Site-selective, reversible, pH-induced N-terminal maleylation and its application for proteomics research.

Abstract

Compared with traditional labelling reagents used in proteomics, maleic anhydride is milder and can be easily removed under certain conditions, thus simplifying chemical derivatization. The proposed strategy combined a site-specific chemical labelling reaction with mass spectrometry. Site-selective, reversible N-terminal maleylation was controlled by pH. Selective maleyl N-terminal labelling was achieved with high efficiency under the optimized reaction conditions. The demaleylation conditions were also optimized. The sequence coverage of histone H4 increased from 77% to 95% after the maleyl labels were removed, and the number of maleylated peptides was five times that of the unlabelled peptides. We further verified the reversible and selective N-terminal labelling properties of maleic anhydride through propionylation labelling at the peptide/protein level. A new method for site-selective maleylation of the N-terminal amino groups of a peptide was explored. Through the optimization experiment, good efficiency was achieved for this labelling reaction. The reversibility of maleylation labelling was also explored and applied for the identification of post-translational modifications of histones. Thus, site-selective, reversible, pH-induced N-terminal labelling using maleic anhydride has greater potential for application in proteomics than any other labelling methods.