An approach to quantitative proteome analysis by labeling tryptophan residues

This report describes a method for quantification and sequence identification of individual proteins in complex mixtures. The method is based on labeling with the chemical reagent 2-nitrobenzenesulfenyl chloride (NBSCl) in conjunction with tandem mass spectrometry. In this method, selective introduction of the 2-nitrobenzenesulfenyl (NBS) moiety onto tryptophan residues is achieved, and a 6 Da mass differential is generated using C-13(6)-labeled NBSCl (NBSCl-Cd-13 and C-12(6)-labeled NBSCl (NBSCl-C-12(6)). The 6 Da mass differential between the NBS-C-12(6)-labeled and the NBS-C-13(6)-labeled peptides assigns a mass signature to all tryptophan-containing peptides in any pool of proteolytic digests for protein identification through peptide mass mapping. Using this strategy, we compared the protein expression in rat sera using a normal (control) rat (Crj:-Wistar) and a hyperglycemic rat (GK/Crj). The stable isotope dilution techniques used in this method provide highly accurate relative quantification. The NBS approach offers a widely applicable means of analyzing protein mixtures derived from biological samples, and the method described here presents an effective and simplified approach to proteome analysis. Copyright (C) 2003 John Wiley Sons, Ltd.