Abstract
Differential precipitation of proteins (DiffPOP) is a simple technique for fractionating complex protein mixtures. Using stepwise addition of acidified methanol, ten distinct subsets of proteins can be selectively precipitated by centrifugation and identified by mass spectrometry-based proteomics. We have previously shown that the ability of a protein to resist precipitation can be altered by drug binding, which enabled us to identify a novel drug-target interaction. Here, we show that the addition of DiffPOP to a standard LC-MS proteomics workflow results in a three-dimensional separation of peptides that increases protein coverage and peptide identifications. Importantly, DiffPOP reveals solubility differences between proteoforms, potentially providing valuable insights that are typically lost in bottom-up proteomics.
Original language | English (US) |
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Pages (from-to) | 2025-2033 |
Number of pages | 9 |
Journal | Journal of the American Society for Mass Spectrometry |
Volume | 34 |
Issue number | 9 |
DOIs | |
State | Published - Sep 6 2023 |
Keywords
- DiffPOP
- MudPIT
- protein separation
- proteoform
- sequence coverage
ASJC Scopus subject areas
- Structural Biology
- Spectroscopy