Fig. 4

Evaluation of in vitro enzyme pathways for production of glycolate (a), lactate (b), and ethyleneglycol (c) (see also Fig. 1). In each case, the substrate for the pathway was xylonolactone and the pathway was composed of 4 enzymes, where the first three enzymes are common and the 4th is varied depending on the desired product. The production of glycolate was followed through generation of NADH, the production of lactate through generation of NADH, and the production of ethylene glycol was followed through generation of NAD⁺ by the final enzymatic step catalysed by Ec AldA (a), LDH (b), and Ec FucO (c) respectively. The reaction conducted at 22 °C contained 50 mM Tris HCl pH 7.0, 10 mM MgCl₂, 0.5 mM DTT, 2 mM NAD⁺ or NADH, 1 mM xylonolactone, 4 µg Cc XylC, 2 µg Cc XylD, 2 µg Ec YagE. In a the amount of Ec AldA was varied 0 µg (black line), 2 µg (magenta line), 4 µg (cyan line), 8 µg (blue line), 16 µg (green line) and 24 µg (red line). In b the amount of LDH was varied 0 µg (black line), 0.1 µg (cyan line), 0.2 µg (blue line), 0.4 µg (green line), 0.8 µg (red line). In c the amount of Ec FucO was varied 0 µg (black line), 1 µg (cyan line), 2 µg (blue line), 4 µg (green line), 8 µg (red line)