Fig. 3

Flavin absorption spectra of GSTACMO (a) and H325K (b) were recorded in 50 mM HEPES–NaOH pH 7.5 upon titration with NADP⁺ (0–250 μM). The inset shows the difference spectra. Absorbance changes at 467 nm for wild type GSTACMO (c) and H325K (d) were plotted as a function of NADP⁺ concentration and the data were fitted to a quadratic binding isotherm (Eq. 1), which gave K_d values of 21.1 ± 4.1 μM (wild type) and 0.50 ± 0.04 μM (H325K)