biostudies-literatureBereta GPNational Science Centre PolandEEA Grants/Norway GrantsEEA Grants/Norway Grants (EEA and Norway Grants)Polish Ministry of Higher EducationResearch Council of Norway and NIHNIGMS NIH HHSe2508369122https://www.ebi.ac.uk/biostudies/studies/S-EPMC12595441biostudies-literatureUnknown122(44)Rheumatoid arthritis is a chronic inflammatory disease driven by abnormal protein modifications. These include citrullination of arginine residues by the calcium-activated enzyme peptidylarginine deiminase 4 (PAD4). However, calcium in body fluids may not fully activate PAD4, suggesting the potential involvement of other activators. In this study, we investigated the ability of glycosaminoglycans (a class of negatively charged polysaccharides) to modulate PAD4 activity. We found that model glycosaminoglycans bind to the enzyme with a nanomolar affinity, increase its calcium sensitivity, and require enzyme dimerization for activation. These effects depend on the size and negative charge of the glycosaminoglycan, and its various natural forms activate PAD4. Thus, our findings elucidate a mecProceedings of the National Academy of Sciences of the United States of AmericaGlycosaminoglycans activate peptidylarginine deiminase 4 by enhancing calcium affinity.PMC12595441R01 GM1293252024/53/B/NZ5/010052961291/SOL/2021/22018/30/M/NZ1/003672019/34/H/NZ1/00674Marzec KNowak JPijanowski LMydel PKaminska MPoreba MBereta GPWilk PBiela APBielecka EKowalczyk DKoziel JGrudnik PWator-Wilk EKantyka TfalseGlycosaminoglycans activate peptidylarginine deiminase 4 by enhancing calcium affinity.Rheumatoid arthritis is a chronic inflammatory disease driven by abnormal protein modifications. These include citrullination of arginine residues by the calcium-activated enzyme peptidylarginine deiminase 4 (PAD4). However, calcium in body fluids may not fully activate PAD4, suggesting the potential involvement of other activators. In this study, we investigated the ability of glycosaminoglycans (a class of negatively charged polysaccharides) to modulate PAD4 activity. We found that model glycosaminoglycans bind to the enzyme with a nanomolar affinity, increase its calcium sensitivity, and require enzyme dimerization for activation. These effects depend on the size and negative charge of the glycosaminoglycan, and its various natural forms activate PAD4. Thus, our findings elucidate a mec2025-01-01T00:00:00Z2025 Nov2026-06-05T14:43:45.427Z2026-05-18T03:08:09.13ZS-EPMC125954414116641710.1073/pnas.2508369122