TY - JOUR
T1 - Isotope-Coded Maleimide Affinity Tags for Proteomics Applications
AU - Wdowiak, Adam Piotr
AU - Duong, Marisa
AU - Joyce, Rohan
AU - Boyatzis, Amber
AU - Walkey, Mark
AU - Nealon, Gareth
AU - Arthur, Peter
AU - Piggott, Matthew
PY - 2021/8/18
Y1 - 2021/8/18
N2 - 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.
AB - 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.
U2 - 10.1021/acs.bioconjchem.1c00206
DO - 10.1021/acs.bioconjchem.1c00206
M3 - Article
C2 - 34160215
VL - 32
SP - 1652
EP - 1666
JO - Bioconjugate Chemistry
JF - Bioconjugate Chemistry
SN - 1043-1802
IS - 8
ER -