Identification of consistent alkylation of cysteine-less peptides in a proteomics experiment

In a proteomics experiment, reduction and alkylation of proteins prior to enzymatic digestion ensures high sequence coverage of that protein during a database search. However, the alkylation procedure uses an excess of an alkylating agent such as iodoacetamide (IAA). Therefore, although other amino acids are alkylated, these modified peptides are not identified in a database search. Here we show that a large proportion of peptides are mono- and di-alkylated by IAA and therefore not identified via a database search. The first alkylation consistently takes place at the N-terminal amino acid. Therefore, we propose that during the database search conducted during a proteomics experiment, one should have the option of searching for any alkylated peptide at the N-terminal amino acid. (C) 2012 Elsevier Inc. All rights reserved.