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Investigating quantitation of phosphorylation using MALDI‐TOF mass spectrometry
Author(s) -
Parker Laurie,
EngelHall Aaron,
Drew Kevin,
Steinhardt George,
Helseth Jr Donald L.,
Jabon David,
McMurry Timothy,
Angulo David S.,
Kron Stephen J.
Publication year - 2008
Publication title -
journal of mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.475
H-Index - 121
eISSN - 1096-9888
pISSN - 1076-5174
DOI - 10.1002/jms.1342
Subject(s) - chemistry , mass spectrometry , phosphopeptide , chromatography , peptide , isobaric labeling , calibration curve , analytical chemistry (journal) , matrix assisted laser desorption/ionization , protein mass spectrometry , desorption , tandem mass spectrometry , biochemistry , detection limit , organic chemistry , adsorption
Despite advances in methods and instrumentation for analysis of phosphopeptides using mass spectrometry, it is still difficult to quantify the extent of phosphorylation of a substrate because of physiochemical differences between unphosphorylated and phosphorylated peptides. Here we report experiments to investigate those differences using MALDI‐TOF mass spectrometry for a set of synthetic peptides by creating calibration curves of known input ratios of peptides/phosphopeptides and analyzing their resulting signal intensity ratios. These calibration curves reveal subtleties in sequence‐dependent differences for relative desorption/ionization efficiencies that cannot be seen from single‐point calibrations. We found that the behaviors were reproducible with a variability of 5–10% for observed phosphopeptide signal. Although these data allow us to begin addressing the issues related to modeling these properties and predicting relative signal strengths for other peptide sequences, it is clear that this behavior is highly complex and needs to be further explored. Copyright © 2007 John Wiley & Sons, Ltd.