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. 2014 Nov 17;15(11):21179–21201. doi: 10.3390/ijms151121179

Figure 6.

Figure 6

8-Anilinonaphthalene-1-sulphonate (ANS) and 5,5'-dithiobis-2-nitrobenzoic acid (DTNB) accessibility assays of WT G6PD and the four clinical variants. (A) ANS fluorescence spectra of WT G6PD and the four clinical mutants were obtained using an excitation wavelength of 395 nm and recording emission spectra from 400 to 600 nm; the final concentrations of ANS and enzyme were 165 μM and 400 μg/mL, respectively. Values obtained from buffer containing ANS without protein (open stars) were subtracted from the recordings with protein; and (B) The derivatization of Cys residues in G6PD (200 µg/mL) in 1 mM DTNB was followed spectrophotometrically at 412 nm. The arrow indicates the time of addition of 5% SDS, after which the number of Cys per monomer was calculated for the enzymes. Each trace is the average of two independent experiments.