title: Structural Analysis of Variable Domain Glycosylation of Anti-Citrullinated Protein Antibodies in Rheumatoid Arthritis Reveals the Presence of Highly Sialylated Glycans.
abstract: Recently, we showed the unexpectedly high abundance of N-linked glycans on the Fab-domain of Anti-Citrullinated Protein Antibodies (ACPA). As N-linked glycans can mediate a variety of biological functions, we now aimed at investigating the structural composition of the Fab-glycans of ACPA-IgG to better understand their mediated biological effects. ACPA-IgG and noncitrulline specific (control) IgG from plasma and/or synovial fluid of nine ACPA positive rheumatoid arthritis patients were affinity purified. The N-linked glycosylation of total, Fc and F(ab')2fragments, as well as heavy and light chains of ACPA-IgG and control IgG were analyzed by UHPLC and MALDI-TOF mass spectrometry. The Fc-glycosylation of ACPA-IgG and IgG was analyzed at the glycopeptide level using LC-MS. The structural analyses revealed that ACPA-IgG molecules contain highly sialylated glycans in their Fab-domain. Importantly, Fab-glycans were estimated to be present on over 90% of ACPA-IgG, which is five times higher than in control IgG isolated from the same patients. This feature was more prominent on ACPA isolated from synovial fluid compared with peripheral blood. These observations provide the first evidence pointing to the ability of ACPA-IgG to mediate novel immunological activities, for example through binding specific lectins via hyper-sialylated Fab-glycans.
authors.affiliation: From the ‡Department of Rheumatology, Leiden University Medical Center, 2300 RC Leiden, the Netherlands; l.hafkenscheid@lumc.nl.
authors.forename: Lise
authors.initials: L
authors.lastname: Hafkenscheid
authors.affiliation: From the ‡Department of Rheumatology, Leiden University Medical Center, 2300 RC Leiden, the Netherlands.|§Center for Proteomics and Metabolomics, Leiden University Medical Center, 2300 RC Leiden, the Netherlands.
authors.forename: Albert
authors.initials: A
authors.lastname: Bondt
authors.affiliation: From the ‡Department of Rheumatology, Leiden University Medical Center, 2300 RC Leiden, the Netherlands.
authors.forename: Hans U
authors.initials: HU
authors.lastname: Scherer
authors.affiliation: From the ‡Department of Rheumatology, Leiden University Medical Center, 2300 RC Leiden, the Netherlands.
authors.forename: Tom W J
authors.initials: TW
authors.lastname: Huizinga
authors.affiliation: §Center for Proteomics and Metabolomics, Leiden University Medical Center, 2300 RC Leiden, the Netherlands.
authors.forename: Manfred
authors.initials: M
authors.lastname: Wuhrer
authors.affiliation: From the ‡Department of Rheumatology, Leiden University Medical Center, 2300 RC Leiden, the Netherlands.
authors.forename: René E M
authors.initials: RE
authors.lastname: Toes
authors.affiliation: From the ‡Department of Rheumatology, Leiden University Medical Center, 2300 RC Leiden, the Netherlands.|§Center for Proteomics and Metabolomics, Leiden University Medical Center, 2300 RC Leiden, the Netherlands.|¶Institut de Pharmacologie et de Biologie Structurale, Université de Toulouse, CNRS, UPS, France.
authors.forename: Yoann
authors.initials: Y
authors.lastname: Rombouts
mesh_terms: D001172:Arthritis, Rheumatoid / Q000276:immunology*; D001323:Autoantibodies / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry*; D007074:Immunoglobulin G / Q000032:analysis / Q000097:blood / Q000276:immunology; D011134:Polysaccharides / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013582:Synovial Fluid / Q000737:chemistry / Q000276:immunology
chemical_list: D001323:Autoantibodies; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D011134:Polysaccharides