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Oxidation of the C117A mutant by modified tRNALeu. Anaerobic enzyme (15 μm) in 100 mm HEPES, pH 7.5, 100 mm NaCl, 10 mm MgCl2, 10% glycerol was reduced by titrating with one equivalent of dithionite. The reduced enzyme was reoxidized by mixing with anaerobic modified tRNALeu. The inset shows the time dependence of flavin oxidation, detected at 450 nm. The trace was fit to a single exponential, giving a rate constant of 8.7 × 10–5 ± 3 × 10–6 s–1.
