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. Author manuscript; available in PMC: 2019 Aug 4.
Published in final edited form as: Nat Chem Biol. 2019 Feb 4;15(4):391–400. doi: 10.1038/s41589-018-0217-y

Figure 1.

Figure 1.

Fumarate is a covalent oncometabolite. (a) Covalent labeling of cysteine residues by fumarate yields the PTM S-succination. (b) Applying S-succinated Cys immunoblotting to establish the concentration of fumarate required for covalent protein labeling. HEK-293 proteomes were treated with fumarate (0, 1, 5, 10 mM) for 15 h prior to western blotting. (c) Applying fumarate alkyne (FA-alkyne, 1) to visualize reactivity of the fumarate chemotype. HEK-293 proteomes were treated with FA-alkyne (0, 0.1, 0.5, 1 mM) for 15 h prior to click chemistry and SDS-PAGE. (d) Applying iodoacetamide alkyne (IA-alkyne, 2) as a competitive probe of covalent fumarate labeling. HEK-293 proteomes were incubated with fumarate for 15 h prior to treatment with 100 μM IA-alkyne for 1 h followed by desalting, click chemistry, and SDS-PAGE. Representative images from two independent experiments are shown in b-d. Uncropped scans of gels and immunoblots are provided in Supplementary Fig. 10.