Detection of dimethylarginines in protein hydrolysates by matrix-assisted laser desorption/ionization mass spectrometry

We report a method to detect the presence of dimethylarginines on proteins. Peptides with dimethylarginines were hydrolyzed in acid. The hydrolysates were subjected to matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometric analysis using a mixture of u-cyano-4-hydroxycinnamic acid and nitrocellulose as matrix. Both asymmetric omega-N-G, N-G-dimethylarginine and symmetric (omega-N-G-N-G-diniethylarginine give a clear signal at m/z 203. Recombinant Shp I p modified by Hint 1p in vivo were isolated by affinity chromatosiraphy followed by electrophoresis on a polyacrylamide gel and subjected to acid hydrolysis. MALDI-TOF analysis of the acid hydrolysates confirmed the presence of dimethylarginines. The detection limit of the method is estimated at approximately 1 pmol of protein. (c) 2005 Elsevier Inc. All rights reserved.