Simultaneous Quantification and Identification Using 18O Labeling with an Ion Trap Mass Spectrometer and the Analysis Software Application ZoomQuant

摘要

Stable isotope labeling with ¹⁸O is a promising technique for obtaining both qualitative and quantitative information from a single differential protein expression experiment. The small 4 Da mass shift produced by incorporation of two molecules of ¹⁸O, and the lack of available methods for automated quantification of large data sets has limited the use of this approach with electrospray ionization-ion trap (ESI-IT) mass spectrometers. In this paper, we describe a method of acquiring ESI-IT mass spectrometric data that provides accurate calculation of relative ratios of peptides that have been differentially labeled using¹⁸O. The method utilizes zoom scans to provide high resolution data. This allows for accurate calculation of ¹⁸O/¹⁶O ratios for peptides even when as much as 50% of a ¹⁸O labeled peptide is present as the singly labeled species. The use of zoom scan data also provides sufficient resolution for calculating accurate ratios for peptides of +3 and lower charge states. Sequence coverage is comparable to that obtained with data acquisition modes that use only MS and MS/MS scans. We have employed a newly developed analysis software tool, ZoomQuant, which allows for the automated analysis of large data sets. We show that the combination of zoom scan data acquisition and analysis using ZoomQuant provides calculation of isotopic ratios accurate to ~21%. This compares well with data produced from ¹⁸O labeling experiments using time of flight (TOF) and Fourier transform-ion cyclotron resonance (FT-ICR) MS instruments.