1. Isotope-Coded Maleimide Affinity Tags for Proteomics Applications
- Author
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Matthew J. Piggott, Mark C. Walkey, Marisa N. Duong, Peter G. Arthur, Adam P. Wdowiak, Rohan D. Joyce, Gareth L. Nealon, and Amber E. Boyatzis
- Subjects
Male ,Models, Molecular ,Proteomics ,Protein Conformation ,Quantitative proteomics ,Biomedical Engineering ,Pharmaceutical Science ,Muscle Proteins ,Bioengineering ,02 engineering and technology ,Mass spectrometry ,01 natural sciences ,Adduct ,Maleimides ,chemistry.chemical_compound ,Mice ,Dogs ,Tandem Mass Spectrometry ,Animals ,Muscle, Skeletal ,Maleimide ,Pharmacology ,Carbon Isotopes ,010405 organic chemistry ,Organic Chemistry ,021001 nanoscience & nanotechnology ,Combinatorial chemistry ,0104 chemical sciences ,chemistry ,Gene Expression Regulation ,Isotope Labeling ,Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ,Electrophile ,Proteome ,Mice, Inbred mdx ,0210 nano-technology ,Linker ,Biotechnology ,Chromatography, Liquid - Abstract
Isotope-coded affinity tags (ICATs) are valuable tools for mass spectrometry-based quantitative proteomics, in particular, for comparison of protein (cysteine-residue) thiol oxidation state in normal, stressed, and diseased tissue. However, the iodoacetamido electrophile used in most commercial ICATs suffers from poor thiol-selectivity and modest rates of adduct formation, which can lead to spurious results. Hence, we designed and synthesized three ICATs containing thiol-selective N-alkylmaleimide electrophiles (isotope-coded maleimide affinity tags = ICMATs) and assessed these as mass spectrometry probes for ratiometric analysis of lysozyme and muscle proteomes. Two ICMAT pairs containing butylene/D8-butylene linkers were effective MS probes, but not ideal for typical proteomics workflows, because peptides bearing these tags frequently did not coelute with HPLC. A switch to a phenylene/13C6-phenylene linker solved this issue without compromising the efficiency of adduct formation.
- Published
- 2021