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1.
Rosa Garcia Sanchez Bärbel Hahn-Hägerdal Marie F Gorwa-Grauslund 《Microbial cell factories》2010,9(1):40
Background
In Saccharomyces cerevisiae galactose is initially metabolized through the Leloir pathway after which glucose 6-phosphate enters glycolysis. Galactose is controlled both by glucose repression and by galactose induction. The gene PGM2 encodes the last enzyme of the Leloir pathway, phosphoglucomutase 2 (Pgm2p), which catalyses the reversible conversion of glucose 1-phosphate to glucose 6-phosphate. Overexpression of PGM2 has previously been shown to enhance aerobic growth of S. cerevisiae in galactose medium. 相似文献2.
Fermentation performance of engineered and evolved xylose-fermenting Saccharomyces cerevisiae strains 总被引:1,自引:0,他引:1
Sonderegger M Jeppsson M Larsson C Gorwa-Grauslund MF Boles E Olsson L Spencer-Martins I Hahn-Hägerdal B Sauer U 《Biotechnology and bioengineering》2004,87(1):90-98
Lignocellulose hydrolysate is an abundant substrate for bioethanol production. The ideal microorganism for such a fermentation process should combine rapid and efficient conversion of the available carbon sources to ethanol with high tolerance to ethanol and to inhibitory components in the hydrolysate. A particular biological problem are the pentoses, which are not naturally metabolized by the main industrial ethanol producer Saccharomyces cerevisiae. Several recombinant, mutated, and evolved xylose fermenting S. cerevisiae strains have been developed recently. We compare here the fermentation performance and robustness of eight recombinant strains and two evolved populations on glucose/xylose mixtures in defined and lignocellulose hydrolysate-containing medium. Generally, the polyploid industrial strains depleted xylose faster and were more resistant to the hydrolysate than the laboratory strains. The industrial strains accumulated, however, up to 30% more xylitol and therefore produced less ethanol than the haploid strains. The three most attractive strains were the mutated and selected, extremely rapid xylose consumer TMB3400, the evolved C5 strain with the highest achieved ethanol titer, and the engineered industrial F12 strain with by far the highest robustness to the lignocellulosic hydrolysate. 相似文献
3.
Almeida JR Röder A Modig T Laadan B Lidén G Gorwa-Grauslund MF 《Applied microbiology and biotechnology》2008,78(6):939-945
Saccharomyces cerevisiae alcohol dehydrogenases responsible for NADH-, and NADPH-specific reduction of the furaldehydes 5-hydroxymethyl-furfural (HMF)
and furfural have previously been identified. In the present study, strains overexpressing the corresponding genes (mut-ADH1 and ADH6), together with a control strain, were compared in defined medium for anaerobic fermentation of glucose in the presence and
absence of HMF. All strains showed a similar fermentation pattern in the absence of HMF. In the presence of HMF, the strain
overexpressing ADH6 showed the highest HMF reduction rate and the highest specific ethanol productivity, followed by the strain overexpressing
mut-ADH1. This correlated with in vitro HMF reduction capacity observed in the ADH6 overexpressing strain. Acetate and glycerol yields per biomass increased considerably in the ADH6 strain. In the other two
strains, only the overall acetate yield per biomass was affected. When compared in batch fermentation of spruce hydrolysate,
strains overexpressing ADH6 and mut-ADH1 had five times higher HMF uptake rate than the control strain and improved specific ethanol productivity. Overall, our results
demonstrate that (1) the cofactor usage in the HMF reduction affects the product distribution, and (2) increased HMF reduction
activity results in increased specific ethanol productivity in defined mineral medium and in spruce hydrolysate. 相似文献
4.
Nádia Skorupa Parachin Magnus Carlquist Marie F. Gorwa-Grauslund 《Applied microbiology and biotechnology》2009,84(3):487-497
In this study, the production of enantiomerically pure (1R,4S,6S)-6-hydroxy-bicyclo[2.2.2]octane-2-one ((−)-2) through stereoselective bioreduction was used as a model reaction for the comparison of engineered Saccharomyces cerevisiae and engineered Escherichia coli as biocatalysts. For both microorganisms, over-expression of the gene encoding the NADPH-dependent aldo-keto reductase YPR1
resulted in high purity of the keto alcohol (−)-2 (>99% ee, 97–98% de). E. coli had three times higher initial reduction rate but S. cerevisiae continued the reduction reaction for a longer time period, thus reaching a higher conversion of the substrate (95%). S. cerevisiae was also more robust than E. coli, as demonstrated by higher viability during bioreduction. It was also investigated whether the NADPH regeneration rate was
sufficient to supply the over-expressed reductase with NADPH. Five strains of each microorganism with varied carbon flux through
the NADPH regenerating pentose phosphate pathway were genetically constructed and compared. S. cerevisiae required an increased NADPH regeneration rate to supply YPR1 with co-enzyme while the native NADPH regeneration rate was
sufficient for E. coli.
Nádia Skorupa Parachin and Magnus Carlquist have contributed equally to the paper. 相似文献
5.
Anders G. Sandström Henrik Almqvist Diogo Portugal-Nunes Dário Neves Gunnar Lidén Marie F. Gorwa-Grauslund 《Applied microbiology and biotechnology》2014,98(17):7299-7318
Carboxylic acids are important bulk chemicals that can be used as building blocks for the production of polymers, as acidulants, preservatives and flavour compound or as precursors for the synthesis of pharmaceuticals. Today, their production mainly takes place through catalytic processing of petroleum-based precursors. An appealing alternative would be to produce these compounds from renewable resources, using tailor-made microorganisms. Saccharomyces cerevisiae has already demonstrated its value for bioethanol production from renewable resources. In this review, we discuss Saccharomyces cerevisiae engineering potential, current strategies for carboxylic acid production as well as the specific challenges linked to the use of lignocellulosic biomass as carbon source. 相似文献
6.
Emmanuel Assidjo Thierry Chianéa Marie-F. Dreyfuss Philippe J.P. Cardot 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》1998,709(2):9081
Sedimentation field-flow fractionation (SdFFF) offers great potential for the separation of submicrometer and micrometer-sized species. The availability of commercial instrumentation and the versatility of this method originated its success. At this stage of development, SdFFF techniques are mature enough for use in analytical research, development and even routine work. However, prior to their use, these techniques like any other methodologies, have to be validated. As the application of SdFFF techniques to cell separation is being constantly developed, we have investigated separation performance according to validation rules classically defined for separation methods (chromatography) in the case of cellular materials. 相似文献
7.
Jeppsson M Bengtsson O Franke K Lee H Hahn-Hägerdal B Gorwa-Grauslund MF 《Biotechnology and bioengineering》2006,93(4):665-673
Xylose fermentation by Saccharomyces cerevisiae requires the introduction of a xylose pathway, either similar to that found in the natural xylose-utilizing yeasts Pichia stipitis and Candida shehatae or similar to the bacterial pathway. The use of NAD(P)H-dependent XR and NAD(+)-dependent XDH from P. stipitis creates a cofactor imbalance resulting in xylitol formation. The effect of replacing the native P. stipitis XR with a mutated XR with increased K(M) for NADPH was investigated for xylose fermentation to ethanol by recombinant S. cerevisiae strains. Enhanced ethanol yields accompanied by decreased xylitol yields were obtained in strains carrying the mutated XR. Flux analysis showed that strains harboring the mutated XR utilized a larger fraction of NADH for xylose reduction. The overproduction of the mutated XR resulted in an ethanol yield of 0.40 g per gram of sugar and a xylose consumption rate of 0.16 g per gram of biomass per hour in chemostat culture (0.06/h) with 10 g/L glucose and 10 g/L xylose as carbon source. 相似文献
8.
Ted Johanson Ndia Skorupa Parachin Paula Adamis Marie-F. Gorwa-Grauslund 《Journal of Molecular Catalysis .B, Enzymatic》2009,59(4):286-291
Stereoselective baker's yeast-catalysed bioreduction of bicyclo [2.2.2]octane-2,6-dione generates (1R, 4S, 6S)-6-hydroxy-bicyclo [2.2.2]octane-2-one (endo-alcohol) with high enantiomeric and diastereomeric excess. In contrast, whole cells and crude membrane fractions of Candida sp. have been reported to produce the unusual (1R, 4S, 6S)-diastereomer (exo-alcohol) as a major product. Previous in silico screening has identified seven membrane or membrane-bound reductases in C. albicans as candidates for the exo-activity. In this work, purification of the corresponding exo-reductase(s) as well as the heterologous cloning of the seven candidate genes was attempted in C. tropicalis. The overexpression of IPF4033 (AYR1) gene generated an increased exo-to-endo ratio and exo-alcohol production in whole cells and membranes of C. tropicalis. In addition, a slight increased exo-to-endo ratio was observed when overexpressing IPF4033 in S. cerevisiae, although the reduction rate and exo-to-endo ratio were several fold lower compared to those obtained with C. tropicalis. 相似文献
9.
Nádia Skorupa Parachin Jenny Schelin Börje Norling Peter Rådström Marie F. Gorwa-Grauslund 《Applied microbiology and biotechnology》2010,87(5):1927-1933
Nowadays, soil diversity is accessed at molecular level by the total DNA extraction of a given habitat. However, high DNA
yields and purity are difficult to achieve due to the co-extraction of enzyme-inhibitory substances that inhibit downstream
applications, such as PCR, restriction enzyme digestion, and DNA ligation. Therefore, there is a need for further development
of sample preparation methods that efficiently can result in pure DNA with satisfactory yield. In this study, the buoyant
densities of soil microorganisms were utilized to design a sample preparation protocol where microbial cells could be separated
from the soil matrix and enzyme-inhibitory substances by flotation. A discontinuous density gradient was designed using a
colloidal solution of non-toxic silanised silica particles (BactXtractor). The method proved to be an efficient alternative
to direct extraction protocols where cell lysis is performed in the presence of soil particles. The environmental DNA extracted
after flotation had high molecular weight and comparable yield as when using available commercial kits (3.5 μg DNA/g soil),
and neither PCR nor restriction enzyme digestion of DNA were inhibited. Furthermore, specific primers enabled recovery of
both prokaryotic and eukaryotic sequences. 相似文献
10.
Gárdonyi M Jeppsson M Lidén G Gorwa-Grauslund MF Hahn-Hägerdal B 《Biotechnology and bioengineering》2003,82(7):818-824
Saccharomyces cerevisiae TMB3001 has previously been engineered to utilize xylose by integrating the genes coding for xylose reductase (XR) and xylitol dehydrogenase (XDH) and overexpressing the native xylulokinase (XK) gene. The resulting strain is able to metabolize xylose, but its xylose utilization rate is low compared to that of natural xylose utilizing yeasts, like Pichia stipitis or Candida shehatae. One difference between S. cerevisiae and the latter species is that these possess specific xylose transporters, while S. cerevisiae takes up xylose via the high-affinity hexose transporters. For this reason, in part, it has been suggested that xylose transport in S. cerevisiae may limit the xylose utilization.We investigated the control exercised by the transport over the specific xylose utilization rate in two recombinant S. cerevisiae strains, one with low XR activity, TMB3001, and one with high XR activity, TMB3260. The strains were grown in aerobic sugar-limited chemostat and the specific xylose uptake rate was modulated by changing the xylose concentration in the feed, which allowed determination of the flux response coefficients. Separate measurements of xylose transport kinetics allowed determination of the elasticity coefficients of transport with respect to extracellular xylose concentration. The flux control coefficient, C(J) (transp), for the xylose transport was calculated from the response and elasticity coefficients. The value of C(J) (transp) for both strains was found to be < 0.1 at extracellular xylose concentrations > 7.5 g L(-1). However, for strain TMB3260 the flux control coefficient was higher than 0.5 at xylose concentrations < 0.6 g L(-1), while C(J) (transp) stayed below 0.2 for strain TMB3001 irrespective of xylose concentration. 相似文献