Peptidylarginine deiminase from Porphyromonas gingivalis citrullinates human fibrinogen and α‐enolase: Implications for autoimmunity in rheumatoid arthritis

Abstract

Objective. To investigate protein citrullination by the periodontal pathogen Porphyromonas gingivalis as a potential mechanism for breaking tolerance to citrullinated proteins in rheumatoid arthritis (RA).Methods. The expression of endogenous citrullinated proteins was analyzed by immunoblotting of cell extracts from P gingivalis and 10 other oral bacteria. P gingivalis-knockout strains lacking the bacterial peptidylarginine deiminases (PADs) or gingipains were created to assess the role of these enzymes in citrullination. Citrullination of human fibrinogen and ␣-enolase by P gingivalis was studied by incubating live wild-type and knockout strains with the proteins and analyzing the products by immunoblotting and mass spectrometry.Results. Endogenous protein citrullination was abundant in P gingivalis but lacking in the other oral bacteria. Deletion of the bacterial PAD gene resulted in complete abrogation of protein citrullination. Inactivation of arginine gingipains, but not lysine gingipains, led to decreased citrullination. Incubation of wild-type P gingivalis with fibrinogen or ␣-enolase caused degradation of the proteins and citrullination of the resulting peptides at carboxy-terminal arginine residues, which were identified by mass spectrometry.Conclusion. Our findings demonstrate that among the oral bacterial pathogens tested, P gingivalis is unique in its ability to citrullinate proteins. We further show that P gingivalis rapidly generates citrullinated host peptides by proteolytic cleavage at Arg-X peptide bonds by arginine gingipains, followed by citrullination of carboxy-terminal arginines by bacterial PAD. Our results suggest a novel model where P gingivalismediated citrullination of bacterial and host proteins provides a molecular mechanism for generating antigens that drive the autoimmune response in RA.Rheumatoid arthritis (RA) is characterized by disease-specific autoimmunity to citrullinated proteins. Citrullination is a posttranslational modification of arginine residues that is mediated by the family of peptidylarginine deiminases (PADs). Citrullinated fibrin(ogen) and ␣-enolase are 2 of the physiologic proteins that are targeted by anti-citrullinated protein antibodies in RA (1-5). Fibrinogen is the precursor of fibrin, and autoanMs. Wegner and Drs.

Citrullination as a plausible link to periodontitis, rheumatoid arthritis, atherosclerosis and Alzheimer’s disease

“…The modification of C-terminal arginine residues is the result of PPAD becoming structurally closer to Rgps on the bacterial surface. This dual enzymatic action of modification was initially reported following production of citrullinated peptides derived from fibrinogen and a-enolase by PPAD [30]. These two proteins are major auto-antigens in RA [3].…”

Animal models of rheumatoid arthritis: How informative are they?

“…The presence of anti-citrullinated protein antibodies (ACPA) is also highly specific to RA (over 90%) and linked with a more progressive, destructive arthritis (Klareskog et al, 2008;Silveira et al, 2007;Snir et al, 2010). ACPA have been described up to 10 years before disease onset and are additionally correlated with RA risk factors such as smoking and periodontal disease (Hensvold et al, 2015;Mahdi et al, 2009;Wegner et al, 2010). RF is generally thought to be absent from rodent models, but aggressive erosive disease is linked to the presence of anticitrullinated cyclic peptide antibodies (anti-CCP) which may be generated within RA pannus ELS.…”

The role of the gastrointestinal tract in the pathogenesis of rheumatic diseases

“…This is suggested by the evidence that the oral bacteria Porphyromonas gingivalis (PG) is capable of inducing the local production of citrullinated protein [8]. PG is a leading pathogen of chronic periodontitis [9] and produces a unique bacterial enzyme, PG peptidyl-arginine deiminase (PPAD), which has the ability to convert arginine residues in proteins to citrulline [8].…”