A novel pyrimidine-based stable-isotope labeling reagent and its application to quantitative analysis using...

As an extension of our previous work, a novel pyrimidine-based stable-isotope labeling reagent, [d(0)]-/[d(6)]-4,6-dimethoxy-2-(methylsulfonyl)pyrimidine (DMMSP), was developed for comparative quantification of proteins by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Our one-step labeling strategy combines several desirable properties such as cysteine-specific labeling, signal amplification and direct analysis with minimum sample handling. All these features not only allow easy interpretation for protein identification and quantification but also ensure rapid and sensitive progression to MS analysis. Using cysteine, Cys-containing peptide, and lysozyme digest as model samples, the labeling methodology was established and the following pilot application for quantitative analysis was accomplished with high confidence, accuracy, efficiency, and reproducibility. The application of DMMSP-labeling strategy is expected to provide a powerful new tool for comparative proteome research, especially for the analysis of low-abundance proteins.