Deconvolution of overlapping isotopic clusters improves quantification of stable isotope-labeled peptides.

ABSTRACT

High-resolution mass spectrometry and the use of stable isotopes have greatly improved our ability to quantify proteomes. Typically, the relative abundance of peptides is estimated by identifying the isotopic clusters and by comparing the peak intensities of peptide pairs. However, when the mass shift between the labeled peptides is small, there can be the possibility for overlap of the isotopic clusters which will hamper quantification accuracy with a typical upwards bias for the heavier peptide. Here, we investigated the impact of the overlapping peak issue with respect to dimethyl based quantification and we confirmed there can be need for correction. In addition, we present a tool that can correct overlapping issues when they arise which is based on modeling isotopic distributions. We demonstrate that our approach leads to improved accuracy and precision of protein quantification.