Abstract
Besides protein identification via mass spectrometric methods, protein and peptide quantification has become more and more important in order to tackle biological questions. Methods like differential gel electrophoresis or enzyme-linked immunosorbent assays have been used to assess protein concentrations, while stable isotope labeling methods are also well established in quantitative proteomics. Recently, we developed metal-coded affinity tagging (MeCAT) as an alternative for accurate and sensitive quantification of peptides and proteins. In addition to absolute quantification via inductively coupled plasma mass spectrometry, MeCAT also enables sequence analysis via electrospray ionization tandem mass spectrometry. In the current study, we developed a new labeling approach utilizing an iodoacetamide MeCAT reagent (MeCAT-IA). The MeCAT-IA approach shows distinct advantages over the previously used MeCAT with maleinimide reactivity such as higher labeling efficiency and the lack of diastereomer formation during labeling. Here, we present a careful characterization of this new method focusing on the labeling process, which yields complete tagging with an excess of reagent of 1.6 to 1, less complex chromatographic behavior, and fragmentation characteristics of the tagged peptides using the iodoacetamide MeCAT reagent.
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We are very grateful to Proteome Factory AG and Thermo Fisher Scientific, especially Torsten Lindemann and Meike Hamester, for the technical support.
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Schwarz, G., Beck, S., Weller, M.G. et al. MeCAT—new iodoacetamide reagents for metal labeling of proteins and peptides. Anal Bioanal Chem 401, 1203–1209 (2011). https://doi.org/10.1007/s00216-011-5189-7
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DOI: https://doi.org/10.1007/s00216-011-5189-7