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Background: D-xylonic acid is a versatile platform chemical with wide potential applications as the water reducer and disperser for cement and the precursor for 1,4-butanediol and 1,2,4-tributantriol. Microbial production of D-xylonic acid with bacteria like Gluconobacter oxydans from cheap lignocellulosic feedstock is generally regarded as one of the most promising and cost-effective methods to industrial production, but xylonic acid productivity is reduced by high substrate inhibition and hydrolysate inhibitors. Results: D-xylonic acid productivity of G. oxydans DSM2003 was improved by overexpressing the mGDH gene, which encoded the membrane-bound glucose dehydrogenase. Using the mutated plasmids based on pBBR1MCS-5 in our previous work, the recombinant strain G. oxydans/pBBR-R3510-mGDH with the significant improvement in D-xylonic acid production and the strengthened tolerance to hydrolysate inhibitors was obtained. The fed-batch biotransformation of D-xylose by this recombinant strain reached the record high titer (588.7 g/L), yield (99.4%), and space-time yield (8.72 g/L/h). Moreover, up to 246.4 g/L D-xylonic acid was produced directly from corn stover hydroxylates without detoxification at a yield of 98.9% and a space-time yield of 11.2 g/L/h. In addition, G. oxydans/pBBR-R3510-mGDH performed strong tolerance to typical inhibitors, i.e., formic acid, furfural, and 5-hydroxymethylfurfural. Conclusion: Through overexpression of mgdh in G. oxydans, we obtained a recombinant strain G. oxydans/pBBR-R3510-mGDH. It was capable to efficiently produce xylonic acid from the corn stover hydrolysate under high concentrations of inhibitors. The high D-xylonic acid productivity made G. oxydans/pBBR-R3510-mGDH an attractive choice for biotechnical production.

Objectives: 3,4-Dihydroxybutyric acid (3,4-DHBA) is a multi-functional C4 platform compound with wide applications in the synthesis of materials and pharmaceuticals. Currently, although the biosynthetic pathway for the production of 3,4-DHBA has been developed, low productivity still hampers its use on large scales. Here, a non-natural four-steps biosynthetic pathway was established in recombinant E. coli with a combinatorial strategy.Results: Firstly, several aldehyde dehydrogenases (ALDHs) were screened and characterized for catalyzing the dehydrogenation of 3,4-dihydroxybutanal (3,4-DHB) to 3,4-DHBA through in vitro enzyme assays. Secondly, a recombinant E. coli was successfully constructed to generate 3,4-DHBA from D-xylose by introducing the pathway containing BsGDH, YagF, PpMdlC and ALDH into E. coli with 3.04 g/L 3,4-DHBA obtained. Then, disruption of competing pathways by deleting xylA, ghrA, ghrB and adhP genes contributed to increase the accumulation of 3,4-DHBA by 87%. Final, fusion expression of PpMdlC and YagF resulted in an enhancement of 3,4-DHBA titer (7.71 g/L), as the highest titer reported so far.Conclusions: These results showed that deleting competing pathways and constructing fusion protein could significantly improve the 3,4-DHBA titer in engineered E. coli.