Bärbel Hahn-Hägerdal

Lund University, Lund, Skåne, Sweden

Are you Bärbel Hahn-Hägerdal?

Claim your profile

Publications (243)726.3 Total impact

  • Source
    Basti Bergdahl · Dominik Heer · Uwe Sauer · Bärbel Hahn-Hägerdal · Ed Wj van Niel
    [Show abstract] [Hide abstract]
    ABSTRACT: 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. 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. 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.
    Full-text · Article · May 2012 · Biotechnology for Biofuels
  • Michael J. O’Donohue · Bärbel Hahn-Hägerdal

    No preview · Article · Mar 2012 · PROCESS BIOCHEMISTRY
  • N.S. Parachin · B. Hahn-Hägerdal · M. Bettiga
    [Show abstract] [Hide abstract]
    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.
    No preview · Chapter · Dec 2011
  • [Show abstract] [Hide abstract]
    ABSTRACT: Ethanolic fermentation of lignocellulose raw materials requires industrial xylose-fermenting strains capable of complete and efficient D-xylose consumption. A central question in xylose fermentation by Saccharomyces cerevisiae engineered for xylose fermentation is to improve the xylose uptake. In the current study, the glucose/xylose facilitator Gxf1 from Candida intermedia, was expressed in three different xylose-fermenting S. cerevisiae strains of industrial origin. The in vivo effect on aerobic xylose growth and the initial xylose uptake rate were assessed. The expression of Gxf1 resulted in enhanced aerobic xylose growth only for the TMB3400 based strain. It displayed more than a 2-fold higher affinity for D-xylose than the parental strain and approximately 2-fold higher initial specific growth rate at 4 g/L D-xylose. Enhanced xylose consumption was furthermore observed when the GXF1-strain was assessed in simultaneous saccharification and co-fermentation (SSCF) of pretreated wheat straw. However, the ethanol yield remained unchanged due to increased by-product formation. Metabolic flux analysis suggested that the expression of the Gxf1 transporter had shifted the control of xylose catabolism from transport to the NAD(+) dependent oxidation of xylitol to xylulose.
    No preview · Article · May 2011
  • Source
    Kim Olofsson · David Runquist · Bärbel Hahn-Hägerdal · Gunnar Lidén
    [Show abstract] [Hide abstract]
    ABSTRACT: Genetically engineered Saccharomyces cerevisiae strains are able to ferment xylose present in lignocellulosic biomass. However, better xylose fermenting strains are required to reach complete xylose uptake in simultaneous saccharification and co-fermentation (SSCF) of lignocellulosic hydrolyzates. In the current study, haploid Saccharomyces cerevisiae strains expressing a heterologous xylose pathway including either the native xylose reductase (XR) from P. stipitis, a mutated variant of XR (mXR) with altered co-factor preference, a glucose/xylose facilitator (Gxf1) from Candida intermedia or both mXR and Gxf1 were assessed in SSCF of acid-pretreated non-detoxified wheat straw. The xylose conversion in SSCF was doubled with the S. cerevisiae strain expressing mXR compared to the isogenic strain expressing the native XR, converting 76% and 38%, respectively. The xylitol yield was less than half using mXR in comparison with the native variant. As a result of this, the ethanol yield increased from 0.33 to 0.39 g g-1 when the native XR was replaced by mXR. In contrast, the expression of Gxf1 only slightly increased the xylose uptake, and did not increase the ethanol production. The results suggest that ethanolic xylose fermentation under SSCF conditions is controlled primarily by the XR activity and to a much lesser extent by xylose transport.
    Full-text · Article · Mar 2011 · AMB Express
  • B. Hahn-Hägerdal · F. Gorwa-Grauslund

    No preview · Article · Nov 2010 · Journal of Biotechnology
  • Source
    D. Runquist · B. Hahn-Hägerdal · M. Bettiga

    Full-text · Article · Nov 2010 · Journal of Biotechnology
  • Source
    David Runquist · Bärbel Hahn-Hägerdal · Maurizio Bettiga
    [Show abstract] [Hide abstract]
    ABSTRACT: Baker's yeast (Saccharomyces cerevisiae) has been genetically engineered to ferment the pentose sugar xylose present in lignocellulose biomass. One of the reactions controlling the rate of xylose utilization is catalyzed by xylose reductase (XR). In particular, the cofactor specificity of XR is not optimized with respect to the downstream pathway, and the reaction rate is insufficient for high xylose utilization in S. cerevisiae. The current study describes a novel approach to improve XR for ethanol production in S. cerevisiae. The cofactor binding region of XR was mutated by error-prone PCR, and the resulting library was expressed in S. cerevisiae. The S. cerevisiae library expressing the mutant XR was selected in sequential anaerobic batch cultivation. At the end of the selection process, a strain (TMB 3420) harboring the XR mutations N272D and P275Q was enriched from the library. The Vmax of the mutated enzyme was increased by an order of magnitude compared to that of the native enzyme, and the NADH/NADPH utilization ratio was increased significantly. The ethanol productivity from xylose in TMB 3420 was increased ∼40 times compared to that of the parent strain (0.32 g/g [dry weight {DW}] × h versus 0.007 g/g [DW] × h), and the anaerobic growth rate was increased from ∼0 h−1 to 0.08 h−1. The improved traits of TMB 3420 were readily transferred to the parent strain by reverse engineering of the mutated XR gene. Since integrative vectors were employed in the construction of the library, transfer of the improved phenotype does not require multicopy expression from episomal plasmids.
    Full-text · Article · Oct 2010 · Applied and Environmental Microbiology

  • No preview · Article · Sep 2010 · ChemInform
  • Source
    Rosa Garcia Sanchez · Bärbel Hahn-Hägerdal · Marie F Gorwa-Grauslund
    [Show abstract] [Hide abstract]
    ABSTRACT: Overexpression of the PGM2 gene encoding phosphoglucomutase (Pgm2p) has been shown to improve galactose utilization both under aerobic and under anaerobic conditions. Similarly, xylose utilization has been improved by overexpression of genes encoding xylulokinase (XK), enzymes from the non-oxidative pentose phosphate pathway (non-ox PPP) and deletion of the endogenous aldose reductase GRE3 gene in engineered Saccharomyces cerevisiae strains carrying either fungal or bacterial xylose pathways. In the present study, we investigated how the combination of these traits affect xylose and galactose utilization in the presence or absence of glucose in S. cerevisiae strains engineered with the xylose reductase (XR)-xylitol dehydrogenase (XDH) pathway. In the absence of PGM2 overexpression, the combined overexpression of XK, the non-ox PPP and deletion of the GRE3 gene significantly delayed aerobic growth on galactose, whereas no difference was observed between the control strain and the xylose-engineered strain when the PGM2 gene was overexpressed. Under anaerobic conditions, the overexpression of the PGM2 gene increased the ethanol yield and the xylose consumption rate in medium containing xylose as the only carbon source. The possibility of Pgm2p acting as a xylose isomerase (XI) could be excluded by measuring the XI activity in both strains. The additional copy of the PGM2 gene also resulted in a shorter fermentation time during the co-consumption of galactose and xylose. However, the effect was lost upon addition of glucose to the growth medium. PGM2 overexpression was shown to benefit xylose and galactose fermentation, alone and in combination. In contrast, galactose fermentation was impaired in the engineered xylose-utilizing strain harbouring extra copies of the non-ox PPP genes and a deletion of the GRE3 gene, unless PGM2 was overexpressed. These cross-reactions are of particular relevance for the fermentation of mixed sugars from lignocellulosic feedstock.
    Full-text · Article · Sep 2010 · Biotechnology for Biofuels
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: In a recent study combining transcriptome analyses of a number of recombinant laboratory and industrial S. cerevisiae strains with improved xylose utilization and their respective control strains, the ORF YLR042c was identified as a downregulated gene and it was shown that the gene deletion improved aerobic growth on xylose in the tested strain background. In the present study, the influence of deleting YLR042c on xylose fermentation was investigated in two different xylose-fermenting strains: TMB3001, which expresses genes from the initial xylose catabolizing pathway, including heterologous xylose reductase (XR) and xylitol dehydrogenase (XDH) and endogenous xylulokinase (XK); and TMB3057, which, in addition to the initial xylose catabolizing pathway, overexpresses the endogenous genes encoding the non-oxidative pentose phosphate pathway enzymes. The deletion of YLR042c led to improved aerobic growth on xylose in both strain backgrounds. However, the effect was more significant in the strain with the poorer growth rate on xylose (TMB3001). Under anaerobic conditions, the deletion of YLR042c increased the specific xylose consumption rate and the ethanol and xylitol yields. In strain TMB3057, xylose consumption was also improved at low concentrations and during co-fermentation of xylose and glucose. The effect of the gene deletion and overexpression was also tested for different carbon sources. Altogether, these results suggest that YLR042c influences xylose and the assimilation of carbon sources other than glucose, and that the effect could be at the level of sugar transport or sugar signalling.
    Full-text · Article · Sep 2010 · Yeast
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Cost-effective fermentation of lignocellulosic hydrolysate to ethanol by Saccharomyces cerevisiae requires efficient mixed sugar utilization. Notably, the rate and yield of xylose and arabinose co-fermentation to ethanol must be enhanced. Evolutionary engineering was used to improve the simultaneous conversion of xylose and arabinose to ethanol in a recombinant industrial Saccharomyces cerevisiae strain carrying the heterologous genes for xylose and arabinose utilization pathways integrated in the genome. The evolved strain TMB3130 displayed an increased consumption rate of xylose and arabinose under aerobic and anaerobic conditions. Improved anaerobic ethanol production was achieved at the expense of xylitol and glycerol but arabinose was almost stoichiometrically converted to arabitol. Further characterization of the strain indicated that the selection pressure during prolonged continuous culture in xylose and arabinose medium resulted in the improved transport of xylose and arabinose as well as increased levels of the enzymes from the introduced fungal xylose pathway. No mutation was found in any of the genes from the pentose converting pathways. To the best of our knowledge, this is the first report that characterizes the molecular mechanisms for improved mixed-pentose utilization obtained by evolutionary engineering of a recombinant S. cerevisiae strain. Increased transport of pentoses and increased activities of xylose converting enzymes contributed to the improved phenotype.
    Full-text · Article · Jun 2010 · Biotechnology for Biofuels
  • Source
    Rosa Garcia Sanchez · Bärbel Hahn-Hägerdal · Marie F Gorwa-Grauslund
    [Show abstract] [Hide abstract]
    ABSTRACT: 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. In the present study we show that overexpression of PGM2 under control of the HXT7'promoter from an integrative plasmid increased the PGM activity 5 to 6 times, which significantly reduced the lag phase of glucose-pregrown cells in an anaerobic galactose culture. PGM2 overexpression also increased the anaerobic specific growth rate whereas ethanol production was less influenced. When PGM2 was overexpressed from a multicopy plasmid instead, the PGM activity increased almost 32 times. However, this increase of PGM activity did not further improve aerobic galactose fermentation as compared to the strain carrying PGM2 on the integrative plasmid. PGM2 overexpression in S. cerevisiae from an integrative plasmid is sufficient to reduce the lag phase and to enhance the growth rate in anaerobic galactose fermentation, which results in an overall decrease in fermentation duration. This observation is of particular importance for the future development of stable industrial strains with enhanced PGM activity.
    Full-text · Article · May 2010 · Microbial Cell Factories
  • D. Runquist · N.S. Parachin · B. Hahn-Hägerdal
    [Show abstract] [Hide abstract]
    ABSTRACT: Lignocellulosic biomass presents an attractive but challenging substrate for bioethanol production. Compared to starch and sucrose based processes, utilization of lignocellulose is complicated by more elaborate pretreatment of the biomass and the presence of toxic inhibitory compounds. In addition, lignocellulosic biomass is composed of several types of sugar polymers, which gives rise to a complex blend of hexose and pentose monomer sugars in the feed medium. Brewer's yeast, Saccharomyces cerevisiae, has been metabolically engineered to utilize the pentose sugars xylose and arabinose; however, successful co-fermentation of all sugars derived from the lignocellulosic biomass is even less straightforward. Mixed substrate fermentation may lead to sequential utilization of sugars, increased byproduct formation and longer overall fermentation times. In this chapter, challenges relating specifically to co-fermentation of lignocelluloses-derived sugars are discussed. Preferential utilization of certain sugars will be analyzed from the perspective of transport and transcriptional control. Practical solutions to improve co-fermentation of lignocelluloses-derived sugars are discussed and suggestions for further improvements are given.
    No preview · Article · May 2010
  • Source
    David Runquist · Bärbel Hahn-Hägerdal · Peter Rådström
    [Show abstract] [Hide abstract]
    ABSTRACT: Baker's yeast (Saccharomyces cerevisiae) has been engineered for xylose utilization to enable production of fuel ethanol from lignocellulose raw material. One unresolved challenge is that S. cerevisiae lacks a dedicated transport system for pentose sugars, which means that xylose is transported by non-specific Hxt transporters with comparatively low transport rate and affinity for xylose. In this study, we compared three heterologous xylose transporters that have recently been shown to improve xylose uptake under different experimental conditions. The transporters Gxf1, Sut1 and At5g59250 from Candida intermedia, Pichia stipitis and Arabidopsis thaliana, respectively, were expressed in isogenic strains of S. cerevisiae and the transport kinetics and utilization of xylose was evaluated. Expression of the Gxf1 and Sut1 transporters led to significantly increased affinity and transport rates of xylose. In batch cultivation at 4 g/L xylose concentration, improved transport kinetics led to a corresponding increase in xylose utilization, whereas no correlation could be demonstrated at xylose concentrations greater than 15 g/L. The relative contribution of native sugar transporters to the overall xylose transport capacity was also estimated during growth on glucose and xylose. Kinetic characterization and aerobic batch cultivation of strains expressing the Gxf1, Sut1 and At5g59250 transporters showed a direct relationship between transport kinetics and xylose growth. The Gxf1 transporter had the highest transport capacity and the highest xylose growth rate, followed by the Sut1 transporter. The range in which transport controlled the growth rate was determined to between 0 and 15 g/L xylose. The role of catabolite repression in regulation of native transporters was also confirmed by the observation that xylose transport by native S. cerevisiae transporters increased significantly during cultivation in xylose and at low glucose concentration.
    Full-text · Article · Mar 2010 · Biotechnology for Biofuels
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Sugarcane bagasse is a lignocellulosic residue obtained from sugarcane milling, and a potentially interesting raw material that can be used for fuel ethanol production. In the present study, bagasse was steam pretreated at temperatures between 180 and 205 °C, with holding times of 5–10 min using SO2 as a catalyst to determine conditions that provide a good recovery of pentoses and a suitable material for enzymatic hydrolysis. Pretreatment conducted at 190 °C for 5 min gave a pentose yield of 57%, with only minor amounts of degradation compounds formed. Commercial cellulolytic enzymes were used to hydrolyze the obtained fiber fractions after pretreatment at different water-insoluble solid contents (2%, 5% and 8% WIS). The overall highest sugar yield achieved from bagasse was 87% at 2% WIS. Fermentation tests were made on both the pentose-rich hemicellulose hydrolysate obtained from the pretreatment, and the enzymatic hydrolysates obtained from the fiber fractions using the xylose-fermenting strain of Saccharomyces cerevisiae TMB3400, as well as the natural xylose-utilizing yeast Pichia stipitis CBS 6054. The pretreatment hydrolysates produced at 2% WIS as well as the enzymatic hydrolysates showed a complete glucose fermentability indicating a low toxicity to the yeasts. The best xylose conversion (more than 60%) was achieved by the strain TMB3400 at 2% WIS.
    Full-text · Article · Feb 2010 · Enzyme and Microbial Technology
  • Source
    Grant Stanley · Bärbel Hahn-Hägerdal

    Preview · Chapter · Jan 2010
  • Source
    David Runquist · Bärbel Hahn-Hägerdal · Maurizio Bettiga
    [Show abstract] [Hide abstract]
    ABSTRACT: Fermentation of xylose to ethanol has been achieved in S. cerevisiae by genetic engineering. Xylose utilization is however slow compared to glucose, and during anaerobic conditions addition of glucose has been necessary for cellular growth. In the current study, the xylose-utilizing strain TMB 3415 was employed to investigate differences between anaerobic utilization of glucose and xylose. This strain carried a xylose reductase (XYL1 K270R) engineered for increased NADH utilization and was capable of sustained anaerobic growth on xylose as sole carbon source. Metabolic and transcriptional characterization could thus for the first time be performed without addition of a co-substrate or oxygen. Analysis of metabolic fluxes showed that although the specific ethanol productivity was an order of magnitude lower on xylose than on glucose, product yields were similar for the two substrates. In addition, transcription analysis identified clear regulatory differences between glucose and xylose. Respiro-fermentative metabolism on glucose during aerobic conditions caused repression of cellular respiration, while metabolism on xylose under the same conditions was fully respiratory. During anaerobic conditions, xylose repressed respiratory pathways, although notably more weakly than glucose. It was also observed that anaerobic xylose growth caused up-regulation of the oxidative pentose phosphate pathway and gluconeogenesis, which may be driven by an increased demand for NADPH during anaerobic xylose catabolism. Co-factor imbalance in the initial two steps of xylose utilization may reduce ethanol productivity by increasing the need for NADP+ reduction and consequently increase reverse flux in glycolysis.
    Full-text · Article · Sep 2009 · Microbial Cell Factories
  • Ana Maria Souto-Maior · David Runquist · Bärbel Hahn-Hägerdal
    [Show abstract] [Hide abstract]
    ABSTRACT: For recombinant xylose-utilizing Saccharomyces cerevisiae, ethanol yield and productivity is substantially lower on xylose than on glucose. In contrast to glucose, xylose is a novel substrate for S. cerevisiae and it is not known how this substrate is recognized on a molecular level. Failure to activate appropriate genes during xylose-utilization has the potential to result in sub-optimal metabolism and decreased substrate uptake. Certain differences in fermentative performance between the two substrates have thus been ascribed to variations in regulatory response. In this study differences in substrate utilization of glucose and xylose was analyzed in the recombinant S. cerevisiae strain TMB3400. Continuous cultures were performed with glucose and xylose under carbon- and nitrogen-limited conditions. Whereas biomass yield and substrate uptake rate were similar during carbon-limited conditions, the metabolic profile was highly substrate dependent under nitrogen-limited conditions. While glycerol production occurred in both cases, ethanol production was only observed for glucose cultures. Addition of acetate and 2-deoxyglucose pulses to a xylose-limited culture was able to stimulate transient overflow metabolism and ethanol production. Application of glucose pulses enhanced xylose uptake rate under restricted co-substrate concentrations. Results are discussed in relation to regulation of sugar metabolism in Crabtree-positive and -negative yeast.
    No preview · Article · Aug 2009 · Journal of Biotechnology
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Xylose fermentation in yeast has been a target of research for years, yet not all the factors that may affect xylose fermentation performance of yeast strains are known. In this study, the mutant S. cerevisiae strain TMB 3400, which has good xylose fermentation properties, was compared with its parental strain to examine the factors behind the improved xylose utilization at protein level. The proteome of the parental and the mutant strains were characterized by difference in gel electrophoresis (DiGE) to quantitatively identify proteins that are expressed at altered levels in the mutant. The most significant changes detected by proteome analysis were the 6-10-fold increased levels of xylose reductase, xylitol dehydrogenase and transketolase (Tkl1) in the mutant, which is in accordance with previous knowledge about xylose metabolism in yeast. The level of acetaldehyde dehydrogenase (Ald6) was also significantly increased. In addition, several proteins homologous to proteins from yeast species other than S. cerevisiae were identified in both strains, demonstrating the genetic heterogeneity of industrial yeast strains. The results were also compared with a previously reported transcription analysis performed with identical experimental set-up; however, very little correlation between the two datasets was observed. The results of the proteome analysis were in good agreement with a parallel study in which rationally designed overexpression of XR, XDH and the non-oxidative pentose phosphate pathway resulted in similar improvement in xylose utilization, which demonstrates the usefulness of proteome analysis for the identification of target genes for further metabolic engineering strategies in industrial yeast strains.
    Full-text · Article · Jul 2009 · Yeast

Publication Stats

16k Citations
726.30 Total Impact Points


  • 1981-2012
    • Lund University
      • • Division of Applied Microbiology
      • • Department of Analytical Chemistry
      • • Department of Chemistry
      Lund, Skåne, Sweden
  • 2006
    • Goethe-Universität Frankfurt am Main
      • Institute of Molecular Biosciences
      Frankfurt, Hesse, Germany
  • 2005
    • Stellenbosch University
      • Department of Microbiology
      Stellenbosch, Western Cape, South Africa
  • 2002
    • University of the Free State
      • Division of Microbiology and Biochemistry
      Bloemfontein, Orange Free State, South Africa
    • Heinrich-Heine-Universität Düsseldorf
      • Institute of Microbiology
      Düsseldorf, North Rhine-Westphalia, Germany
  • 1989-1995
    • The University of Calgary
      • Department of Biological Sciences
      Calgary, Alberta, Canada