Fig. 3.

Enzymatic activities of recombinant EcDQD/SDHs. Recombinant proteins were assayed at 30 °C in 4 mM 3-DHQ at pH 7 (a), 12 mM 3-DHS and 0.4 mM NAD(P)H at pH 6.5 (b), 12 mM shikimate and 2 mM NAD(P)⁺ at pH 9 (c), 30 mM 3-DHS and 6 mM NAD(P)⁺ at pH 10.5 (d), 8 mM 3-DHQ and 0.4 mM NAD(P)H at pH 7.5 (e), or 6 mM quinate and 1 mM NAD(P)⁺ at pH 9 (f). Enzymatic activities were determined by measuring the production of 3-DHS (a) or gallate (d) by HPLC or by monitoring the consumption (b, e) or production (c, f) of NAD(P)H spectrophotometrically. e*Activity was determined by measuring the quinate production by HPLC and not by monitoring NADPH consumption because the reactions from 3-DHQ to shikimate through 3-DHS also consume NADPH. Data are presented as the mean ± SD (n = 3). Activities less than 10 nkat mg⁻¹ (a, b, e) or 1 nkat mg⁻¹ (c, d, f) are indicated as “not detected (ND)”