Abstract

In this study, we present a new isotope-coded carbamidomethylation (iCCM)-based quantitative proteomics, as acomplementary strategy for conventional isotope labeling strategies, with providing the simplicity, ease of use, and robustness. In iCCM-based quantification, two proteome samples can be separately isotope-labeled by means of covalently reaction of allcysteinyl residues in proteins with iodoacetamide (IAA) and its isotope (IAA-13C2, D2), denoted as CM and iCCM, respectively,leading to a mass shift of all cysteinyl residues to be + 4 Da. To evaluate iCCM-based isotope labeling in proteomic quantification,6 protein standards (i.e., bovine serum albumin, serotransferrin, lysozyme, beta-lactoglobulin, beta-galactosidase, andalpha-lactalbumin) isotopically labeled with IAA and its isotope, mixed equally, and followed by proteolytic digestion. Theresulting CM-/iCCM-labeled peptide mixtures were analyzed using a nLC-ESI-FT orbitrap-MS/MS. From our experimentalresults, we found that the efficiency of iCCM-based quantification is more superior to that of mTRAQ, as a conventional nonisobariclabeling method, in which both of a number of identified peptides from 6 protein standards and the less quantitative variationsin the relative abundance ratios of heavy-/light-labeled corresponding peptide pairs. Finally, we applied the developediCCM-based quantitative method to lung cancer serum proteome in order to evaluate the potential in biomarker discovery study