(A) Cyclic
voltammetry measuring bidirectionality of IDH1-catalyzed
2OG/isocitrate interconversion at different 2OG/isocitrate ratios.
(B) Cyclic voltammetry measuring the ability of R132H to catalyze
isocitrate oxidation. Conditions (A and B): electrode area 0.03 cm2, scan rate 1 mV/s, temperature 25 °C, volume 4 mL, 10
mM MgCl2. (A) (FNR+IDH1+CA)@ITO/PGE electrode, 15 μM
NADP+, 15 μM NADPH; HEPES and NaHCO3 at
0.10 M; rotated at 1000 rpm. Magenta trace: 10 mM 2OG + 200 μM
isocitrate; red trace: 10 mM each: 2OG, isocitrate. (B) (FNR+R132H)@ITO/PGE
electrode, 5 μM NADP+; 20 mM each: MES, TAPS, CHES;
stationary electrode. Purple trace: 10 mM each: 2OG, isocitrate. Enzyme
loading ratios (molar): (A) FNR/IDH1/CA; 1/0.5/0.25; (B) FNR/R132H;
1/2.5.