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Molecules 2015, 20(4), 6592-6600; doi:10.3390/molecules20046592

Phenylglyoxal-Based Visualization of Citrullinated Proteins on Western Blots

1
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
2
Department of Synthetic Organic Chemistry, Institute for Molecules and Materials, Radboud University Nijmegen, P.O. Box 9101, NL-6500 HB Nijmegen, The Netherlands
*
Author to whom correspondence should be addressed.
Academic Editor: Arnaud Gautier
Received: 20 March 2015 / Revised: 1 April 2015 / Accepted: 2 April 2015 / Published: 14 April 2015
(This article belongs to the Special Issue Advances in Click Chemistry)
View Full-Text   |   Download PDF [631 KB, uploaded 14 April 2015]   |  

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. View Full-Text
Keywords: citrulline; peptidylarginine deiminases; anti-modified citrulline antibodies; phenylglyoxal; azide-alkyne cycloaddition; rheumatoid arthritis citrulline; peptidylarginine deiminases; anti-modified citrulline antibodies; phenylglyoxal; azide-alkyne cycloaddition; rheumatoid arthritis
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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MDPI and ACS Style

Hensen, S.M.M.; Boelens, W.C.; Bonger, K.M.; van Cruchten, R.T.P.; van Delft, F.L.; Pruijn, G.J.M. Phenylglyoxal-Based Visualization of Citrullinated Proteins on Western Blots. Molecules 2015, 20, 6592-6600.

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