Modification of Cysteine Residues for Mass Spectrometry-Based Proteomic Analysis: Facts and Artifacts

Mass spectrometric proteomic analysis at the sample preparation stage involves the artificial reduction of disulfide bonds in proteins formed between cysteine residues. Such bonds, when preserved in their native state, complicate subsequent enzymatic hydrolysis and interpretation of the research results. To prevent the re-formation of the disulfide bonds, cysteine residues are protected by special groups, most often by alkylation. In this review, we consider the methods used to modify cysteine residues during sample preparation, as well as possible artifacts of this stage. Particularly, side reactions of the alkylating agents with other amino acid residues are described. The most common alkylating compound used to protect cysteine residues in mass spectrometric proteomic analysis is iodoacetamide. However, an analysis of the literature in this area indicates that this reagent causes more side reactions than other agents used, such as chloroacetamide and acrylamide. The latter can be recommended for wider use. In the review we also discuss the features of the cysteine residue modifications and their impact on the efficiency of the search for post-translational modifications and protein products of genes containing single nucleotide substitutions.