Notable Improvement of 3-Hydroxypropionic Acid and 1,3-Propanediol Coproduction Using Modular Coculture Engineering and Pathway Rebalancing

Abstract

Sustainable bioconversion of glycerol into 3hydroxypropionic acid (3-HP) and 1,3-propanediol (1,3-PDO) is often bottlenecked by the accumulation of a toxic metabolic intermediate, 3-hydroxypropanaldehyde (3-HPA), mostly due to the imbalanced expression of the two key enzymes, namely, aldehyde dehydrogenase (AldH) and 1,3-PDO oxidoreductase (PDOR). Herein, we attempted to overcome it by modular coculture engineering using Lactobacillus reuteri and recombinant Escherichia coli strains through overexpressing an AldH (GabD4) and a PDOR (PduQ) in E. coli and pathway rebalancing via 5′untranslated region (5′-UTR) engineering in the gabD4 gene. We then used the L. reuteri−E. coli coculture systems in resting cell biocatalysis of glycerol under several technological configurations to produce 3-HP and 1,3-PDO. The conventional approach, which is a two-step process comprising resting cell production (first step) and biotransformation of glycerol with resting cells (second step), produced 51.36 g/L 3-HP and 34.27 g/L 1,3-PDO from 120 g/L glycerol, leaving more than 30 g/L glycerol unconverted. Subsequently, we did experiments in a modified approach, including one more biotransformation step with the two-step approach to convert the residual glycerol. This three-step approach increased glycerol consumption to 140 g/L and produced 71.62 g/L 3-HP and 47.68 g/L 1,3-PDO. Finally, the sequential feeding of glycerol and fresh cells notably improved glycerol consumption to 240 g/L and titers of 3-HP and 1,3-PDO to 125.93 and 88.46 g/L, respectively, with a net titer of 214.39 g/L, which was the highest titer ever reported. Therefore, modular coculture engineering with an appropriate technological configuration could have the potential for improved production of value-added chemicals, particularly 3-HP and 1,3-PDO.

Recent advances in biological production of 1,3-propanediol: new routes and engineering strategies

“…By cultivating L. reuteri, which can secrete 3-HPA, with E. coli overexpressing the aldehyde dehydrogenase gene gabD4 and the 1,3-PDO dehydrogenase gene pduQ, the engineered system can accumulate 125.93 g L −1 3-HP and 88.46 g L −1 1,3-PDO in fed-batch fermentation. 61 Another cocultivation system consisted of two recombinant E. coli strains, one overexpressing dhaB1B2 from C. butyricum and another overexpressing dhaT from K. pneumoniae. This co-fermentation system took glycerol and glucose as substrates and produced 41.65 g L −1 1,3-PDO with a yield of 0.67 mol mol −1 glycerol.…”

Heterologous Production of Acrylic Acid: Current Challenges and Perspectives

“…For this reason, the work regarding 3-HP production in this organism has been patented [44,45]. Additionally, to improve the concentrations, a co-culture engineering strategy with Lactobacillus reuteri and E. coli was recently used [48]. L. reuteri is able to naturally synthesize 3-HP but with high accumulation of 3-HPA.…”

“…In the past years, AA heterologous production has significantly evolved due to all the efforts made in the past focussing on the biosynthesis of 3-hydroxypropionic acid (3-HP), a possible intermediary compound in AA biosynthesis. 3-HP bio-based production has been intensively studied, and up to 125.93 g/L concentrations have already been obtained using different pathways and hosts [35][36][37][38][39][40][41][42][43][44][45][46][47][48][49][50][51][52][53]. This acid can be converted to AA by catalysis (dehydrogenation) or, more recently, by fermentation.…”

Design of a self-stable and population-controllable co-culture system in E. coli and its applications

“…Maintaining the stability of the strain population ratio can be di cult [3][4][5][6][7] . Despite this, many remarkable achievements using modular co-cultivation engineering have been reported [11][12][13][14][15][16][17] . These studies showed the advantages of modularity, but the population ratio did not maintain long-term balance, and each strain grew relatively independently because there was no growth-related relationship among the cocultured strains.…”