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Cell Dry Weight (cell + dry_weight)
Selected AbstractsEnhancement of the NAD(P)(H) Pool in Escherichia coli for BiotransformationENGINEERING IN LIFE SCIENCES (ELECTRONIC), Issue 4 2007F. Heuser Abstract In pyridine nucleotide-dependent, reductive whole cell biotransformation with resting cells of Escherichia coli, the availability of intracellular NAD(P)(H) is a pivotal point for an efficient and highly productive substrate conversion. The question whether an increase of the intracellular NAD(P)(H) concentration could increase the productivity was discussed controversially in the past. This is the first report on an E. coli strain with an increased NAD(P)(H) pool which was tested in a reductive biotransformation system for an increased productivity. Biotransformation was performed with a strain overexpressing a gene encoding an (R)-specific alcohol dehydrogenase for the stereospecific, NADPH-dependent reduction of methyl acetoacetate (MAA) to (R)-methyl-3-hydroxybutanoate (MHB). Cofactor regeneration was implemented via glucose oxidation by coexpression of a gene encoding glucose dehydrogenase. The specific MHB productivity (mmol mg,1 cell dry weight,1h,1) enabled a comparison between the E. coli,BL21(DE3) wild-type and a genetically modified strain. The enhancement of the NAD(P)(H) pool was achieved by genetic manipulation of the NAD(H) biosynthetic pathways. After simultaneous overexpression of the pncB and nadE genes, encoding nicotinic acid phosphoribosyltransferase and NAD synthetase, measurements of the total NAD(P)(H) pool, sizes showed a 7-fold and 2-fold increased intracellular concentration of NAD(H) and NADP(H), respectively. However, the implementation of an E.,coli strain carrying a genomically integrated pncB gene with an upstream T7,promoter for biotransformation did not result in reproducible increased specific cell productivity. [source] Screening, Characterization and Application of Cyanide-resistant Nitrile HydratasesENGINEERING IN LIFE SCIENCES (ELECTRONIC), Issue 6 2004T. Gerasimova Abstract Two new bacterial strains, Pseudomonas marginales MA32 and Pseudomonas putida MA113, containing nitrile hydratases resistant to cyanide were isolated from soil samples by an enrichment procedure. In contrast to known nitrile hydratases, which rapidly lose activity at low to moderate cyanide concentrations, the enzymes described in this paper tolerate up to 50 mM cyanide. They show a broad substrate spectrum including not only small substrates like acrylonitrile but also nitriles with longer side chains and even nitriles with quarternary alpha-carbon atoms. Both characteristics are essential for the transformation of ketone cyanohydrins, which are much more instable and therefore releasing much higher amounts of prussic acid than cyanohydrins formed from aldehydes. P. marginales MA32 was used as a whole cell biocatalyst for the hydration of acetone cyanohydrin to ,-Hydroxyisobutyramide, which is a precursor of methacrylamide, an important pre-polymer. After optimization of the process conditions a maximum amide concentration of more than 1.6 M could be reached within 5 hours with 5,g/L biocatalyst referred to cell dry weight. [source] Process optimization of the integrated synthesis and secretion of ectoine and hydroxyectoine under hyper/hypo-osmotic stressBIOTECHNOLOGY & BIOENGINEERING, Issue 1 2010C. Fallet Abstract The synthesis and secretion of the industrial relevant compatible solutes ectoine and hydroxyectoine using the halophile bacterium Chromohalobacter salexigens were studied and optimized. For this purpose, a cascade of two continuously operated bioreactors was used. In the first bioreactor, cells were grown under constant hyperosmotic conditions and thermal stress driving the cells to accumulate large amounts of ectoines. To enhance the overall productivity, high cell densities up to 61,g,L,1 were achieved using a cross-flow ultrafiltration connected to the first bioreactor. In the coupled second bioreactor the concentrated cell broth was subjected to an osmotic and thermal down-shock by addition of fresh distilled water. Under these conditions, the cells are forced to secrete the accumulated intracellular ectoines into the medium to avoid bursting. The cultivation conditions in the first bioreactor were optimized with respect to growth temperature and medium salinity to reach the highest synthesis (productivity); the second bioreactor was optimized using a multi-objective approach to attain maximal ectoine secretion with simultaneous minimization of cell death and product dilution caused by the osmotic and thermal down-shock. Depending on the cultivation conditions, intracellular ectoine and hydroxyectoine contents up to 540 and 400,mg per g cell dry weight, respectively, were attained. With a maximum specific growth rate of 0.3,h,1 in defined medium, productivities of approximately 2.1,g,L,1,h,1 secreted ectoines in continuous operation were reached. Biotechnol. Bioeng. 2010;107: 124,133. © 2010 Wiley Periodicals, Inc. [source] The utilization of glycogen accumulating organisms for mixed culture production of polyhydroxyalkanoatesBIOTECHNOLOGY & BIOENGINEERING, Issue 4 2009*Article first published online: 15 JUN 200, Simon Bengtsson Abstract Production of polyhydroxyalkanoates (PHAs) by an open mixed culture enriched in glycogen accumulating organisms (GAOs) under alternating anaerobic,aerobic conditions with acetate as carbon source was investigated. The culture exhibited a stable enrichment performance over the 450-day operating period with regards to phenotypic behavior and microbial community structure. Candidatus Competibacter phosphatis dominated the culture at between 54% and 70% of the bacterial biomass throughout the study, as determined by fluorescence in situ hybridization. In batch experiments under anaerobic conditions, PHA containing 3-hydroxybutyrate (3HB) and 27,mol-% 3-hydroxyvalerate (3HV) was accumulated up to 49% of cell dry weight utilizing the glycogen pool stored in the SBR cycle. Under aerobic and ammonia limited conditions, PHA comprising only 3HB was accumulated to 60% of cell dry weight. Glycogen was consumed during aerobic PHA accumulation as well as under anaerobic conditions, but with different stoichiometry. Under aerobic conditions 0.31 C-mol glycogen was consumed per consumed C-mol acetate compared to 0.99 under anaerobic conditions. Both the PHA biomass content and the specific PHA production rate obtained were similar to what is typically obtained using the more commonly applied aerobic dynamic feeding strategy. Biotechnol. Bioeng. 2009; 104: 698,708 © 2009 Wiley Periodicals, Inc. [source] Carbon metabolism and product inhibition determine the epoxidation efficiency of solvent-tolerant Pseudomonas sp. strain VLB120,CBIOTECHNOLOGY & BIOENGINEERING, Issue 6 2007Jin-Byung Park Abstract Utilization of solvent tolerant bacteria as biocatalysts has been suggested to enable or improve bioprocesses for the production of toxic compounds. Here, we studied the relevance of solvent (product) tolerance and inhibition, carbon metabolism, and the stability of biocatalytic activity in such a bioprocess. Styrene degrading Pseudomonas sp. strain VLB120 is shown to be solvent tolerant and was engineered to produce enantiopure (S)-styrene oxide from styrene. Whereas glucose as sole source for carbon and energy allowed efficient styrene epoxidation at rates up to 97 µmol/min/(g cell dry weight), citrate was found to repress epoxidation by the engineered Pseudomonas sp. strain VLB120,C emphasizing that carbon source selection and control is critical. In comparison to recombinant Escherichia coli, the VLB120,C-strain tolerated higher toxic product levels but showed less stable activities during fed-batch cultivation in a two-liquid phase system. Epoxidation activities of the VLB120,C-strain decreased at product concentrations above 130 mM in the organic phase. During continuous two-liquid phase cultivations at organic-phase product concentrations of up to 85 mM, the VLB120,C-strain showed stable activities and, as compared to recombinant E. coli, a more efficient glucose metabolism resulting in a 22% higher volumetric productivity. Kinetic analyses indicated that activities were limited by the styrene concentration and not by other factors such as NADH availability or catabolite repression. In conclusion, the stability of activity of the solvent tolerant VLB120,C-strain can be considered critical at elevated toxic product levels, whereas the efficient carbon and energy metabolism of this Pseudomonas strain augurs well for productive continuous processing. Biotechnol. Bioeng. 2007;98: 1219,1229. © 2007 Wiley Periodicals, Inc. [source] Genetic modification of the marine-derived yeast Yarrowia lipolytica with high-protein content using a GPI-anchor-fusion expression systemBIOTECHNOLOGY PROGRESS, Issue 5 2009Fang Wang Abstract Yarrowia lipolytica SWJ-1b isolated from the marine fish gut was found to contain 47.6 g of crude protein per 100 g of cell dry weight and had potential use as single cell protein. When the gene encoding enhanced green fluorescent protein (EGFP) was inserted into the surface display plasmid pINA1317-YlCWP110 and expressed in uracil mutant of Y. lipolytica SWJ-1b, the corresponding protein was successfully displayed on the cell surface, and 100% of the yeast cells exhibited the anchored target proteins. We found that yeast cells displaying EGFP were similar to those of Y. lipolytica SWJ-1b. Furthermore, C18:1 and C18:3 fatty acids biosynthesis in the marine yeast cells displaying the heterologous EGFP was weakly enhanced compared with that in its wild-type. The results suggest that the marine-derived Y. lipolytica SWJ-1b can be armed with the heterologous protein by the genetic modification and further used as single cell protein. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009 [source] Effect of Overexpression of a Soluble Pyridine Nucleotide Transhydrogenase (UdhA) on the Production of Poly(3-hydroxybutyrate) in Escherichia coliBIOTECHNOLOGY PROGRESS, Issue 2 2006Ailen M. Sánchez A soluble pyridine nucleotide transhydrogenase (UdhA) has been used to increase the productivity and yield of PHB in vivo. By inducing a high level of UdhA, which can transfer reducing equivalents between NAD and NADP, we have increased NADPH availability, resulting in high yield and productivity of PHB in Escherichia coli. Coexpression of the phboperon from Alcaligenes eutrophusH16 and the native udhAfrom E. coli from high copy plasmids resulted in an increase in PHB yield from 49 to 66% g of PHB per gram of total cell dry weight and an increase in final concentration from 3.52 to 6.42 g/L; the PHB concentration of the udhA carrying strain is almost twice that of the control strain expressing only the phb operon. The results of this study demonstrate the effectiveness of cofactor manipulation and its application as a tool in metabolic engineering. [source] Static Magnetic Fields Enhancement of Saccharomyces cerevisae Ethanolic FermentationBIOTECHNOLOGY PROGRESS, Issue 1 2004Mauricy Alves da Motta Magnetic effects induced in ethanolic fermentation by Saccharomycescerevisiae strain DAUFPE-1012 were studied during a 24 h exposure to 220 mT steady magnetic fields (SMF) at 23 ± 1 °C, produced by NdFeB rod magnets. The magnets were attached diametrically opposed (N to S) to a cylindrical tube reactor. The biomass growth in the reactor culture media (yeast extract + glucose 2%) during 24 h was monitored by measurements of optical density, which was correlated to cell dry weight. Ethanol concentration and glucose level were measured every 2 h. The pH of the culture media was maintained between 4 and 5. As a result, biomass (g/L) increased 2.5-fold and ethanol concentration 3.4-fold in magnetized cultures ( n = 8) as compared with SMF nonexposed cultures ( n = 8). Glucose consumption was higher in magnetized cultures, which correlated to the ethanol yield. [source] Continuous Cultivation of the Diatom Nitzschia laevis for Eicosapentaenoic Acid Production: Physiological Study and Process OptimizationBIOTECHNOLOGY PROGRESS, Issue 1 2002Zhi-You Wen The continuous cultures of the diatom Nitzschia laevis were performed at different dilution rates ( D) and feed glucose concentrations ( S0) to investigate cellular physiological responses and its production potential of eicosapentaenoic acid (EPA). Steady-state cell dry weight, residual glucose concentration, cell growth yield, specific glucose consumption rate, and fatty acid profiles were investigated within the range of D from 0.1 to 1.0 day,1 ( S0 fixed at 20 g/L) and the range of S0 from 5 to 35 g/L ( D fixed at 0.3 day,1), respectively. The highest EPA productivity of 73 mg L -1 day -1 was obtained at D = 0.5 day,1 and S0 = 20 g/L. However, when the continuous culture achieved high productivities of EPA at certain dilution rates and feed glucose concentrations, glucose in the feed could not be consumed completely. Accordingly, the continuous culture was evaluated in terms of both EPA productivity ( P) and glucose assimilation efficiency ( E). The parameter ,, defined as the product of P and E, was used as an overall performance index. Since , is a function of the two independent variables Dand S0, we employed a central composite design to optimize D and S0 for the highest , value. Based on the experimental results of the design, a second-order polynomial equation was established to represent the relationship between , and D and S0. The optimal values of D and S0 were subsequently determined as 0.481 day,1 and 15.56 g/L, respectively by the empirical model. The verification experiment confirmed the validity of the model. Under the optimal conditions, , value reached 46.5 mg L -1 day -1, suggesting a considerably high efficiency of the continuous culture of N. laevis in terms of EPA production and glucose utilization. [source] Toxicity Effect of Pb(II) on Two Different Kinds of Microbes Measured by MicrocalorimetryCHINESE JOURNAL OF CHEMISTRY, Issue 3 2009Nan GAI Abstract In this work, microcalorimetric technique was used to analyze Pb(II) toxic action on the metabolic activities of Candida humicola and Bacillus subtilis. The experimental results revealed that Pb(II) had a stimulating effect on C. humicola and B. subtilis growth at a relatively low concentration (10.0 g·mL,1); while, C. humicola and B. subtilis were inhibited completely when the concentrations were up to 320.0 and 160.0 g·mL,1, respectively, and the relationships between growth rate constant (k) and doses of Pb(II) were approximately linear for the two microbes at certain concerntrations. At the same time, their cell dry weight and turbidity (OD600) during growth were also obtained. Their thermogenic curves of the growth coincided well with their turbidity curves, elucidating that the microcalorimetric method agreed with the routine microbiology methods. All of these corroborate the validity and sensitivity of the microcalorimetric technique to investigate the toxic effect of Pb(II) on soil microorganisms. [source] | ||||||||||