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Demethylation assay assessing kinetic activity of mutant vs WT FTO proteins. FTO activity is proportional to the concentration present in the reaction. Demethylase activity is likely to be related to the ability of FTO to function as a sensor for cellular metabolism (29). The R316Q mutant is enzymatically dead across all concentrations tested. The A163T and L44V mutants showed demethylase activity toward methylated uridine in a dose-dependent manner but with different affinities.
