Engineering of glycerol utilization in Gluconobacter oxydans 621H for biocatalyst preparation in a low-cost way

Abstract

BackgroundWhole cells of Gluconobacter oxydans are widely used in various biocatalytic processes. Sorbitol at high concentrations is commonly used in complex media to prepare biocatalysts. Exploiting an alternative process for preparation of biocatalysts with low cost substrates is of importance for industrial applications.Results G. oxydans 621H was confirmed to have the ability to grow in mineral salts medium with glycerol, an inevitable waste generated from industry of biofuels, as the sole carbon source. Based on the glycerol utilization mechanism elucidated in this study, the major polyol dehydrogenase (GOX0854) and the membrane-bound alcohol dehydrogenase (GOX1068) can competitively utilize glycerol but play no obvious roles in the biocatalyst preparation. Thus, the genes related to these two enzymes were deleted. Whole cells of G. oxydans ∆GOX1068∆GOX0854 can be prepared from glycerol with a 2.4-fold higher biomass yield than that of G. oxydans 621H. Using whole cells of G. oxydans ∆GOX1068∆GOX0854 as the biocatalyst, 61.6 g L−1 xylonate was produced from 58.4 g L−1 xylose at a yield of 1.05 g g−1.ConclusionThis process is an example of efficient preparation of whole cells of G. oxydans with reduced cost. Besides xylonate production from xylose, other biocatalytic processes might also be developed using whole cells of metabolic engineered G. oxydans prepared from glycerol.Electronic supplementary materialThe online version of this article (10.1186/s12934-018-1001-0) contains supplementary material, which is available to authorized users.

Selective oxidation of alkyl and aryl glyceryl monoethers catalysed by an engineered and immobilised glycerol dehydrogenase

“…23 The best candidates to catalyse such selective oxidation would be the glycerol dehydrogenases (GlyDH) due to their natural substrate specicity. Biotechnological applications of these enzymes however are mainly limited to the whole cell production of dihydroxyacetone 24 and valuable diols through metabolic engineering. 25 Besides, this family of enzymes has also been widely applied in biosensing as the active phase of glycerol sensors.…”

Coculture of Gluconobacter oxydans and Escherichia coli for 3,4-Dihydroxybutyric Acid Production from Xylose

“…When cultivated using glucose as carbon source, G. oxydans converted most of the substrate into gluconate via mGDH; , gluconate was then metabolized for biomass generation through the pentose phosphate pathway or the Entner–Doudoroff pathway (Figure A). ,, When cultivated with xylose, G. oxydans oxidized some of the xylose to xylonate via mGDH, , generating electrons for energy but a limited increase in biomass. When both glucose and xylose were provided, the increase in biomass was greater than that with either substrate alone (Figure A and Figure S1).…”

“…There are at least 10 membrane-bound dehydrogenases encoded in the G. oxydans 621H genome, , each oxidizing a wide range of compounds. Utilizing the enzymatic activities of two or more membrane-bound dehydrogenases results in the combination of biosynthetic processes and may yield new applications for G. oxydans .…”

“…11 G. oxydans 621H was cultivated in complex medium with shaking at 200 rpm and 30 °C. 20 Antibiotics were supplemented at following concentrations when required: 50 μg/mL kanamycin (Km), 40 μg/mL chloramphenicol (Cm), 50 μg/mL spectinomycin (Spe), and 50 μg/mL cefoxitin (Cef), respectively.…”

Repurposing the Endogenous Type I-E CRISPR/Cas System for Gene Repression in Gluconobacter oxydans WSH-003

“…A series of appropriate artificial poly­(A/T) tails was reported that could enhance the mRNA abundance of the sldhAB gene coding sorbitol dehydrogenase . Several shuttle plasmids were constructed for gene overexpression via inserting the par-rep gene of cryptic plasmid from G. oxydans into pUC18 or pUC19. , Two markerless gene knockout methods were established based on 5-fluorouracil and sucrose as counter-selection reagents. − These investigations contributed to the metabolic engineering of G. oxydans . However, the genetic tools for industrial strains G. oxydans , compared with Escherichia coli, Saccharomyces cerevisiae , and other model microorganisms, are relatively scarce.…”