Change search
Refine search result
12 1 - 50 of 91
CiteExportLink to result list
Permanent link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Rows per page
  • 5
  • 10
  • 20
  • 50
  • 100
  • 250
Sort
  • Standard (Relevance)
  • Author A-Ö
  • Author Ö-A
  • Title A-Ö
  • Title Ö-A
  • Publication type A-Ö
  • Publication type Ö-A
  • Issued (Oldest first)
  • Issued (Newest first)
  • Created (Oldest first)
  • Created (Newest first)
  • Last updated (Oldest first)
  • Last updated (Newest first)
  • Standard (Relevance)
  • Author A-Ö
  • Author Ö-A
  • Title A-Ö
  • Title Ö-A
  • Publication type A-Ö
  • Publication type Ö-A
  • Issued (Oldest first)
  • Issued (Newest first)
  • Created (Oldest first)
  • Created (Newest first)
  • Last updated (Oldest first)
  • Last updated (Newest first)
Select
The maximal number of hits you can export is 250. When you want to export more records please use the 'Create feeds' function.
  • 1.
    Agnhage, Tove
    et al.
    University of Borås, Swedish School of Textiles.
    Nierstrasz, Vincent
    University of Borås, Swedish School of Textiles.
    Perwuelz, A.
    Guan, J.P.
    Chen, G.Q.
    Eco-design innovative methods for fabric finishing2014Conference paper (Other academic)
  • 2. Allas, Ular
    et al.
    Toom, Lauri
    Selyutina, Anastasia
    Maeorg, Uno
    Medina, Ricardo
    Merits, Andres
    Rinken, Ago
    Hauryliuk, Vasili
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). University of Tartu, Institute of Technology, Nooruse 1, Tartu 50411, Estonia.
    Kaldalu, Niilo
    Tenson, Tanel
    Antibacterial activity of the nitrovinylfuran G1 (Furvina) and its conversion products2016In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, 36844Article in journal (Refereed)
    Abstract [en]

    2-Bromo-5-(2-bromo-2-nitrovinyl) furan (G1 or Furvina) is an antimicrobial with a direct reactivity against thiol groups. It is active against Gram-positive and Gram-negative bacteria, yeasts and filamentous fungi. By reacting with thiol groups it causes direct damage to proteins but, as a result, is very short-living and interconverts into an array of reaction products. Our aim was to characterize thiol reactivity of G1 and its conversion products and establish how much of antimicrobial and cytotoxic effects are due to the primary activity of G1 and how much can be attributed to its reaction products. Stability of G1 in growth media as well as its conversion in the presence of thiols was characterized. The structures of G1 decomposition products were determined using NMR and mass-spectroscopy. Concentration-and time-dependent killing curves showed that G1 is bacteriostatic for Escherichia coli at the concentration of 16 mu g/ml and bactericidal at 32 mu g/ml. However, G1 is inefficient against non-growing E. coli. Addition of cysteine to medium reduces the antimicrobial potency of G1. Nevertheless, the reaction products of G1 and cysteine enabled prolonged antimicrobial action of the drug. Therefore, the activity of 2-bromo-5-(2-bromo-2-nitrovinyl) furan is a sum of its immediate reactivity and the antibacterial effects of the conversion products.

  • 3.
    Berglund, P.
    et al.
    University of Toronto, Canada.
    Stabile, M. R.
    Gold, M.
    Jones, J. B.
    Mitchinson, C.
    Bott, R. R.
    Graycar, T. P.
    Altering the specificity of subtilisin B. lentus by combining site-directed mutagenesis and chemical modification1996In: Bioorganic and Medicinal Chemistry Letters, ISSN 0960894X (ISSN), Vol. 6, no 21, 2507-2512 p.Article in journal (Refereed)
    Abstract [en]

    The thiol side chain of the M222C mutant of the subtilisin from Bacillus lentus (SBL) has been chemically modified by methyl-, aminoethyl-, and sulfonatoethylthiosulfonate reagents. Introduction of charged residues into the active site of the enzyme reduced the catalytic efficiency with Suc-AAPF-pNA as the substrate, but resulted in better binding of sterically demanding boronic acid inhibitors.

  • 4.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    BIO-AMINES: Searching for a Novel Approach to Biocatalytic Transaminations – a Vinnova Sponsored Project2009In: Book of abstracts, 2009Conference paper (Other academic)
  • 5.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    Biocatalysis with Promiscuous Enzymes2007In: 2007 European BioPerspectives / [ed] Dechema, 2007Conference paper (Refereed)
  • 6.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    Enzyme Catalytic Promiscuity and Rational Design2006In: Book of abstracts, 2006Conference paper (Refereed)
  • 7.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    omega-Transaminases: Tailored for Chiral Amine Synthesis2010In: Biocat2010 / [ed] Ralf Grote, Garabed Antranikian, Hamburg, Germany: TuTech Verlag , 2010Conference paper (Refereed)
  • 8.
    Berglund, Per
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Christiernin, M.
    Hedenström, E.
    Enantiorecognition of chiral acids by Candida rugosa lipase: Two substrate binding modes evidenced in an organic medium2001In: American Chemical Society Symposium Series (ACS), ISSN 0097-6156, E-ISSN 1947-5918, Vol. 776, 263-273 p.Article in journal (Refereed)
    Abstract [en]

    We have identified the existence of different modes of binding the enantiomers of 2-methyl-branched carboxylic acids to a lipase active site by rational substrate engineering. Similar to hydrolysis, previously investigated, we have now evidence for differential binding modes in the Candida rugosa lipase-catalyzed esterifications in cyclohexane. The relevance of considering two different binding modes to understand lipase enantiorecognition is demonstrated by introducing bulky substituents on a chiral carboxylic acid which impose a different orientation of the substrate acyl chain in the active site of Candida rugosa lipase. With this substrate engineering approach based on molecular modeling it is thus possible to markedly alter the enantioselectivity of the lipase. Examples from hydrolysis and new results from esterifications in an organic solvent are presented and discussed.

  • 9.
    Berglund, Per
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Hedenström, Erik
    Mid Sweden university.
    Preparation of 2-, 3-, and 4-methylcarboxylic acids and the corresponding alcohols of high enantiopurity by lipase-catalyzed esterification2001In: Enzymes in Nonaqueous Solvents: Methods and Protocols / [ed] Vulfson, E. N.; Halling, P. J.; Holland, H. L., Totowa: Humana Press , 2001, 307-317 p.Chapter in book (Refereed)
  • 10.
    Berglund, Per
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Holmquist, M.
    Hult, K.
    Reversed enantiopreference of Candida rugosa lipase supports different modes of binding enantiomers of a chiral acyl donor1998In: Journal of Molecular Catalysis - B Enzymatic, ISSN 13811177 (ISSN), Vol. 5, no 1-4, 283-287 p.Article in journal (Refereed)
    Abstract [en]

    Molecular modelling identifies two different productive modes of binding the enantiomers of a 2-methyldecanoic acid ester to the active site of Candida rugosa lipase (CRL). The fast reacting S-enantiomer occupies the previously identified acyl-binding tunnel of the enzyme, whereas the R- enantiomer leaves the tunnel empty. The modelling suggested that if both enantiomers were forced to bind to the active site leaving the tunnel empty, the enzyme would reverse its enantiopreference to become R-enantioselective. To test this hypothesis, we designed a structural analogue to 2- methyldecanoic acid, 2-methyl-6-(2-thienyl)hexanoic acid, which was expected to be too bulky to fit its acyl moiety into the acyl-binding tunnel. The CRL- catalysed hydrolysis of the ethyl ester of this substrate resulted in the preferential conversion of the R-enantiomer as predicted by molecular modelling. This represents the first kinetic evidence supporting the existence of two different modes of binding the enantiomers of a 2- methyldecanoic acid ester to the active site of CRL. We have shown that a rational 3D based approach in combination with substrate engineering can be used to predict and control the stereochemical outcome of a lipase catalysed reaction.

  • 11.
    Berglund, Per
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Hult, Karl
    KTH, School of Biotechnology (BIO), Biochemistry.
    Biocatalytic synthesis of enantiopure compounds using lipases: Chapter 212000In: Stereoselective Biocatalysis / [ed] Patel, R. N., New York: Marcel Dekker, 2000, 633-657 p.Chapter in book (Refereed)
  • 12.
    Berglund, Per
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Svedendahl, Maria
    KTH, School of Biotechnology (BIO), Biochemistry.
    Engelmark Cassimjee, Karim
    KTH, School of Biotechnology (BIO), Biochemistry.
    Branneby, Cecilia
    Cambrex Karlskoga AB.
    Abedi, Vahak
    AstraZeneca.
    Wells, Andrew
    AstraZeneca.
    Federsel, Hans-Jürgen
    AstraZeneca.
    Omega-Transaminases Redesigned for Chiral Amine Synthesis2011In: BIT Life Sciences’ 2nd Symposium on Enzymes & Biocatalysis, Dalian, China: BIT Life Sciences , 2011Conference paper (Refereed)
  • 13.
    Berglund, Per
    et al.
    Mid Sweden University.
    Vörde, Carin
    Hogberg, Hans-Erik
    Esterification of 2-methylalkanoic acids Catalysed by Lipase from Candida rugosa: Enantioselectivity as a Function of water Activity and Alcohol Chain Length1994In: Biocatalysis and Biotransformation, ISSN 1024-2422, E-ISSN 1029-2446, Vol. 9, no 1-4, 123-130 p.Article in journal (Refereed)
  • 14.
    Bi, Ran
    KTH, School of Chemical Science and Engineering (CHE), Fibre and Polymer Technology, Wood Chemistry and Pulp Technology. KTH, School of Chemical Science and Engineering (CHE), Centres, Wallenberg Wood Science Center.
    Lignocellulose Degradation by Soil Micro-organisms2016Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Lignocellulosic biomass is a sustainable resource with abundant reserves. Compared to petroleum ‐ based products, the biomass ‐ derived polymers and chemicals give better environmental profiles. A lot of research interest is focused on understanding the lignocellulose structures.

    Lignin, among the three major wood components, represents most difficulty for microbial degradation because of its complex structure and because cross ‐ linking to hemicellulose makes wood such a compact structure. Nevertheless, wood is naturally degraded by wood ‐ degrading micro ‐ organisms and modified and partly degraded residual of lignin goes into soil. Therefore soil serves as a good environment in which to search for special lignin ‐ degraders. In this thesis, different types of lignin have been used as sole carbon sources to screen for lignin ‐ degrading soil micro ‐ organisms. Eleven aerobic and three anaerobic microbe strains have been isolated and identified as able to grow on lignin. The lignin degradation patterns of selected strains have been studied and these partly include an endwise cleavage of  β‐ O ‐ 4 bonds in lignin and is more complex than simple hydrolytic degradation.

    As lignin exists in wood covalently bonded to hemicellulose, one isolated microbe strain, Phoma herbarum, has also been studied with regards to its ability to degrade covalent lignin polysaccharide networks (LCC). The results show that its culture filtrate can attack lignin ‐ polysaccharide networks in a manner different from that of the commercial enzyme product, Gammanase, possibly by selective cleavage of phenyl glucoside bonds. The effects on LCC of Phoma herbarum also enhance polymer extractability. Hot ‐ water extraction of a culture filtrate of Phoma herbarum ‐ treated fiberized spruce wood material gave an amount of extracted galactoglucomannan more than that given by the Gammanase ‐ treated material and non ‐ enzyme ‐ treated material.

    Over millions of years of natural evolution, micro ‐ organisms on the one hand develop so that they can degrade all wood components to get energy for growth, while plants on the other hand also continuously develop to defend from microbial attack. Compared with lignin and cellulose, hemicelluloses as major components of plant cell walls, are much more easily degraded, but hemicelluloses differ from cellulose in that they are acetylated to different extents. The biological functions of acetylation are not completely understood, but it is suggested is that one function is to decrease the microbial degradability of cell walls. By cultivation of soil micro ‐ organisms using mannans acetylated to deffernent degrees as sole carbon source on agar plates, we were able to see significant trends where the resistance towards microbial degradation of glucomannan and galactomannan increased with increasing degree of acetylation. Possible mechanisms and the technological significance of this are discussed. Tailoring the degree of acetylation of polysaccharide materials might slow down the biodegradation, making it possible to design a material with a degradation rate suited to its application.

  • 15. Billig, S.
    et al.
    Agrawal, P.B.
    Birkemeyer, C.
    Nierstrasz, Vincent
    University of Borås, Swedish School of Textiles.
    Warmoeskerken, M.M.C.G.
    Zimmerman, W.
    Biodegradation of diverse PET materials by polyester hydrolases from Thermobifida fusca and Fusarium solani2014Conference paper (Refereed)
  • 16.
    Blissing, Annica
    Linköping University, Department of Physics, Chemistry and Biology, Chemistry. Linköping University, Faculty of Science & Engineering.
    Thiopurine S-methyltransferase - characterization of variants and ligand binding2017Licentiate thesis, comprehensive summary (Other academic)
    Abstract [en]

    Thiopurine S-methyltransferase (TPMT) belongs to the Class I S-adenosylmethionine-dependent methyltransferase (SAM-MT) super family of structurally related proteins. Common to the members of this large protein family is the catalysis of methylation reactions using S-adenosylmethionine (SAM) as a methyl group donor, although SAM-MTs act on a wide range of different substrates and carry out numerous biologically important functions. While the natural function of TPMT is unknown, this enzyme is involved in the metabolism of thiopurines, a class of pharmaceutical substances administered in treatment of immune-related disorders. Specifically, methylation by TPMT inactivates thiopurines and their metabolic intermediates, which reduces the efficacy of clinical treatment and increases the risk of adverse side effects. To further complicate matters, TPMT is a polymorphic enzyme with over 40 naturally occurring variants known to date, most of which exhibit lowered methylation activity towards thiopurines. Consequently, there are individual variations in TPMTmediated thiopurine inactivation, and the administered dose has to be adjusted prior to clinical treatment to avoid harmful side effects.

    Although the clinical relevance of TPMT is well established, few studies have investigated the molecular causes of the reduced methylation activity of variant proteins. In this thesis, the results of biophysical characterization of two variant proteins, TPMT*6 (Y180F) and TPMT*8 (R215H), are presented. While the properties of TPMT*8 were indistinguishable from those of the wild-type protein, TPMT*6 was found to be somewhat destabilized. Interestingly, the TPMT*6 amino acid substitution did not affect the functionality or folding pattern of the variant protein. Therefore, the decreased in vivo functionality reported for TPMT*6 is probably caused by increased proteolytic degradation in response to the reduced stability of this protein variant, rather than loss of function.

    Also presented herein are novel methodological approaches for studies of TPMT and its variants. Firstly, the advantages of using 8-anilinonaphthalene-1-sulfonic acid (ANS) to probe TPMT tertiary structure and active site integrity are presented. ANS binds exclusively to the native state of TPMT with high affinity (KD ~ 0.2 μm) and a 1:1 ratio. The stability of TPMT was dramatically increased by binding of ANS, which was shown to co-localize with the structurally similar adenine moiety of the cofactor SAM. Secondly, an enzyme activity assay based on isothermal titration calorimetry (ITC) is presented. Using this approach, the kinetics of 6-MP and 6-TG methylation by TPMT has been characterized.

  • 17.
    Blomqvist, J.
    et al.
    Department of Microbiology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    South, E.
    Department of Microbiology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Tiukova, L.
    Department of Microbiology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Momeni, M. H.
    Department of Molecular Biology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Hansson, H.
    Department of Molecular Biology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Ståhlberg, J.
    Department of Molecular Biology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Horn, S. J.
    NDepartment of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, Ås, Norway.
    Schnürer, Johan
    Department of Microbiology, Swedish University of Agricultural Sciences (SLU), Uppsala, Sweden.
    Passoth, V.
    Department of Microbiology, Uppsala Biocenter, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Fermentation of lignocellulosic hydrolysate by the alternative industrial ethanol yeast Dekkera bruxellensis2011In: Letters in Applied Microbiology, ISSN 0266-8254, E-ISSN 1472-765X, Vol. 53, no 1, 73-78 p.Article in journal (Refereed)
    Abstract [en]

    Aim: Testing the ability of the alternative ethanol production yeast Dekkera bruxellensis to produce ethanol from lignocellulose hydrolysate and comparing it to Saccharomyces cerevisiae.

    Methods and Results: Industrial isolates of D. bruxellensis and S. cerevisiae were cultivated in small-scale batch fermentations of enzymatically hydrolysed steam exploded aspen sawdust. Different dilutions of hydrolysate were tested. None of the yeasts grew in undiluted or 1 : 2 diluted hydrolysate [final glucose concentration always adjusted to 40 g l(-1) (0.22 mol l(-1))]. This was most likely due to the presence of inhibitors such as acetate or furfural. In 1 : 5 hydrolysate, S. cerevisiae grew, but not D. bruxellensis, and in 1 : 10 hydrolysate, both yeasts grew. An external vitamin source (e.g. yeast extract) was essential for growth of D. bruxellensis in this lignocellulosic hydrolysate and strongly stimulated S. cerevisiae growth and ethanol production. Ethanol yields of 0 42 +/- 0 01 g ethanol (g glucose)(-1) were observed for both yeasts in 1 : 10 hydrolysate. In small-scale continuous cultures with cell recirculation, with a gradual increase in the hydrolysate concentration, D. bruxellensis was able to grow in 1 : 5 hydrolysate. In bioreactor experiments with cell recirculation, hydrolysate contents were increased up to 1 : 2 hydrolysate, without significant losses in ethanol yields for both yeasts and only slight differences in viable cell counts, indicating an ability of both yeasts to adapt to toxic compounds in the hydrolysate.

    Conclusions: Dekkera bruxellensis and S. cerevisiae have a similar potential to ferment lignocellulose hydrolysate to ethanol and to adapt to fermentation inhibitors in the hydrolysate.

    Significance and Impact of the study: This is the first study investigating the potential of D. bruxellensis to ferment lignocellulosic hydrolysate. Its high competitiveness in industrial fermentations makes D. bruxellensis an interesting alternative for ethanol production from those substrates.

  • 18.
    Branneby, Cecilia
    et al.
    Cambrex Karlskoga AB.
    Svedendahl, Maria
    KTH, School of Biotechnology (BIO), Biochemistry.
    Hult, Karl
    KTH, School of Biotechnology (BIO), Biochemistry.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    Lipase-Catalyzed Aldol and Michael-Type Reactions2006Conference paper (Refereed)
  • 19.
    Branneby, Cecilia
    et al.
    Cambrex Karlskoga AB.
    Svedendahl, Maria
    KTH, School of Biotechnology (BIO), Biochemistry.
    Hult, Karl
    KTH, School of Biotechnology (BIO), Biochemistry.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    Lipase-Catalyzed Aldol and Michael-Type Reactions2005In: Book of abstracts, 2005Conference paper (Refereed)
  • 20.
    Broberg, Anders
    et al.
    Department of Chemistry, Swedish University of Agricultural Sciences, Sweden.
    Jacobsson, Karin
    Department of Microbiology, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Ström, Katrin
    Department of Microbiology, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Schnürer, Johan
    Department of Microbiology, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Metabolite profiles of lactic acid bacteria in grass silage2007In: Applied and Environmental Microbiology, ISSN 0099-2240, E-ISSN 1098-5336, Vol. 73, no 17, 5547-5552 p.Article in journal (Refereed)
    Abstract [en]

    The metabolite production of lactic acid bacteria JAB) on silage was investigated. The aim was to compare the production of antifungal metabolites in silage with the production in liquid cultures previously studied in our laboratory. The following metabolites were found to be present at elevated concentrations in silos inoculated with LAB strains: 3-hydroxydecanoic acid, 2-hydroxy-4-methylpentanoic acid, benzoic acid, catechol, hydrocinnamic acid, salicylic acid, 3-phenyllactic acid, 4-hydroxybenzoic acid, (trans, trans)-3,4-dihydroxycyclohexane-1-carboxylic acid, p-hydrocoumaric acid, vanillic acid, azelaic acid, hydroferulic acid, p-coumaric acid, hydrocaffeic acid, ferulic acid, and caffeic acid. Among these metabolites, the antifungal compounds 3-phenyllactic acid and 3-hydroxydecanoic acid were previously isolated in our laboratory from liquid cultures of the same LAB strains by bioassay-guided fractionation. It was concluded that other metabolites, e.g., p-hydrocoumaric acid, hydroferulic acid, and p-coumaric acid, were released from the grass by the added LAB strains. The antifungal activities of the identified metabolites in 100 mM lactic acid were investigated. The MICs against Pichia anomala, Penicillium roqueforti, and Aspergillus fumigatus were determined, and 3-hydroxydecanoic acid showed the lowest MIC (0.1 mg ml(-1) for two of the three test organisms).

  • 21.
    Carvalho, Alexandra T P
    et al.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
    Barrozo, Alexandre
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Doron, Dvir
    Kilshtain, Alexandra Vardi
    Major, Dan Thomas
    Kamerlin, Lynn Shina Caroline
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
    Challenges in computational studies of enzyme structure, function and dynamics2014In: Journal of Molecular Graphics and Modelling, ISSN 1093-3263, E-ISSN 1873-4243, Vol. 54, 62-79 p.Article, review/survey (Refereed)
    Abstract [en]

    In this review we give an overview of the field of Computational enzymology. We start by describing the birth of the field, with emphasis on the work of the 2013 chemistry Nobel Laureates. We then present key features of the state-of-the-art in the field, showing what theory, accompanied by experiments, has taught us so far about enzymes. We also briefly describe computational methods, such as quantum mechanics-molecular mechanics approaches, reaction coordinate treatment, and free energy simulation approaches. We finalize by discussing open questions and challenges.

  • 22.
    Cavka, Adnan
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Biorefining of lignocellulose: Detoxification of inhibitory hydrolysates and potential utilization of residual streams for production of enzymes2013Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Lignocellulosic biomass is a renewable resource that can be utilized for the production of biofuels, chemicals, and bio-based materials. Biochemical conversion of lignocellulose to advanced biofuels, such as cellulosic ethanol, is generally performed through microbial fermentation of sugars generated by thermochemical pretreatment of the biomass followed by an enzymatic hydrolysis of the cellulose. The aims of the research presented in this thesis were to address problems associated with pretreatment by-products that inhibit microbial and enzymatic biocatalysts, and to investigate the potential of utilizing residual streams from pulp mills and biorefineries to produce hydrolytic enzymes.

    A novel method to detoxify lignocellulosic hydrolysates to improve the fermentability was investigated in experiments with the yeast Saccharomyces cerevisiae. The method is based on treatment of lignocellulosic slurries and hydrolysates with reducing agents, such as sodium dithionite and sodium sulfite. The effects of treatment with sodium borohydride were also investigated. Treatment of a hydrolysate of Norway spruce by addition of 10 mM dithionite resulted in an increase of the balanced ethanol yield from 0.03 to 0.35 g/g. Similarly, the balanced ethanol yield of a hydrolysate of sugarcane bagasse increased from 0.06 to 0.28 g/g after treatment with 10 mM dithionite. In another study with a hydrolysate of Norway spruce, addition of 34 mM borohydride increased the balanced ethanol yield from 0.02 to 0.30 g/g, while the ethanol productivity increased from 0.05 to 0.57 g/(L×h). While treatment with sulfur oxyanions had a positive effect on microbial fermentation and enzymatic hydrolysis, treatment with borohydride resulted in an improvement only for the microbial fermentation. The chemical effects of treatments of hydrolysates with sodium dithionite, sodium sulfite, and sodium borohydride were investigated using liquid chromatography-mass spectrometry (LC-MS). Treatments with dithionite and sulfite were found to rapidly sulfonate inhibitors already at room temperature and at a pH that is compatible with enzymatic hydrolysis and microbial fermentation. Treatment with borohydride reduced inhibitory compounds, but the products were less hydrophilic than the products obtained in the reactions with the sulfur oxyanions.

    The potential of on-site enzyme production using low-value residual streams, such as stillage, was investigated utilizing recombinant Aspergillus niger producing xylanase and cellulase. A xylanase activity of 8,400 nkat/ml and a cellulase activity of 2,700 nkat/ml were reached using stillages from processes based on waste fiber sludge. The fungus consumed a large part of the xylose, the acetic acid, and the oligosaccharides that were left in the stillages after fermentation with S. cerevisiae. In another study, the capability of two filamentous fungi (A. niger and Trichoderma reesei) and three yeasts (S. cerevisiae, Pichia pastoris, and Yarrowia lipolytica) to grow on inhibitory lignocellulosic media were compared. The results indicate that the two filamentous fungi had the best capability to utilize different nutrients in the media, while the S. cerevisiae strain exhibited the best tolerance against the inhibitors. Utilization of different nutrients would be especially important in enzyme production using residual streams, while tolerance against inhibitors is desirable in a consolidated bio-process in which the fermenting microorganism also contributes by producing enzymes.

  • 23.
    Cavka, Adnan
    et al.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Alriksson, Björn
    Processum Biorefinery Initiative AB, 891 22 Örnsköldsvik, Sweden.
    Rose, Shaunita H.
    Department of Microbiology, Stellenbosch University, Stellenbosch, 7602, South Africa.
    van Zyl, Willem H.
    Department of Microbiology, Stellenbosch University, Stellenbosch, 7602, South Africa.
    Jönsson, Leif J.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Production of cellulosic ethanol and enzyme from waste fiber sludge using SSF, recycling of hydrolytic enzymes and yeast, and recombinant cellulase-producing Aspergillus niger2014In: Journal of Industrial Microbiology & Biotechnology, ISSN 1367-5435, E-ISSN 1476-5535, Vol. 41, no 8, 1191-1200 p.Article in journal (Refereed)
    Abstract [en]

    Bioethanol and enzymes were produced from fiber sludges through sequential microbial cultivations. After a first simultaneous saccharification and fermentation (SSF) with yeast, the bioethanol concentrations of sulfate and sulfite fiber sludges were 45.6 and 64.7 g/L, respectively. The second SSF, which included fresh fiber sludges and recycled yeast and enzymes from the first SSF, resulted in ethanol concentrations of 38.3 g/L for sulfate fiber sludge and 24.4 g/L for sulfite fiber sludge. Aspergillus niger carrying the endoglucanase-encoding Cel7B gene of Trichoderma reesei was grown in the spent fiber sludge hydrolysates. The cellulase activities obtained with spent hydrolysates of sulfate and sulfite fiber sludges were 2,700 and 2,900 nkat/mL, respectively. The high cellulase activities produced by using stillage and the significant ethanol concentrations produced in the second SSF suggest that onsite enzyme production and recycling of enzyme are realistic concepts that warrant further attention.

  • 24.
    Cavka, Adnan
    et al.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Jönsson, Leif J.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Comparison of the growth of filamentous fungi and yeasts in lignocellulose-derived mediaManuscript (preprint) (Other academic)
  • 25.
    Chen, Shan
    et al.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Land, Henrik
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Svedendahl Humble, Maria
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Stabilization of an amine transaminase for biocatalysis2016In: Journal of Molecular Catalysis B: Enzymatic, ISSN 1381-1177, E-ISSN 1873-3158, Vol. 124, 20-28 p.Article in journal (Refereed)
    Abstract [en]

    The amine transaminase from Chromobacterium violaceum (Cv-ATA) is a well-known enzyme to achievechiral amines of high enantiomeric excess in laboratory scales. However, the low operational stabilityof Cv-ATA limits the enzyme applicability on larger scales. In order to improve the operational stabilityof Cv-ATA, and thereby extending its applicability, factors (additives, co-solvents, organic solvents anddifferent temperatures) targeting enzyme stability and activity were explored in order to find out how tostore and apply the enzyme. The present investigation shows that the melting point of Cv-ATA is improvedby adding sucrose or glycerol, separately. Further, by storing the enzyme at higher concentrations and inco-solvents, such as; 50% glycerol, 20% methanol or 10% DMSO, the active dimeric structure of Cv-ATAis retained. Enzyme stored in 50% glycerol at −20◦C was e.g., still fully active after 6 months. Finally,the enzyme performance was improved 5-fold by a co-lyophilization with surfactants prior to usage inisooctane.

  • 26.
    Engelmark Cassimjee, Karim
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    Affinity Tag Purification Method and Immobilization of the Promiscuous Enzyme Alanine Racemase2006Conference paper (Refereed)
  • 27.
    Engelmark Cassimjee, Karim
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    Affinity Tag Purification Method of the Promiscuous Enzyme Alanine Racemase2006In: Book of abstracts, 2006Conference paper (Other academic)
  • 28.
    Engelmark Cassimjee, Karim
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    Immobilization Method for the Promiscuous Enzyme Alanine Racemase2007In: BIOTRANS Oviedo 2007 / [ed] Vicente Gotor, 2007Conference paper (Refereed)
  • 29.
    Engelmark Cassimjee, Karim
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Branneby, Cecilia
    Cambrex Karlskoga AB.
    Sjöstrand, Ulf
    Cambrex Karlskoga AB.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    High Yield Transamination with Isopropyl Amine as Donor, by Employment of YADH and in situ Cofactor Regeneration2009Conference paper (Refereed)
  • 30.
    Engelmark Cassimjee, Karim
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Branneby, Cecilia
    Cambrex Karlskoga AB.
    Sjöstrand, Ulf
    Cambrex Karlskoga AB.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    High Yield Transamination with Isopropyl Amine as Donor, by Employment of YADH and in situ Cofactor Regeneration2009In: Book of abstracts, 2009Conference paper (Refereed)
  • 31.
    Engelmark Cassimjee, Karim
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Kourist, Robert
    University of Greifswald, Germany.
    Lindberg, Diana
    Uppsala university, SE.
    Wittrup Larsen, Marianne
    KTH, School of Biotechnology (BIO), Biochemistry.
    Widersten, Mikael
    Uppsala university, SE.
    Bornscheuer, Uwe T
    University of Greifswald, DE.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    A One Step Enzyme Extraction and Immobilization Method for Organic and Aqueous Solvents2008In: Biocat2008 / [ed] Ralf Grote, Garabed Antranikian, Hamburg, Germany: TuTech Innovation GmbH , 2008Conference paper (Refereed)
  • 32.
    Engelmark Cassimjee, Karim
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry (closed 20130101).
    Kourist, Robert
    University of Greifswald, Germany.
    Lindberg, Diana
    Uppsala university, SE.
    Wittrup Larsen, Marianne
    KTH, School of Biotechnology (BIO), Biochemistry (closed 20130101).
    Widersten, Mikael
    Uppsala university, SE.
    Bornscheuer, Uwe T
    University of Greifswald, DE.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry (closed 20130101).
    A One Step General Enzyme Immobilization Method for Organic and Aqueous Solvents2008In: Book of Abstracts, 2008Conference paper (Refereed)
  • 33.
    Engelmark Cassimjee, Karim
    et al.
    KTH, School of Biotechnology (BIO), Biochemistry.
    Svedendahl, Maria
    KTH, School of Biotechnology (BIO), Biochemistry.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry.
    Rational Redesign of omega-Transaminase2010In: Biocat2010, Hamburg, Germany: TuTech Verlag , 2010Conference paper (Refereed)
  • 34.
    Gullfot, Fredrika
    KTH, School of Biotechnology (BIO), Glycoscience.
    Synthesis of xyloglucan oligo- and polysaccharides with glycosynthase technology2009Licentiate thesis, comprehensive summary (Other academic)
    Abstract [en]

    Xyloglucans are polysaccharides found as storage polymers in seeds and tubers, and as cross-linking glycans in the cell wall of plants. Their structure is complex with intricate branching patterns, which contribute to the physical properties of the polysaccharide including its binding to and interaction with other glycans such as cellulose.

    Xyloglucan is widely used in bulk quantities in the food, textile and paper making industries. With an increasing interest in technically more advanced applications of xyloglucan, such as novel biocomposites, there is a need to understand and control the properties and interactions of xyloglucan with other compounds, to decipher the relationship between xyloglucan structure and function, and in particular the effect of different branching patterns. However, due to the structural heterogeneity of the polysaccharide as obtained from natural sources, relevant studies have not been possible to perform in practise. This fact has stimulated an interest in synthetic methods to obtain xyloglucan mimics and analogs with well-defined structure and decoration patterns.

    Glycosynthases are hydrolytically inactive mutant glycosidases that catalyse the formation of glycosidic linkages between glycosyl fluoride donors and glycoside acceptors. Since its first conception in 1998, the technology is emerging as a useful tool in the synthesis of large, complex polysaccharides. This thesis presents the generation and characterisation of glycosynthases based on xyloglucanase scaffolds for the synthesis of well-defined homogenous xyloglucan oligo- and polysaccharides with regular substitution patterns.

  • 35.
    Guo, Fei
    et al.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Transaminase biocatalysis: optimization and application2017In: Green Chemistry, ISSN 1463-9262, E-ISSN 1463-9270Article in journal (Refereed)
    Abstract [en]

    Transaminases (TAs) are one of the most promising biocatalysts in organic synthesis for the preparation of chiral amino compounds. The concise reaction, excellent enantioselectivity, environmental friendliness and compatibility with other enzymatic or chemical systems have brought TAs to the attention of scientists working in the area of biocatalysis. However, to utilize TAs in a more efficient and economical way, attempts have to be made to optimize their performance. The demand for various substrate specificities, stability under non-physiological conditions and higher conversions in reversible reactions have been targeted and investigated thoroughly. A number of both protein- and process-based strategies have been developed to improve TAs and systems involving TAs. Moreover, by combination with other enzymes in cascade reactions or even in more complex systems, so called synthetic biology and systems biocatalysis, TAs can be biocatalysts with immense potential in the industrial production of high-value chemical products. This review will highlight strategies for optimization of TAs and will discuss a number of elegant systems for improving their performance. Transaminase biocatalysis has been, and will continue to be, one of the most interesting topics in green organic synthesis.

  • 36. Gustafsson, Kerstin
    et al.
    Blidberg, Eva
    Elfgren, I. K.
    Hellström, Anna
    Kylin, Henrik
    Gorokhova, E.
    Direct and indirect effects of the fungicide azoxystrobin in outdoor brackish water microcosms2010In: Ecotoxicology, ISSN 0963-9292, E-ISSN 1573-3017, Vol. 19, no 2, 431-444 p.Article in journal (Refereed)
    Abstract [en]

    The effects of the strobilurin fungicide azoxystrobin were studied in brackish water microcosms, with natural plankton communities and sediment. Two experiments were conducted: Experiment 1 (nominal conc. 0, 15 and 60 mu g/L, 24-L outdoor microcosms for 21 days) and a second, follow-up, Experiment 2 (nominal conc. 0, 3, 7.5, 15 mu g/L, 4-L indoor microcosms for 12 days). The microcosms represent a simplified brackish water community found in shallow semi-enclosed coastal areas in agricultural districts in the Baltic Sea region. Measured water concentrations of the fungicide (Experiment 1) were, on average, 83 and 62% of nominal concentrations directly after application, and 25 and 30% after 21 days, for the low and high dose treatments, respectively, corresponding to mean DT50-values of 15.1 and 25.8 days, for low and high dose treatments, respectively. In Experiment 1, direct toxic effects on calanoid copepods at both test concentrations were observed. Similarly, in Experiment 2, the copepod abundance was significantly reduced at all tested concentrations. There were also significant secondary effects on zooplankton and phytoplankton community structure, standing stocks and primary production. Very few ecotoxicological studies have investigated effects of plant protection products on Baltic organisms in general and effects on community structure and function specifically. Our results show that azoxystrobin is toxic to brackish water copepods at considerably lower concentrations than previously reported from single species tests on freshwater crustaceans, and that direct toxic effects on this ecologically important group may lead to cascade effects altering lower food webs and ecosystem functioning.

  • 37.
    Gustavsson, Martin
    et al.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Muraleedharan, Madhu Nair
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Larsson, Gen
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Surface Expression of omega-Transaminase in Escherichia coli2014In: Applied and Environmental Microbiology, ISSN 0099-2240, E-ISSN 1098-5336, Vol. 80, no 7, 2293-2298 p.Article in journal (Refereed)
    Abstract [en]

    Chiral amines are important for the chemical and pharmaceutical industries, and there is rapidly growing interest to use transaminases for their synthesis. Since the cost of the enzyme is an important factor for process economy, the use of whole-cell biocatalysts is attractive, since expensive purification and immobilization steps can be avoided. Display of the protein on the cell surface provides a possible way to reduce the mass transfer limitations of such biocatalysts. However, transaminases need to dimerize in order to become active, and furthermore, they require the cofactor pyridoxal phosphate; consequently, successful transaminase surface expression has not been reported thus far. In this work, we produced an Arthrobacter citreus omega-transaminase in Escherichia coli using a surface display vector based on the autotransporter adhesin involved in diffuse adherence (AIDA-I), which has previously been used for display of dimeric proteins. The correct localization of the transaminase in the E. coli outer membrane and its orientation toward the cell exterior were verified. Furthermore, transaminase activity was detected exclusively in the outer membrane protein fraction, showing that successful dimerization had occurred. The transaminase was found to be present in both full-length and proteolytically degraded forms. The removal of this proteolysis is considered to be the main obstacle to achieving sufficient whole-cell transaminase activity.

  • 38.
    Hassan, Noor
    et al.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Geiger, Barbara
    Gandini, Rosaria
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Patel, Bharat K. C.
    Kittl, Roman
    Haltrich, Dietmar
    Nguyen, Thu-Ha
    Divne, Christina
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Tan, Tien Chye
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Engineering a thermostable Halothermothrix orenii beta-glucosidase for improved galacto-oligosaccharide synthesis2016In: Applied Microbiology and Biotechnology, ISSN 0175-7598, E-ISSN 1432-0614, Vol. 100, no 8, 3533-3543 p.Article in journal (Refereed)
    Abstract [en]

    Lactose is produced in large amounts as a by-product from the dairy industry. This inexpensive disaccharide can be converted to more useful value-added products such as galacto-oligosaccharides (GOSs) by transgalactosylation reactions with retaining beta-galactosidases (BGALs) being normally used for this purpose. Hydrolysis is always competing with the transglycosylation reaction, and hence, the yields of GOSs can be too low for industrial use. We have reported that a beta-glucosidase from Halothermothrix orenii (HoBGLA) shows promising characteristics for lactose conversion and GOS synthesis. Here, we engineered HoBGLA to investigate the possibility to further improve lactose conversion and GOS production. Five variants that targeted the glycone (-1) and aglycone (+1) subsites (N222F, N294T, F417S, F417Y, and Y296F) were designed and expressed. All variants show significantly impaired catalytic activity with cellobiose and lactose as substrates. Particularly, F417S is hydrolytically crippled with cellobiose as substrate with a 1000-fold decrease in apparent k(cat), but to a lesser extent affected when catalyzing hydrolysis of lactose (47-fold lower k(cat)). This large selective effect on cellobiose hydrolysis is manifested as a change in substrate selectivity from cellobiose to lactose. The least affected variant is F417Y, which retains the capacity to hydrolyze both cellobiose and lactose with the same relative substrate selectivity as the wild type, but with similar to 10-fold lower turnover numbers. Thin-layer chromatography results show that this effect is accompanied by synthesis of a particular GOS product in higher yields by Y296F and F417S compared with the other variants, whereas the variant F417Y produces a higher yield of total GOSs.

  • 39.
    Hendil-Forssell, Peter
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Rational engineering of esterases for improved amidase specificity in amide synthesis and hydrolysis2016Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Biocatalysis is an ever evolving field that uses enzymes or microorganisms for chemical synthesis. By utilizing enzymes that generally have evolved for specific reactions under mild conditions and temperatures, biocatalysis can be a more environmentally friendly option compared to traditional chemistry.

    Amide-type chemistries are important and bond formation avoiding poor atom economy is of high priority in organic chemistry. Biocatalysis could potentially be a solution but restricted substrate scope is a limitation. Esterases/lipases usually display broad substrate scope and catalytic promiscuity but are poor at hydrolyzing amides compared to amidases/proteases. The difference between the two enzyme classes is hypothesized to reside in one key hydrogen bond present in amidases, which facilitates the transition state for nitrogen inversion during catalysis.

    In this thesis the work has been focused on introducing a stabilizing hydrogen bond acceptor in esterases, mimicking that found in amidases, to develop better enzymatic catalysts for amide-based chemistries.

    By two strategies, side-chain or water interaction, variants were created in three esterases that displayed up to 210-times increased relative amidase specificity compared to the wild type. The best variant displayed reduced activation enthalpy corresponding to a weak hydrogen bond. The results show an estimated lower limit on how much the hydrogen bond can be worth to catalysis.

    MsAcT catalyze kinetically controlled N-acylations in water. An enzymatic one-pot one-step cascade was developed for the formation of amides from aldehydes in water that gave 97% conversion. In addition, engineered variants of MsAcT with increased substrate scope could synthesize an amide in water with 81% conversion, where the wild type gave no conversion. Moreover, variants of MsAcT displayed up to 32-fold change in specificity towards amide synthesis and a switch in reaction preference favoring amide over ester synthesis.

  • 40.
    Hendil-Forssell, Peter
    et al.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Semlitsch, Stefan
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Martinelle, Mats
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Engineering the esterase/acyltransferase from Mycobacterium smegmatis: extended substrate scope for amide synthesis in waterManuscript (preprint) (Other academic)
    Abstract [en]

    Some esterases/lipases display high acyl transfer activity, favoring alcoholysis over hydrolysis, which make them valuable catalysts for synthesis reactions in aqueous media. An esterase from Mycobacterium smegmatis, MsAcT, has been characterized as an efficient catalyst for ester synthesis in water. The acyl donor specificity for MsAcT was however found to be very narrow and the enzyme displayed no activity towards esters with larger acyl group than butyrate. With rational engineering, the narrow acyl donor specificity of wild type MsAcT enzyme was altered and variants displaying extended substrate scope were generated. A double mutant, T93A/F154A, could accommodate methyl nonanoate as substrate, i.e. five carbons longer acyl group as compared to wild type, without compromising the acyl transfer capabilities. With similar selectivity towards a broad range of acyl donors (propionate to nonanoate) this is a more applicable catalyst than the wild type. Furthermore, the T93A/F154A variant was an efficient catalyst for synthesis of N-benzylhexanamide in water using methyl hexanoate as acyl donor, which is not a substrate for the wild type enzyme. The conversion reached 81% and the enzyme variant could potentially be used to produce amides in water with a wide variety of acyl donors.

  • 41.
    Hendil-Forssell, Peter
    et al.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Semlitsch, Stefan
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Martinelle, Mats
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Rational engineering of an esterase/acyltransferase for improved amidase specificity in amide synthesis and hydrolysisManuscript (preprint) (Other academic)
    Abstract [en]

    The esterase/acyltransferase from Mycobacterium smegmatis, MsAcT, display high acyltransfer capacity in water media with demonstrations found for both ester and amide syntheses. However, it has recently been discovered that esterases in contrast to amidases lack a key hydrogen bond in the transition state, donated by the scissile NH-group of the substrate. Esterases with improved amidase performance have been achieved with the introduction of amino-acid side chains or water network as hydrogen bond acceptors. Using the esterase from Mycobacterium smegmatis, MsAcT, the influence of this hydrogen bond was studied in both amide hydrolysis and synthesis, using a rational engineering approach. Two positions were selected for mutagenesis and enzyme variants with improved performance in amide synthesis and hydrolysis were generated. Compared to the wild-type, variant F154A had the highest absolute increase in amidase specificity (11-fold) and I194Q had the greatest change in relative amidase versus esterase reaction specificity (160-fold). The relative reaction specificities for amide over ester synthesis followed a similar trend as that of hydrolysis and the best variant was I194Q with a 32-fold increase compared to wt. Based on MD-simulations water seems to play an important role in the transition state as a hydrogen bond bridge between the NH-group of the amide substrate and the enzyme.

  • 42. Holmquist, M.
    et al.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Biochemistry (closed 20130101).
    Creation of a synthetically useful lipase with higher than wild-type enantioselectivity and maintained catalytic activity1999In: Organic Letters, ISSN 1523-7060, E-ISSN 1523-7052, Vol. 1, no 5, 763-765 p.Article in journal (Refereed)
    Abstract [en]

    Formula presented Wild type I: 89.9% ee (E=32) Wild type II: 79.8% ee (E=10) Lipase hybrid: 95.4% ee (E=54) We have found that two Geotrichum candidum lipase isozymes have remarkably different abilities to differentiate between enantiomers of ethyl 2-methyldecanoate. By rational recombination of selected portions of the two isozymes, we have created a novel lipase with an enantioselectivity superior to that of the best wild-type parent isozyme. Site-directed mutagenesis identified two key amino acid residues responsible for the improved enantioselectivity without compromised total activity of the reengineered enzyme.

  • 43.
    Holmquist, Mats
    et al.
    KTH, Superseded Departments, Biochemistry and Biotechnology.
    Berglund, Per
    KTH, Superseded Departments, Biochemistry and Biotechnology.
    Improved lipase enantioselectivity by combinatorial and rational redesign.2000In: Abstract of Papers of the American Chemical Society, ISSN 0065-7727, Vol. 219, no 1, U163-U163 p.Article in journal (Refereed)
  • 44. Hu, Y.
    et al.
    Zhu, Z.
    Nielsen, Jens
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab. Chalmers University of Technology, Sweden.
    Siewers, V.
    Heterologous transporter expression for improved fatty alcohol secretion in yeast2018In: Metabolic engineering, ISSN 1096-7176, E-ISSN 1096-7184, Vol. 45, 51-58 p.Article in journal (Refereed)
    Abstract [en]

    The yeast Saccharomyces cerevisiae is an attractive host for industrial scale production of biofuels including fatty alcohols due to its robustness and tolerance towards harsh fermentation conditions. Many metabolic engineering strategies have been applied to generate high fatty alcohol production strains. However, impaired growth caused by fatty alcohol accumulation and high cost of extraction are factors limiting large-scale production. Here, we demonstrate that the use of heterologous transporters is a promising strategy to increase fatty alcohol production. Among several plant and mammalian transporters tested, human FATP1 was shown to mediate fatty alcohol export in a high fatty alcohol production yeast strain. An approximately five-fold increase of fatty alcohol secretion was achieved. The results indicate that the overall cell fitness benefited from fatty alcohol secretion and that the acyl-CoA synthase activity of FATP1 contributed to increased cell growth as well. This is the first study that enabled an increased cell fitness for fatty alcohol production by heterologous transporter expression in yeast, and this investigation indicates a new potential function of FATP1, which has been known as a free fatty acid importer to date. We furthermore successfully identified the functional domain of FATP1 involved in fatty alcohol export through domain exchange between FATP1 and another transporter, FATP4. This study may facilitate a successful commercialization of fatty alcohol production in yeast and inspire the design of novel cell factories.

  • 45.
    Håkansson Hederos, Sofia
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Catalysis and Site-Specific Modification of Glutathione Transferases Enabled by Rational Design2005Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    This thesis describes the rational design of a novel enzyme, a thiolester hydrolase, derived from human glutathione transferase (GST) A1-1 by the introduction of a single histidine residue. The first section of the thesis describes the design and the determination of the reaction mechanism. The design was based on the crystal structure of human GST A1-1 complexed with S-benzylglutathione. The resulting enzyme, A216H, catalyzed the hydrolysis of the non-natural substrate GSB, a thiolester of glutathione and benzoic acid. The reaction followed saturation kinetics with a kcat of 0.00078 min-1 and KM of 5 μM. The rate constant ratio, (kcat/KM)/kuncat, was found to be more than 107 M-1. The introduction of a single His residue in position 216 opened up a novel reaction pathway in human GST A1-1 and is a nice example of catalytic promiscuity. The substrate requirements were investigated and A216H was found to be selective since only two out of 18 GS-thiolesters tested were substrates for A216H. The reaction mechanism of the A216H-catalyzed hydrolysis of GSB was determined and found to proceed via an acyl intermediate at Y9. The hydrolysis was catalyzed by H216 that acts as a general base and the deacylation was found to be the rate-determining step. The Y9-intermediate could be selectively trapped by oxygen nucleophiles and primary alcohols, in particular 1-propanol and trifluoroethanol, were the most efficient. In addition, saturation kinetics was obtained in the acyl transfer reaction with 1-propanol indicating the presence of a second binding site in A216H.

    The second section of this thesis describes the site-specific covalent modification of human GST A1-1. The addition of GSB to the wild-type protein results in a site-specific benzoylation of only one tyrosine residue, Y9, out of ten present in the protein (one out of totally 51 nucleophiles). The reaction was tested with five GST classes (Alpha, Mu, Pi, Theta and Omega) and found to be specific for the Alpha class isoenzymes. The covalent modification reaction was further refined to target a single lysine residue, K216, providing a more stable linkage in the form of an amide bond. The reaction was found to be versatile and approximately 50% of the GS-thiolesters tested acylated K216, including a fluorophore.

  • 46.
    Johansson, Fredrik
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Microscale measurement of kinetic binding properties of monoclonal antibodies in solution using Gyrolab2011Independent thesis Advanced level (degree of Master (One Year)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    The number of monoclonal antibodies approved for therapeutic use has increased rapidlyover the last decade. As a consequence, precise and robust kinetic characterization techniquesare crucial in order to select the best suitable candidates. A kinetic characterization methodwas developed in Gyrolab with automated sample transfers. The characterization wasperformed in solution in a mixing CD, containing an integrated nanoliter mixing chamberwith affinity binding columns. Association rate constants were determined for four anti-TSHantibodies with values ranging from 3x105 M-1s-1 to 10x105 M-1s-1. The antibodies wereranked according to kass. Reproducibility

  • 47. Kang, Min-Kyoung
    et al.
    Zhou, Yongjin J.
    Buijs, Nicolaas A.
    Nielsen, Jens
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Functional screening of aldehyde decarbonylases for long-chain alkane production by Saccharomyces cerevisiae2017In: Microbial Cell Factories, ISSN 1475-2859, E-ISSN 1475-2859, Vol. 16, 74Article in journal (Refereed)
    Abstract [en]

    Background: Low catalytic activities of pathway enzymes are often a limitation when using microbial based chemical production. Recent studies indicated that the enzyme activity of aldehyde decarbonylase (AD) is a critical bottleneck for alkane biosynthesis in Saccharomyces cerevisiae. We therefore performed functional screening to identify efficient ADs that can improve alkane production by S. cerevisiae. Results: A comparative study of ADs originated from a plant, insects, and cyanobacteria were conducted in S. cerevisiae. As a result, expression of aldehyde deformylating oxygenases (ADOs), which are cyanobacterial ADs, from Synechococcus elongatus and Crocosphaera watsonii converted fatty aldehydes to corresponding Cn-1 alkanes and alkenes. The CwADO showed the highest alkane titer (0.13 mg/L/OD600) and the lowest fatty alcohol production (0.55 mg/L/OD600). However, no measurable alkanes and alkenes were detected in other AD expressed yeast strains. Dynamic expression of SeADO and CwADO under GAL promoters increased alkane production to 0.20 mg/L/OD600 and no fatty alcohols, with even number chain lengths from C8 to C14, were detected in the cells. Conclusions: We demonstrated in vivo enzyme activities of ADs by displaying profiles of alkanes and fatty alcohols in S. cerevisiae. Among the AD enzymes evaluated, cyanobacteria ADOs were found to be suitable for alkane biosynthesis in S. cerevisiae. This work will be helpful to decide an AD candidate for alkane biosynthesis in S. cerevisiae and it will provide useful information for further investigation of AD enzymes with improved activities.

  • 48. Kocík, Jaroslav
    et al.
    Samikannu, Ajaikumar
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Bourajoini, Hasna
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Pham, Tung Ngoc
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Mikkola, Jyri-Pekka
    Umeå University, Faculty of Science and Technology, Department of Chemistry. Faculty of Science and Engineering, Industrial Chemistry & Reaction Engineering, Johan Gadolin Process Chemistry Centre, Åbo Akademi University, Åbo-Turku, Finland.
    Hájek, Martin
    Čapek, Libor
    Screening of active solid catalysts for esterification of tall oil fatty acids with methanol2017In: Journal of Cleaner Production, ISSN 0959-6526, E-ISSN 1879-1786, Vol. 155, no 1, 34-38 p.Article in journal (Refereed)
    Abstract [en]

    The paper is focused on the description of the activity/selectivity of mesoporous silica based materials loaded with various types of active species in the esterification of tall oil free fatty acids. The metals such as aluminium, molybdenum, gallium and zinc, including their combinations were impregnated on the mesoporous silica, which was tested in esterification reaction. All these catalysts preserved its tall oil free fatty conversion in the first and the second catalytic cycles. However, while only insignificant amount of gallium or molybdenum was lost from the solid catalyst into the liquid phases, zinc leached from every studied solid catalyst. In contrast to impregnated gallium on mesoporous silica, which exhibited higher acidity and higher tall oil free fatty acids conversion in the first catalytic cycle, but its value was not preserved in the second catalytic test.

  • 49.
    Kurtovic, Sanela
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Modén, Olof
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Shokeer, Abeer
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Mannervik, Bengt
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Structural determinants of glutathione transferases with azathioprine activity identified by DNA shuffling of alpha class members2008In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 375, no 5, 1365-1379 p.Article in journal (Refereed)
    Abstract [en]

    A library of alpha class glutathione transferases (GSTs), composed of chimeric enzymes derived from human (A1-1, A2-2 and A3-3), bovine (A1-1) and rat (A2-2 and A3-3) cDNA sequences was constructed by the method of DNA shuffling. The GST variants were screened in bacterial lysates for activity with the immunosuppressive agent azathioprine, a prodrug that is transformed into its active form, 6-mercaptopurine, by reaction with the tripeptide glutathione catalyzed by GSTs. Important structural determinants for activity with azathioprine were recognized by means of primary structure analysis and activities of purified enzymes chosen from the screening. The amino acid sequences could be divided into 23 exchangeable segments on the basis of the primary structures of 45 chosen clones. Segments 2, 20, 21, and 22 were identified as primary determinants of the azathioprine activity representing two of the regions forming the substrate-binding H-site. Segments 21 and 22 are situated in the C-terminal helix characterizing alpha class GSTs, which is instrumental in their catalytic function. The study demonstrates the power of DNA shuffling in identifying segments of primary structure that are important for catalytic activity with a targeted substrate. GSTs in combination with azathioprine have potential as selectable markers for use in gene therapy. Knowledge of activity-determining segments in the structure is valuable in the protein engineering of glutathione transferase for enhanced or suppressed activity.

  • 50.
    Land, Henrik
    et al.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Hendil-Forssell, Peter
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Martinelle, Mats
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Berglund, Per
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    One-pot biocatalytic amine transaminase/acyl transferase cascade for aqueous formation of amides from aldehydes or ketones2016In: catalysis science & technology, ISSN 2044-4753, Vol. 6, 2897-2900 p.Article in journal (Refereed)
    Abstract [en]

    An efficient one-pot one-step biocatalytic amine transaminase/acyl transferase cascade for the formation of amides from the corresponding aldehydes and ketones in aqueous solution has been developed. N-benzyl-2-methoxyacetamide has been synthesized utlilizing the developed cascade in conversions up to 97%. The cascade was also evaluated for the synthesis of chiral amides.

12 1 - 50 of 91
CiteExportLink to result list
Permanent link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf