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. Author manuscript; available in PMC: 2021 Feb 23.
Published in final edited form as: ACS Chem Biol. 2017 Oct 17;12(12):2922–2926. doi: 10.1021/acschembio.7b00734

Figure 4.

Figure 4.

Profiling ac4C using its specific reactivity. (a) Scheme for chemical deacetylation of ac4C. (b) Effect of hydroxylamine on ac4C free nucleoside monitored by UV spectroscopy. Reaction conditions: 0.25 mM ac4C [free nucleoside], 50 mM hydroxylamine, Tris at pH 7.0. (c) Effect of hydroxylamine on ac4C in RNA monitored by anti-ac4C dot blot. Reaction conditions: 50 mM hydroxylamine, Tris at pH 8.0, 65 °C, 1 h. (d) Effect of hydroxylamine (50 mM, pH 7.0) on overall cytidine and uridine base content in total RNA analyzed by HPLC. (e) Effect of hydroxylamine (50 mM, pH 7.0, 65 °C, 1 h) on ac4C in total cellular RNA analyzed by LC-MS/MS. (f) Anti-ac4C antibody can detect chemical ablation of ac4C in total cellular RNA.