Open AccessIsobaric Quantitative Protein Interaction Reporter Technology for Comparative Interactome Studies
Author(s) -
Juan D. Chavez,
Andrew Keller,
Jared P. Mohr,
James E. Bruce
Publication year - 2020
Publication title -
analytical chemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.117
H-Index - 332
eISSN - 1520-6882
pISSN - 0003-2700
DOI - 10.1021/acs.analchem.0c03128
Subject(s) - chemistry , stable isotope labeling by amino acids in cell culture , quantitative proteomics , tandem mass spectrometry , linker , isobaric process , proteome , isotope , tandem , mass spectrometry , isobaric labeling , tandem mass tag , peptide , fragmentation (computing) , proteomics , computational biology , biophysics , protein mass spectrometry , chromatography , biochemistry , physics , thermodynamics , materials science , quantum mechanics , composite material , biology , computer science , gene , operating system
Chemical cross-linking with mass spectrometry (XL-MS) has emerged as a useful tool for the large-scale study of protein structures and interactions from complex biological samples including intact cells and tissues. Quantitative XL-MS (qXL-MS) provides unique information on protein conformational and interaction changes resulting from perturbations such as drug treatment and disease state. Previous qXL-MS studies relied on the incorporation of stable isotopes into the cross-linker (primarily deuterium) or metabolic labeling with SILAC. Here, we introduce isobaric quantitative protein interaction reporter (iqPIR) technology which utilizes stable isotopes selectively incorporated into the cross-linker design, allowing for isobaric cross-linked peptide pairs originating from different samples to display distinct quantitative isotope signatures in tandem mass spectra. This enables improved quantitation of cross-linked peptide levels from proteome-wide samples because of the reduced complexity of tandem mass spectra relative to MS 1 spectra. In addition, because of the isotope incorporation in the reporter and the residual components of the cross-linker that remain on released peptides, each fragmentation spectrum can offer multiple independent opportunities and, therefore, improved confidence for quantitative assessment of the cross-linker pair level. Finally, in addition to providing information on solvent accessibility of lysine sites, dead end iqPIR cross-linked products can provide protein abundance and/or lysine site modification level information all from a single in vivo cross-linking experiment.