title: Phenylglyoxal-based visualization of citrullinated proteins on Western blots.
abstract: Citrullination is the conversion of peptidylarginine to peptidylcitrulline, which is catalyzed by peptidylarginine deiminases. This conversion is involved in different physiological processes and is associated with several diseases, including cancer and rheumatoid arthritis. A common method to detect citrullinated proteins relies on anti-modified citrulline antibodies directed to a specific chemical modification of the citrulline side chain. Here, we describe a versatile, antibody-independent method for the detection of citrullinated proteins on a membrane, based on the selective reaction of phenylglyoxal with the ureido group of citrulline under highly acidic conditions. The method makes use of 4-azidophenylglyoxal, which, after reaction with citrullinated proteins, can be visualized with alkyne-conjugated probes. The sensitivity of this procedure, using an alkyne-biotin probe, appeared to be comparable to the antibody-based detection method and independent of the sequence surrounding the citrulline.
authors: Department of Biomolecular Chemistry, Institute for Molecules and Materials, Radboud Institute for Molecular Life Sciences and Netherlands Proteomics Centre, Radboud University Nijmegen, P.O. Box 9101, NL-6500 HB Nijmegen, The Netherlands. sannehensen@gmail.com.
authors: Sanne M M
authors: SM
authors: Hensen
authors: Department of Biomolecular Chemistry, Institute for Molecules and Materials, Radboud Institute for Molecular Life Sciences and Netherlands Proteomics Centre, Radboud University Nijmegen, P.O. Box 9101, NL-6500 HB Nijmegen, The Netherlands. w.boelens@ncmls.ru.nl.
authors: Wilbert C
authors: WC
authors: Boelens
authors: Department of Biomolecular Chemistry, Institute for Molecules and Materials, Radboud Institute for Molecular Life Sciences and Netherlands Proteomics Centre, Radboud University Nijmegen, P.O. Box 9101, NL-6500 HB Nijmegen, The Netherlands. k.bonger@science.ru.nl.
authors: Kimberly M
authors: KM
authors: Bonger
authors: Department of Biomolecular Chemistry, Institute for Molecules and Materials, Radboud Institute for Molecular Life Sciences and Netherlands Proteomics Centre, Radboud University Nijmegen, P.O. Box 9101, NL-6500 HB Nijmegen, The Netherlands. remcovancruchten@gmail.com.
authors: Remco T P
authors: RT
authors: van Cruchten
authors: Department of Synthetic Organic Chemistry, Institute for Molecules and Materials, Radboud University Nijmegen, P.O. Box 9101, NL-6500 HB Nijmegen, The Netherlands. f.vandelft@synaffix.com.
authors: Floris L
authors: FL
authors: van Delft
authors: Department of Biomolecular Chemistry, Institute for Molecules and Materials, Radboud Institute for Molecular Life Sciences and Netherlands Proteomics Centre, Radboud University Nijmegen, P.O. Box 9101, NL-6500 HB Nijmegen, The Netherlands. G.Pruijn@ncmls.ru.nl.
authors: Ger J M
authors: GJ
authors: Pruijn
mesh_terms: D000818:Animals; D015153:Blotting, Western / Q000379:methods; D002384:Catalysis; D002956:Citrulline / Q000737:chemistry*; D006801:Humans; D006867:Hydrolases / Q000378:metabolism; D007202:Indicators and Reagents / Q000737:chemistry; D010658:Phenylglyoxal / Q000737:chemistry*; D000076342:Protein-Arginine Deiminases; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D013194:Staining and Labeling
chemical_list: D007202:Indicators and Reagents; D011506:Proteins; D002956:Citrulline; D006867:Hydrolases; D000076342:Protein-Arginine Deiminases; D010658:Phenylglyoxal