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. Author manuscript; available in PMC: 2020 Nov 15.
Published in final edited form as: ACS Chem Biol. 2019 May 13;14(11):2430–2440. doi: 10.1021/acschembio.8b01083

Figure 3.

Figure 3.

Optimizing RNF4 covalent ligands using gel-based ABPP. (A) Derivatives of TRH 1–23 were tested against IA-rhodamine labeling of RNF4 using gel-based ABPP. Silver stained gels are also shown to visualize total protein content. (B) CCW16 was tested against IA-rhodamine labeling of RNF4 using gel-based ABPP. For parts A and B, covalent ligands were preincubated with pure RNF4 protein for 30 min prior to IA-rhodamine labeling for 1 h. Proteins were subjected to SDS/PAGE and visualized by in-gel fluorescence. In part B, gels were quantified by densitometry to calculate IC50 values. The gel shown in part B is a representative gel from n = 3. (C) Covalent docking of CCW 16 bound to either C132 or C135 in RNF4.