Gums & Jointsa FP7 EU collaborative project, with an aim to investigate protein citrullination as a link between periodontal disease and rheumatoid arthritis main page
Following synthesis, proteins undergo the process of post-translational modification (PTM), thus extending their range of function through the modification of amino acids with various functional groups (1,2). These modifications have a critical influence on protein structure and biological function, especially in the context of aging, physiological stress and inflammation. Those processes are either enzyme mediated (phosphorylation, glycosylation) or occurs spontaneously (carbamylation, isoaspartylation).
The role of PTM in the generation of neo-epitopes on 'self' proteins that are subsequently responsible for the pathogenesis of autoimmune diseases, such as multiple sclerosis, diabetes mellitus, systemic lupus erythematosus, and rheumatoid arthritis, has only recently been recognized (3-6).
Citrullination or deimination is the term used for the post-translational modification of the amino acid arginine in a protein into the amino acid citrulline. This reaction, shown below, is performed by enzymes called peptidylarginine deiminases (PADs). Up to date at least 5 human PAD enzymes had been identified. The conversion of arginine into citrulline can have important consequences for the structure and function of proteins, since arginine is positively charged at a neutral pH, whereas citrulline is uncharged. This increases the hydrophobicity of the protein, leading to changes in protein folding.In the reaction from arginine to citrulline, one of the terminal nitrogen atoms of the arginine sidechain is replaced by an oxygen. The reaction uses one water molecule and yields ammonia as a side-product:
P.gingivalis peptidylarginine deiminase (PPAD) as a potential virulence factor RA is fueled by disease-specific autoantibodies to citrullinated proteins, products of physiological post-translational modification of proteins by endogenous peptidylarginine deiminases (PADs). Factors which trigger the breakdown of tolerance to citrullinated proteins are unknown. It has been shown that PPAD can efficiently deiminate C-terminal Arg residues of bradykinin to inactivate the activity of this kinin (our unpublished results). Several other biologically active peptides (e.g. anaphylatoxins) have C- terminal arginine residues important for their activity. Therefore, citrullination of these peptides by PPAD may directly affect the inflammatory response. Further, citrullination of C-terminal arginines in peptides generated through the action of Arg-specific gingipain should facilitate P. gingivalis adherence to host connective tissues and to other dental plaque bacteria since these interactions are compromised by peptides bearing C-terminal arginine. Finally, generated by PPAD, citrullinated proteins and peptides can trigger autoimmune reaction to endogenously modified proteins and initiate RA as described above.
Carbamylation (homocitrullination) is a PTM that has been studied for many years in the context of uremia (7-9). It involves the non-enzymatic reaction of urea-derived cyanate with free NH2- groups on lysine (Lys) residues to yield homocitrulline (Hcit). This process can be mediated in vivo by myeloperoxidase (MPO), the enzyme responsible for the inflammation-driven carbamylation of proteins via the MPO/H2O2/SCN- system.Thus, MPO has recently attracted attention as a potential trigger factor for atherogenesis and inflammation (10).
Modification Altered Amino Acids
Phosphorylation Serine, tyrosine Glycosylation Aspargine, serine Hydroxylation Proline, lysine Acetylation Serine, lysine Methylation arginine, lysine
Citrulination Arginine Deamidation Asparagine Carbamylation Lysine N-terminal Cyclization N-terminal glutaminyl
Hcit residues affect the charge distribution within a peptide in a way that may result in impaired or even loss of function. Loss of enzymatic function upon carbamylation has been reported for matrix metalloproteinase 2, tissue inhibitor of metalloproteinase-2 and insulin (11,12). Plasma SCN- levels are known to be significantly higher in smokers, leading to increased carbamylation of proteins (10) and, furthermore, carbamylation has emerged as a potential pathogenic factor in renal insufficiency, cardiovascular disease and cataracts . Extracellular matrix proteins such as collagen and fibrinogen are thought to accumulate structural damage elicited by PTMs due to their long half-life and low turnover rates. Notably, it has been shown that collagen is easily carbamylated in vivo, and that such modification may be directly linked to granulocyte activation and protease release (13).
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