Urea Artifacts Interfere with Immuno-Purification of Lysine Acetylation

Comprehensive
analysis of post-translational modifications (PTMs)
often depends on the purification of modified peptides prior to LC–MS/MS.
The implementation of these enrichment methods requires thorough knowledge
of the experimental conditions to achieve optimal selectivity and
sensitivity. In this regard, large-scale analysis of lysine acetylation,
a key PTM for multiple cellular processes, makes use of monoclonal
pan-antibodies designed against this moiety. We report that the immuno-purification
of lysine-acetylated peptides is hampered by the copurification of
lysine carbamylated peptides, a frequent urea artifact. This specific
interaction can be explained by the similar chemical structures of
lysine acetylation and lysine carbamylation. As an alternative, we
propose a sample preparation protocol based on sodium deoxycholate
that eliminates these artifacts and dramatically improves the selectivity
and sensitivity of this immuno-purification assay.