The study of protein kinetics requires an accurate measurement of isotopic ratios of peptides. Although Fourier transform-ion cyclotron resonance (FT-ICR) mass spectrometers yield accurate mass measurements of analytes, the isotopologue ratios are consistently lower than predicted. Recently, we demonstrated that the magnitude of the spectral error in the FT-ICR mass spectrometer is proportional to the scan duration of ions. Here, we present a novel isotopic ratio extrapolation (IRE) method for obtaining accurate isotopic ratio measurements. Accuracy is achieved by performing scans with different duration and extrapolation of the data to the initial moment of the ion rotation; IRE minimizes the absolute isotopic ratio error to .1%. We demonstrate the application of IRE in protein turnover studies using (2)H(2)O-metabolic labeling. Overall, this technique allows accurate measurements of the isotopic ratios of proteolytic peptides, a critical step for enabling routine studies of proteome dynamics.
Figure Measured and extrapolated isotopic ratios (natural abundance) of the peptide AWAVAR. Data were collected at 3 different acquisition times with multiple injections (n=4). The Mi/M0 intercepts represent the extrapolated isotopic ratios. Error bars represent standard deviation from the mean value.
Results from: Ilchenko S, Previs SF, Rachdaoui N, Willard B, McCullough AJ, Kasumov T. J Am Soc Mass Spectrom. 2013 Feb;24(2):309-12. doi: 10.1007/s13361-012-0536-2. Epub 2013 Jan 3.