pH dependence of kinetic parameters on pyruvate reduction. White, grey, and black circles indicate FNLDH, PALDH, and ECLDH, respectively. The kcat(a), S0.5(b), and kcat/S0.5 data (c) are plotted logarithmically, and the Hill coefficient (d) is plotted linearly. The buffers used for the assay were sodium acetate buffer (pH 4.5, 5.0, and 5.5; circles), MES-NaOH buffer (pH 5.5, 6.0, and 6.5; triangles), MOPS-NaOH buffer (pH 6.5, 7.0, and 7.5; squares), HEPES-NaOH buffer (pH 7.0, 7.5 and 8.0; diamonds), and Bicine-NaOH buffer (pH 8.0, 8.5, and 9.0; hexagons). We adjusted the pH of each buffer solution prior to the addition of substrate and cofactor, and confirmed that the pH value is not affected even in the presence of high concentrations of substrate.