.
authors.forename: Gunnar
authors.initials: G
authors.lastname: Houen
mesh_terms: D000595:Amino Acid Sequence; D001172:Arthritis, Rheumatoid / Q000097:blood / Q000473:pathology*; D001323:Autoantibodies / Q000097:blood / Q000276:immunology*; D001710:Biotin / Q000737:chemistry / Q000378:metabolism; D016022:Case-Control Studies; D002956:Citrulline / Q000737:chemistry / Q000378:metabolism*; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology*; D000091344:Filaggrin Proteins; D006801:Humans; D007381:Intermediate Filament Proteins / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000276:immunology; D010456:Peptides, Cyclic / Q000737:chemistry / Q000276:immunology
chemical_list: D001323:Autoantibodies; D000939:Epitopes; C000717249:FLG protein, human; D000091344:Filaggrin Proteins; D007381:Intermediate Filament Proteins; D010455:Peptides; D010456:Peptides, Cyclic; D002956:Citrulline; D001710:Biotin