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. Author manuscript; available in PMC: 2019 May 8.
Published in final edited form as: J Am Chem Soc. 2018 Apr 2;140(14):4757–4760. doi: 10.1021/jacs.7b13141

Figure 2.

Figure 2

A chemo-proteomic probe for protein lysine crotonylation. (a) Scheme depicting the application of TCEP analog 4 in the detection of protein crotonylation. (b) Semisynthetic mononucleosomes bearing indicated modifications were reacted with probe 4 (4 mM) before being analyzed by western-blot employing a biotin detection system (dye-conjugated streptavidin). For comparison, the same nucleosomes were analyzed by western blotting employing commonly used pan anti-lysine crotonyl (α-pCr) and pan anti-lysine acetyl (α-pAc) antibodies. A coomassie stained SDS-PAGE gel of the samples was used as a loading control. (c) Mass analysis of H3K18Cr conjugated with probe 4. Following the reaction, the protein was digested with ArgC and analyzed by LC-MS/MS. In addition to the expected y- and b-ions, fragmentation of the probe itself (lower right panel) is observed and denoted as x-ions.