An advanced strategy for comprehensive profiling of ADP-ribosylation sites using mass spectrometry-based proteomics.

An advanced strategy for comprehensive profiling of ADP-ribosylation sites using mass spectrometry-based proteomics. Mol Cell Proteomics. 2019 Feb 23;: Authors: Hendriks IA, Larsen SC, Nielsen ML Abstract ADP-ribosylation is a widespread post-translational modification (PTM) with crucial functions in many cellular processes. Here, we describe an in-depth ADP-ribosylome using our Af1521-based proteomics methodology for comprehensive profiling of ADP-ribosylation sites, by systematically assessing complementary proteolytic digestions and precursor fragmentation through application of electron-transfer higher-energy collisional dissociation (EThcD) and electron transfer dissociation (ETD), respectively. While ETD spectra yielded higher identification scores, EThcD generally proved superior to ETD in identification and localization of ADP-ribosylation sites regardless of protease employed. Notwithstanding, the propensities of complementary proteases and fragmentation methods expanded the detectable repertoire of ADP-ribosylation to an unprecedented depth. This system-wide profiling of the ADP-ribosylome in HeLa cells subjected to DNA damage uncovered >11,000 unique ADP-ribosylated peptides mapping to >7,000 ADP-ribosylation sites, in total modifying over one-third of the human nuclear proteome and highlighting the vast scope of this PTM. High-resolution MS/MS spectra enabled identification of dozens of proteins concomitantly modifi...
Source: Molecular and Cellular Proteomics : MCP - Category: Molecular Biology Authors: Tags: Mol Cell Proteomics Source Type: research