| 1. |
- Skorupa-Parachin, Nádia, 1982, et al.
(författare)
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A Microbial Perspective on Ethanolic Lignocellulose Fermentation
- 2011
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Ingår i: Comprehensive Biotechnology, 2nd Edition. - 9780080885049 ; 6, s. 605-614
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Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract
- Bioethanol is an alternative to fossil transportation fuel. It is produced from sugar- and starch-containing crops but current efforts have turned to ethanol from agricultural and forestry waste. These materials are not expected to compete with food and feed production and net emission of carbon dioxide is lower. Several ethanol-producing microorganisms have been assessed at laboratory scale, including Gram-positive and Gram-negative bacteria, eukaryotes such as yeasts and filamentous fungi, but few have so far been used at industrial scale. In this article, the advantages and disadvantages of the different microorganisms including co-cultures are discussed with respect to ethanol production from lignocellulose raw materials. The complexity of lignocellulose materials may require development of different microorganisms for different applications, so that 'tailor-made' strains for different lignocellulose raw materials may be more efficient than one 'super-microorganism' for any raw material.
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| 2. |
- Akerberg, C, et al.
(författare)
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Modelling the influence of pH, temperature, glucose and lactic acid concentrations on the kinetics of lactic acid production by Lactococcus lactis ssp. lactis ATCC 19435 in whole-wheat flour
- 1998
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Ingår i: Applied Microbiology and Biotechnology. - : Springer Science and Business Media LLC. - 1432-0614 .- 0175-7598. ; 49:6, s. 682-690
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Tidskriftsartikel (refereegranskat)abstract
- A kinetic model of the fermentative production of lactic acid from glucose by Lactococcus lactis ssp. lactis ATCC 19435 in whole-wheat flour has been developed. The model consists of terms for substrate and product inhibition as well as for the influence of pH and temperature. Experimental data from fermentation experiments under different physical conditions were used to fit and verify the model. Temperatures above 30 degrees C and pH levels below 6 enhanced the formation of byproducts and D-lactic acid. By-products were formed in the presence of maltose only, whereas D-lactic acid was formed independently of the presence of maltose although the amount formed was greater when maltose was present. The lactic acid productivity was highest between 33 degrees C and 35 degrees C and at pH 6. In the concentration interval studied (up to 180 g l(-1) glucose and 89 g l(-1) lactic acid) simulations showed that both substances were inhibiting. Glucose inhibition was small compared with the inhibition due to lactic acid.
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| 3. |
- Almeida, Joao, et al.
(författare)
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Carbon fluxes of xylose-consuming Saccharomyces cerevisiae strains are affected differently by NADH and NADPH usage in HMF reduction.
- 2009
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Ingår i: Applied Microbiology and Biotechnology. - : Springer Science and Business Media LLC. - 1432-0614 .- 0175-7598. ; 84, s. 751-761
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Tidskriftsartikel (refereegranskat)abstract
- Industrial Saccharomyces cerevisiae strains able to utilize xylose have been constructed by overexpression of XYL1 and XYL2 genes encoding the NADPH-preferring xylose reductase (XR) and the NAD(+)-dependent xylitol dehydrogenase (XDH), respectively, from Pichia stipitis. However, the use of different co-factors by XR and XDH leads to NAD(+) deficiency followed by xylitol excretion and reduced product yield. The furaldehydes 5-hydroxymethyl-furfural (HMF) and furfural inhibit yeast metabolism, prolong the lag phase, and reduce the ethanol productivity. Recently, genes encoding furaldehyde reductases were identified and their overexpression was shown to improve S. cerevisiae growth and fermentation rate in HMF containing media and in lignocellulosic hydrolysate. In the current study, we constructed a xylose-consuming S. cerevisiae strain using the XR/XDH pathway from P. stipitis. Then, the genes encoding the NADH- and the NADPH-dependent HMF reductases, ADH1-S110P-Y295C and ADH6, respectively, were individually overexpressed in this background. The performance of these strains, which differed in their co-factor usage for HMF reduction, was evaluated under anaerobic conditions in batch fermentation in absence or in presence of HMF. In anaerobic continuous culture, carbon fluxes were obtained for simultaneous xylose consumption and HMF reduction. Our results show that the co-factor used for HMF reduction primarily influenced formation of products other than ethanol, and that NADH-dependent HMF reduction influenced product formation more than NADPH-dependent HMF reduction. In particular, NADH-dependent HMF reduction contributed to carbon conservation so that biomass was produced at the expense of xylitol and glycerol formation.
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| 4. |
- Almeida, Joao, et al.
(författare)
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Developing Saccharomyces cerevisiae strains for second generation bioethanol: Improving xylose fermentation and inhibitor tolerance
- 2009
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Ingår i: International Sugar Journal. - 0020-8841. ; 111:1323, s. 172-180
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Tidskriftsartikel (refereegranskat)abstract
- The 2nd generation bioethanol will be produced from lignocellulose biomass including agricultural residues such as bagasse, straw, and stover, forest products residues and dedicated energy crops. The bioconversion of lignocellulose raw materials to ethanol requires microorganisms that are able (i) to ferment both hexose and pentose sugars present in lignocellulose and (ii) to tolerate and to remain active in presence of inhibiting compounds generated during lignocellulose pretreatment and hydrolysis. In this review we focus on the development of strains of baker's yeast Saccharomyces cerevisiae. We discuss genetic and metabolic engineering strategies that have been used to improve, independently, xylose fermentation and strain tolerance. Then, we deliberate strategies for simultaneous improvement of xylose utilization and inhibitor tolerance, with the view that a new generation of xylose-utilizing S. cerevisiae strains harboring beneficial traits for xylose consumption and inhibitor tolerance is expected to be available in the foreseeable future.
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| 5. |
- Almeida, Joao, et al.
(författare)
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Increased tolerance and conversion of inhibitors in lignocellulosic hydrolysates by Saccharomyces cerevisiae
- 2007
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Ingår i: Journal of Chemical Technology and Biotechnology. - : Wiley. - 0268-2575 .- 1097-4660. ; 82:4, s. 340-349
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Forskningsöversikt (refereegranskat)abstract
- During hydrolysis of lignocellulosic biomass, monomeric sugars and a broad range of inhibitory compounds are formed and released. These inhibitors, which can be organized around three main groups, furans, weak acids and phenolics, reduce ethanol yield and productivity by affecting the microorganism performance during the fermentation step. Among the microorganisms that have been evaluated for lignocellulosic hydrolysate ethanol fermentation, the yeast Saccharomyces cerevisiae appears to be the least sensitive. In order to overcome the effect of inhibitors, strategies that include improvement of natural tolerance of microorganism and use of fermentation control strategies have been developed. An overview of the origin, effects and mechanisms of action of known inhibitors on S. cerevisiae is given. Fermentation control strategies as well as metabolic, genetic and evolutionary engineering strategies to obtain S. cerevisiae strains with improved tolerance are discussed.
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| 6. |
- Axelsson, Anders, et al.
(författare)
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Performance of batch and continuous reactors with co-immobilized yeast and beta-galactosidase
- 1991
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Ingår i: Journal of Chemical Technology and Biotechnology. - : Wiley. - 0268-2575 .- 1097-4660. ; 52:2, s. 227-241
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Tidskriftsartikel (refereegranskat)abstract
- The anaerobic fermentation of deproteinized whey with beta-galactosidase coimmobilized with Saccharomyces cerevisiae in calcium alginate gel beads for the production of ethanol has been studied in a continuous horizontal packed bed reactor (HPBR). The results are compared with batch experiments in a stirred tank reactor. The immobilized yeast cells are exposed to conditions that vary with time and location in the reactor, making a true steady state impossible. In spite of a very low specific growth rate-of the order of 0.01 h-1 in the first section of the HPBR-the yeast cell growth, accompanied by bead expansion in this section, was high enough to create a cell concentration gradient along the reactor. The continuous reactor is preferable to the batch reactor as the galactose conversion is more efficient. The highest volumetric productivity obtained in the HPBR was 125 mol ethanol m-3 h-1 (6 g ethanol dm-3 h-1) at a substrate concentration of 164 mol m-3 lactose (56 g dm-3) and a dilution rate of 0.21 h-1, corresponding to a space velocity of 0.51 dm3 dm-3 gel h-1. The ethanol yield from consumed glucose and galactose was 80%. The ethanol yield from lactose was only 70%, as only 75% of the galactose was consumed while all the lactose and glucose were converted.
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| 7. |
- Bengtsson, Oskar, et al.
(författare)
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Xylose reductase from Pichia stipitis with altered coenzyme preference improves ethanolic xylose fermentation by recombinant Saccharomyces cerevisiae
- 2009
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Ingår i: Biotechnology for Biofuels. - : Springer Science and Business Media LLC. - 1754-6834. ; 2
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Tidskriftsartikel (refereegranskat)abstract
- Background: Xylose reductase (XR) and xylitol dehydrogenase (XDH) from Pichia stipitis are the two enzymes most commonly used in recombinant Saccharomyces cerevisiae strains engineered for xylose utilization. The availability of NAD+ for XDH is limited during anaerobic xylose fermentation because of the preference of XR for NADPH. This in turn results in xylitol formation and reduced ethanol yield. The coenzyme preference of P. stipitis XR was changed by site-directed mutagenesis with the aim to engineer it towards NADH-preference. Results: XR variants were evaluated in S. cerevisiae strains with the following genetic modifications: overexpressed native P. stipitis XDH, overexpressed xylulokinase, overexpressed non-oxidative pentose phosphate pathway and deleted GRE3 gene encoding an NADPH dependent aldose reductase. All overexpressed genes were chromosomally integrated to ensure stable expression. Crude extracts of four different strains overexpressing genes encoding native P. stipitis XR, K270M and K270R mutants, as well as Candida parapsilosis XR, were enzymatically characterized. The physiological effects of the mutations were investigated in anaerobic xylose fermentation. The strain overexpressing P. stipitis XR with the K270R mutation gave an ethanol yield of 0.39 g (g consumed sugars)(-1), a xylitol yield of 0.05 g (g consumed xylose)(-1) and a xylose consumption rate of 0.28 g (g biomass)(-1) h(-1) in continuous fermentation at a dilution rate of 0.12 h(-1), with 10 g l(-1) glucose and 10 g l(-1) xylose as carbon sources. Conclusion: The cofactor preference of P. stipitis XR was altered by site-directed mutagenesis. When the K270R XR was combined with a metabolic engineering strategy that ensures high xylose utilization capabilities, a recombinant S. cerevisiae strain was created that provides a unique combination of high xylose consumption rate, high ethanol yield and low xylitol yield during ethanolic xylose fermentation.
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| 8. |
- Bergdahl, Basti, et al.
(författare)
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Dynamic metabolomics differentiates between carbon and energy starvation in recombinant Saccharomyces cerevisiae fermenting xylose
- 2012
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Ingår i: Biotechnology for Biofuels. - : Springer Science and Business Media LLC. - 1754-6834. ; 5:34
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Tidskriftsartikel (refereegranskat)abstract
- Background: The concerted effects of changes in gene expression due to changes in the environment are ultimately reflected in the metabolome. Dynamics of metabolite concentrations under a certain condition can therefore give a description of the cellular state with a high degree of functional information. We used this potential to evaluate the metabolic status of two recombinant strains of Saccharomyces cerevisiae during anaerobic batch fermentation of a glucose/xylose mixture. Two isogenic strains were studied, differing only in the pathways used for xylose assimilation: the oxidoreductive pathway with xylose reductase (XR) and xylitol dehydrogenase (XDH) or the isomerization pathway with xylose isomerase (XI). The isogenic relationship between the two strains ascertains that the observed responses are a result of the particular xylose pathway and not due to unknown changes in regulatory systems. An increased understanding of the physiological state of these strains is important for further development of efficient pentose-utilizing strains for bioethanol production. Results: Using LC-MS/MS we determined the dynamics in the concentrations of intracellular metabolites in central carbon metabolism, nine amino acids, the purine nucleotides and redox cofactors. The general response to the transition from glucose to xylose was increased concentrations of amino acids and TCA-cycle intermediates, and decreased concentrations of sugar phosphates and redox cofactors. The two strains investigated had significantly different uptake rates of xylose which led to an enhanced response in the XI-strain. Despite the difference in xylose uptake rate, the adenylate energy charge remained high and stable around 0.8 in both strains. In contrast to the adenylate pool, large changes were observed in the guanylate pool. Conclusions: The low uptake of xylose by the XI-strain led to several distinguished responses: depletion of key metabolites in glycolysis and NADPH, a reduced GTP/GDP ratio and accumulation of PEP and aromatic amino acids. These changes are strong indicators of carbon starvation. The XR/XDH-strain displayed few such traits. The coexistence of these traits and a stable adenylate charge indicates that xylose supplies energy to the cells but does not suppress a response similar to carbon starvation. Particular signals may play a role in the latter, of which the GTP/GMP ratio could be a candidate as it decreased significantly in both strains.
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| 9. |
- Bettiga, Maurizio, et al.
(författare)
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Arabinose and xylose fermentation by recombinant Saccharomyces cerevisiae expressing a fungal pentose utilization pathway
- 2009
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Ingår i: Microbial Cell Factories. - : Springer Science and Business Media LLC. - 1475-2859. ; 8
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Tidskriftsartikel (refereegranskat)abstract
- Background: Sustainable and economically viable manufacturing of bioethanol from lignocellulose raw material is dependent on the availability of a robust ethanol producing microorganism, able to ferment all sugars present in the feedstock, including the pentose sugars L-arabinose and D-xylose. Saccharomyces cerevisiae is a robust ethanol producer, but needs to be engineered to achieve pentose sugar fermentation. Results: A new recombinant S. cerevisiae strain expressing an improved fungal pathway for the utilization of L-arabinose and D-xylose was constructed and characterized. The new strain grew aerobically on L-arabinose and D-xylose as sole carbon sources. The activities of the enzymes constituting the pentose utilization pathway(s) and product formation during anaerobic mixed sugar fermentation were characterized. Conclusion: Pentose fermenting recombinant S. cerevisiae strains were obtained by the expression of a pentose utilization pathway of entirely fungal origin. During anaerobic fermentation the strain produced biomass and ethanol. L-arabitol yield was 0.48 g per gram of consumed pentose sugar, which is considerably less than previously reported for D-xylose reductase expressing strains co-fermenting L-arabinose and D-xylose, and the xylitol yield was 0.07 g per gram of consumed pentose sugar.
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| 10. |
- Bettiga, Maurizio, et al.
(författare)
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Comparing the xylose reductase/xylitol dehydrogenase and xylose isomerase pathways in arabinose and xylose fermenting Saccharomyces cerevisiae strains
- 2008
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Ingår i: Biotechnology for Biofuels. - : Springer Science and Business Media LLC. - 1754-6834. ; 1
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Tidskriftsartikel (refereegranskat)abstract
- Background: Ethanolic fermentation of lignocellulosic biomass is a sustainable option for the production of bioethanol. This process would greatly benefit from recombinant Saccharomyces cerevisiae strains also able to ferment, besides the hexose sugar fraction, the pentose sugars, arabinose and xylose. Different pathways can be introduced in S. cerevisiae to provide arabinose and xylose utilisation. In this study, the bacterial arabinose isomerase pathway was combined with two different xylose utilisation pathways: the xylose reductase/xylitol dehydrogenase and xylose isomerase pathways, respectively, in genetically identical strains. The strains were compared with respect to aerobic growth in arabinose and xylose batch culture and in anaerobic batch fermentation of a mixture of glucose, arabinose and xylose. Results: The specific aerobic arabinose growth rate was identical, 0.03 h-1, for the xylose reductase/xylitol dehydrogenase and xylose isomerase strain. The xylose reductase/xylitol dehydrogenase strain displayed higher aerobic growth rate on xylose, 0.14 h-1, and higher specific xylose consumption rate in anaerobic batch fermentation, 0.09 g (g cells)-1 h-1 than the xylose isomerase strain, which only reached 0.03 h-1 and 0.02 g (g cells)-1h-1, respectively. Whereas the xylose reductase/xylitol dehydrogenase strain produced higher ethanol yield on total sugars, 0.23 g g-1 compared with 0.18 g g-1 for the xylose isomerase strain, the xylose isomerase strain achieved higher ethanol yield on consumed sugars, 0.41 g g-1 compared with 0.32 g g-1 for the xylose reductase/xylitol dehydrogenase strain. Anaerobic fermentation of a mixture of glucose, arabinose and xylose resulted in higher final ethanol concentration, 14.7 g l-1 for the xylose reductase/ xylitol dehydrogenase strain compared with 11.8 g l-1 for the xylose isomerase strain, and in higher specific ethanol productivity, 0.024 g (g cells)-1 h-1 compared with 0.01 g (g cells)-1 h-1 for the xylose reductase/ xylitol dehydrogenase strain and the xylose isomerase strain, respectively. Conclusion: The combination of the xylose reductase/xylitol dehydrogenase pathway and the bacterial arabinose isomerase pathway resulted in both higher pentose sugar uptake and higher overall ethanol production than the combination of the xylose isomerase pathway and the bacterial arabinose isomerase pathway. Moreover, the flux through the bacterial arabinose pathway did not increase when combined with the xylose isomerase pathway. This suggests that the low activity of the bacterial arabinose pathway cannot be ascribed to arabitol formation via the xylose reductase enzyme.
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