Carbamylation reduces the capacity of IgG for hexamerization and complement activation.
R LubbersS C OostindieD J DijkstraP W H I ParrenM K VerheulL AbendsteinT H SharpA de RuG M C JanssenP A van VeelenE T J van den BremerB BleijlevensB-J de KreukF J BeurskensLeendert Adrianus TrouwPublished in: Clinical and experimental immunology (2020)
Carbamylation is a post-translational modification that can be detected on a range of proteins, including immunoglobulin (Ig)G, in several clinical conditions. Carbamylated IgG (ca-IgG) was reported to lose its capacity to trigger complement activation, but the mechanism remains unclear. Because C1q binds with high affinity to hexameric IgG, we analyzed whether carbamylation of IgG affects binding of C1q, hexamerization and complement-dependent cytotoxicity (CDC). Synovial tissues of rheumatoid arthritis (RA) patients were analyzed for the presence of ca-IgG in vivo. Synovial tissues from RA patients were analyzed for the presence of ca-IgG using mass spectrometry (MS). Monomeric or hexameric antibodies were carbamylated in vitro and quality in solution was controlled. The capacity of ca-IgG to activate complement was analyzed in enzyme-linked immunosorbent (ELISAs) and cellular CDC assays. Using MS, we identified ca-IgG to be present in the joints of RA patients. Using in vitro carbamylated antibodies, we observed that ca-IgG lost its capacity to activate complement in both solid-phase and CDC assays. Mixing ca-IgG with non-modified IgG did not result in effective inhibition of complement activation by ca-IgG. Carbamylation of both monomeric IgG and preformed hexameric IgG greatly impaired the capacity to trigger complement activation. Furthermore, upon carbamylation, the preformed hexameric IgG dissociated into monomeric IgG in solution, indicating that carbamylation influences both hexamerization and C1q binding. In conclusion, ca-IgG can be detected in vivo and has a strongly reduced capacity to activate complement which is, in part, mediated through a reduced ability to form hexamers.
Keyphrases
- rheumatoid arthritis
- mass spectrometry
- end stage renal disease
- newly diagnosed
- gene expression
- multiple sclerosis
- cell cycle
- peritoneal dialysis
- systemic lupus erythematosus
- systemic sclerosis
- binding protein
- quality improvement
- interstitial lung disease
- transcription factor
- ankylosing spondylitis
- liquid chromatography