Engineering yield and rate of reductive biotransformation in Escherichia coli by partial cyclization of the pentose phosphate pathway and PTS-independent glucose transport

Optimization of yields and productivities in reductive whole-cell biotransformations is an important issue for the industrial application of such processes. In a recent study with Escherichia coli, we analyzed the reduction of the prochiral β-ketoester methyl acetoacetate by an R-specific alcohol dehydrogenase (ADH) to the chiral hydroxy ester (R)-methyl 3-hydroxybutyrate (MHB) using glucose as substrate for the generation of NADPH. Deletion of the phosphofructokinase gene pfkA almost doubled the yield to 4.8 mol MHB per mole of glucose, and it was assumed that this effect was due to a partial cyclization of the pentose phosphate pathway (PPP). Here, this partial cyclization was confirmed by 13C metabolic flux analysis, which revealed a negative net flux from glucose 6-phosphate to fructose 6-phosphate catalyzed by phosphoglucose isomerase. For further process optimization, the genes encoding the glucose facilitator (glf) and glucokinase (glk) of Zymomonas mobilis were overexpressed in recombinant E. coli strains carrying ADH and deletions of either pgi (phosphoglucose isomerase), or pfkA, or pfkA plus pfkB. In all cases, the glucose uptake rate was increased (30–47%), and for strains Δpgi and ΔpfkA also, the specific MHB production rate was increased by 15% and 20%, respectively. The yield of the latter two strains slightly dropped by 11% and 6%, but was still 73% and 132% higher compared to the reference strain with intact pgi and pfkA genes and expressing glf and glk. Thus, metabolic engineering strategies are presented for improving yield and rate of reductive redox biocatalysis by partial cyclization of the PPP and by increasing glucose uptake, respectively

Fossile Flora der Steinkohlen-Formation Westphalens einschliesslich Piesberg bei Osnabrueck

von E. von Roeh

Assessment of smectite-rich claystones from Northpatagonia for their use as liner materials in landfills

Two types of Northpatagonic claystones (a commercially available sodium bentonite and selected smectite-rich claystones from the Upper Cretaceous Allen Formation) are considered as potential materials for landfill liners. To check the suitability of the claystones, physical (hydraulic conductivity, compaction, consolidation, and swelling), chemical (cation exchange capacity and bulk chemical analysis) and mineralogical analysis (X-ray diffraction and scanning electron microscope) were performed. Both claystone types showed very similar properties, which allow us to classify the smectite-rich claystones as bentonites. The high compressibility of the claystones makes them unsuitable to use them directly as liner materials. On the other hand, in combination with well graded sand, these materials satisfy the requirements stipulated by internationally accepted standards for compacted clay liners in municipal waste disposal repositories. In this way, a new applicability for clay materials of regional provenance was found

Suitability of a Cretaceous Natural Na-Bentonite as Construction Material for Landfill Liners

A natural Cretaceous sodium (Na) bentonite from North Patagonia, Argentine, is considered a potential construction material for landfill liners. It is studied alone and in mixtures with sand. Physical and chemical properties of the Na-bentonite were determined to verify the suitability of this material for the construction of low permeability mineral liners. Hydraulic conductivity measurements were conducted on pure bentonite and on mixtures of poorly graded sand with 3, 6, 9 and 12% of Na-bentonite. Compaction and hydraulic conductivity tests were performed using deionized water and a 1000 mol/m3 CaCl2 solution to determine the effects of a high concentrated saline solution on the hydraulic properties of compacted sand-bentonite mixtures. The experimental results confirm that the pure powdered bentonite meets current specifications to be used as part of a geosynthetic clay liner (GCL), and that ageing effects may affect the hydraulic behavior of this material. Compacted sand-bentonite mixtures required a minimum bentonite content of 6% in order to achieve hydraulic conductivities lower than 1 x 10-9 m/s which is a regular requirement for hydraulic containment liners. However, in contact with a strong saline solution (1000 mol/m3 CaCl2), the permeability of the mixtures increased between three and five orders of magnitude. This indicates that a higher percent of bentonite is required to reach an acceptable hydraulic conductivity value when the mixtures are permeated with a CaCl2 solution. The results show that a new applicability was found for bentonites of regional provenance as well as high reserves which have a naturally very high amount of sodium