Metabolic engineering of 3‐hydroxypropionic acid biosynthesis in Escherichia coli

Abstract

3-Hydroxypropionic acid (3-HP) can be produced in microorganisms as a versatile platform chemical. However, owing to the toxicity of the intermediate product 3-hydroxypropionaldehyde (3-HPA), the minimization of 3-HPA accumulation is critical for enhancing the productivity of 3-HP. In this study, we identified a novel aldehyde dehydrogenase, GabD4 from Cupriavidus necator, and found that it possessed the highest enzyme activity toward 3-HPA reported to date. To augment the activity of GabD4, several variants were obtained by site-directed and saturation mutagenesis based on homology modeling. Escherichia coli transformed with the mutant GabD4_E209Q/E269Q showed the highest enzyme activity, which was 1.4-fold higher than that of wild type GabD4, and produced up to 71.9 g L(-1) of 3-HP with a productivity of 1.8 g L(-1) h(-1) . To the best of our knowledge, these are the highest 3-HP titer and productivity values among those reported in the literature. Additionally, our study demonstrates that GabD4 can be a key enzyme for the development of industrial 3-HP-producing microbial strains, and provides further insight into the mechanism of aldehyde dehydrogenase activity.

Biobased organic acids production by metabolically engineered microorganisms

“…A recent study has demonstrated that the second enzyme is crucial for 3-HP production 168 Food biotechnology as the accumulation of the intermediate product 3-HPA is toxic to the host cells [27 ]. By selecting the most efficient aldehyde dehydrogenase, GabD4 derived from Cupriavidus necator, combined with directed mutagenesis this enzyme activity was further increased and the highest 3-HP titer and productivity was achieved, up to 71.9 g/L of 3-HP with a productivity of 1.8 g/L/h using glucose and glycerol as co-carbon sources.…”

Biological Production of 3-Hydroxypropionic Acid: An Update on the Current Status

“…One solution to the problem could be the selection of an efficient aldehyde dehydrogenase that would rapidly act on 3-hydroxypropionaldehyde and convert it to 3HP. Based on this strategy, Chu et al [34] tried 17 candidate aldehyde dehydrogenases for their activity against 3-hydroxypropionaldehyde, with the gabD4 from Cupriavidus necator turning out to be the most effective one. In an attempt to further improve the aldehyde dehydrogenase selected, the authors performed site-directed and saturation mutagenesis, based on homologous modeling.…”

Bio-based 3-hydroxypropionic Acid: A Review

“…The second highest productivity of 3-HP in Table 2 is 1.8 g*L -1 *h -1 [20]. In the study a novel aldehyde dehydrogenase, GabD4 from Cupriavidus necator was identified and found that it possessed the highest enzyme activity on 3-HPA reported to [43] date.…”

“…The former strategy would reduce the overall rate of production. Therefore, the latter is considered more desirable [10,20].…”

“…The initial glycerol concentration was 80 g*L -1 and concentrated glycerol (700 g*L -1 ) was added to the culture broth as a feeding substrate to maintain the glycerol concentration. The recombinant E. coli produced 71.9 g*L -1 of 3-HP with a productivity of 1.8 g*L -1 *h -1 [20]. The third highest productivity of 3-HP in Table 2 is 1.75 g*L -1 *h -1 [33].…”