Impact of expression of EMP enzymes on glucose metabolism in Zymomonas mobilis.

Zymomonas mobilis is the only known microorganism that utilizes the Entner-Doudoroff (ED) pathway anaerobically. In this work, we investigated whether the overexpression of a phosphofructokinase (PFK), the only missing Embden-Meyerhof-Parnas (EMP) pathway enzyme, could establish the pathway in this organism. Introduction of a pyrophosphate-dependent PFK, along with co-expression of homologous fructose-1,6-bisphosphate aldolase and triosephosphate isomerase, did not result in an EMP flux to any appreciable level.

Engineering efficient xylose metabolism into an acetic acid-tolerant Zymomonas mobilis strain by introducing adaptation-induced mutations.

The impact of the two adaptation-induced mutations in an improved xylose-fermenting Zymomonas mobilis strain was investigated. The chromosomal mutation at the xylose reductase gene was critical to xylose metabolism by reducing xylitol formation. Together with the plasmid-borne mutation impacting xylose isomerase activity, these two mutations accounted for 80 % of the improvement achieved by adaptation.

Transcriptome profiling of a curdlan-producing Agrobacterium reveals conserved regulatory mechanisms of exopolysaccharide biosynthesis.

Background: The ability to synthesize exopolysaccharides (EPS) is widespread among microorganisms, and microbial EPS play important roles in biofilm formation, pathogen persistence, and applications in the food and medical industries. Although it is well established that EPS synthesis is invariably in response to environmental cues, it remains largely unknown how various environmental signals trigger activation of the biochemical synthesis machinery.

Results: We report here the transcriptome profiling of Agrobacterium sp.

Discovery and characterization of a xylose reductase from Zymomonas mobilis ZM4.

Formation of xylitol, a byproduct from xylose fermentation, is a major limiting factor in ethanol production from xylose in engineered Zymomonas strains, yet the postulated xylose reductase remains elusive. We report here the discovery of xylose reductase in Zymomonas mobilis and, for the first time, to associate the enzyme function with its gene. Besides xylose and xylulose, the enzyme was active towards benzaldehyde, furfural, 5-hydroxymethyl furfural, and acetaldehyde, exhibiting nearly 150-times higher affinity with benzaldehyde than xylose.

Adaptation yields a highly efficient xylose-fermenting Zymomonas mobilis strain.

Zymomonas mobilis is a superb ethanol producer with productivity exceeding yeast strains by several fold. Although metabolic engineering was successfully applied to expand its substrate range to include xylose, xylose fermentation lagged far behind glucose. In addition, xylose fermentation was often incomplete when its initial concentration was higher than 5%.

Citrate stimulates oligosaccharide synthesis in metabolically engineered Agrobacterium sp.

Agrobacterium sp. ATCC 31749 was previously shown to be an advantageous host for oligosaccharide production. Unexpectedly, the addition of citrate to the oligosaccharide synthesis reaction resulted in up to a sixfold improvement in the production N-aceytl-lactosamine, a disaccharide.

Recombinant synthesis of hyaluronan by Agrobacterium sp.

Hyaluronan (HA) is a sugar polymer of a repeating disaccharide, beta1-3 D-N-acetylglucosamine (GlcNAc) beta1-4 D-glucuronic acid (GlcA). It finds applications in numerous biomedical procedures such as ophthalmic surgery and osteoarthritis treatment. Until recently, the only commercial sources were extraction of rooster combs and from fermentation of pathogenic Streptococcus.

A window into biocatalysis and biotransformations.

Eight papers were presented in this year's symposium "Advances in Biocatalysis" at the 232nd ACS National Meeting, accentuating the most recent development in biocatalysis. Researchers from both industry and academia are addressing several fundamental problems in biocatalysis, including the limited number of commercially available enzymes that can be provided in bulk quantities, the limited enzyme stability and activity in nonaqueous environments, and the permeability issue and cell localization problems in whole-cell systems. A trend that can be discerned from these eight talks is the infusion of new tools and technologies in addressing various challenges facing biocatalysis.

Permeability issues in whole-cell bioprocesses and cellular membrane engineering.

Nutrient uptake and waste excretion are among the many important functions of the cellular membrane. While permitting nutrients into the cell, the cellular membrane system evolves to guide against noxious agents present in the environment from entering the intracellular milieu. The semipermeable nature of the membrane is at odds with biomolecular engineers in their endeavor of using microbes as cell factory.

A type B feruloyl esterase from Aspergillus nidulans with broad pH applicability.

A hypothetical protein AN1772.2 of Aspergillus nidulans was found to have a 56% identity with a known type C ferulic acid esterase (FAE) from Talaromyces stipitatus. In addition, it contained a 13-amino acid conserved region flanking the characteristic G-X-S-X-G motif of a serine esterase, suggesting a FAE function for the protein.

A complete enzymatic recovery of ferulic acid from corn residues with extracellular enzymes from Neosartorya spinosa NRRL185.

An economic ferulic acid recovery from biomass via biological methods is of interest for a number of reasons. Ferulic acid is a precursor to vanillin synthesis. It is also a known antioxidant with potential food and medical applications.

Metabolic engineering of Agrobacterium sp. for UDP-galactose regeneration and oligosaccharide synthesis.

Curdlan-producing Agrobacterium sp. is unique in possessing a highly efficient UDP-glucose regeneration system. A broad-host-range expression strategy was successfully developed to exploit the unique metabolic capability for UDP-galactose regeneration during oligosaccharide synthesis.

Metabolic engineering of microbes for oligosaccharide and polysaccharide synthesis.

Metabolic engineering has recently been embraced as an effective tool for developing whole-cell biocatalysts for oligosaccharide and polysaccharide synthesis. Microbial catalysts now provide a practical means to derive many valuable oligosaccharides, previously inaccessible through other methods, in sufficient quantities to support research and clinical applications. The synthesis process based upon these microbes is scalable as it avoids expensive starting materials.

Engineering the E. coli UDP-glucose synthesis pathway for oligosaccharide synthesis.

A metabolic engineering strategy was successfully applied to engineer the UDP-glucose synthesis pathway in E. coli. Two key enzymes of the pathway, phosphoglucomutase and UDP-glucose pyrophosphorylase, were overexpressed to increase the carbon flux toward UDP-glucose synthesis.