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  • 1.
    Andersson, Anders
    et al.
    KTH, Superseded Departments, Biotechnology.
    Keskitalo, J.
    Sjödin, A.
    Bhalerao, Rupali
    KTH, Superseded Departments, Biotechnology.
    Sterky, Fredrik
    KTH, Superseded Departments, Biotechnology.
    Wissel, K.
    Tandre, K.
    Aspeborg, Henrik
    KTH, Superseded Departments, Biotechnology.
    Moyle, R.
    Ohmiya, Y.
    Brunner, A.
    Gustafsson, P.
    Karlsson, J.
    Lundeberg, Joakim
    KTH, Superseded Departments, Biotechnology.
    Nilsson, O.
    Sandberg, G.
    Strauss, S.
    Sundberg, B.
    Uhlén, Mathias
    KTH, Superseded Departments, Biotechnology.
    Jansson, S.
    Nilsson, Peter
    KTH, Superseded Departments, Biotechnology.
    A transcriptional timetable of autumn senescence2004In: Genome Biology, ISSN 1465-6906, E-ISSN 1474-760X, Vol. 5, no 4, p. R24-Article in journal (Refereed)
    Abstract [en]

    Background: We have developed genomic tools to allow the genus Populus ( aspens and cottonwoods) to be exploited as a full-featured model for investigating fundamental aspects of tree biology. We have undertaken large-scale expressed sequence tag ( EST) sequencing programs and created Populus microarrays with significant gene coverage. One of the important aspects of plant biology that cannot be studied in annual plants is the gene activity involved in the induction of autumn leaf senescence. Results: On the basis of 36,354 Populus ESTs, obtained from seven cDNA libraries, we have created a DNA microarray consisting of 13,490 clones, spotted in duplicate. Of these clones, 12,376 (92%) were confirmed by resequencing and all sequences were annotated and functionally classified. Here we have used the microarray to study transcript abundance in leaves of a free-growing aspen tree ( Populus tremula) in northern Sweden during natural autumn senescence. Of the 13,490 spotted clones, 3,792 represented genes with significant expression in all leaf samples from the seven studied dates. Conclusions: We observed a major shift in gene expression, coinciding with massive chlorophyll degradation, that reflected a shift from photosynthetic competence to energy generation by mitochondrial respiration, oxidation of fatty acids and nutrient mobilization. Autumn senescence had much in common with senescence in annual plants; for example many proteases were induced. We also found evidence for increased transcriptional activity before the appearance of visible signs of senescence, presumably preparing the leaf for degradation of its components.

  • 2.
    Aspeborg, Henrik
    KTH, Superseded Departments, Biotechnology.
    Discovery of fiber-active enzymes in Populus wood2004Doctoral thesis, comprehensive summary (Other scientific)
    Abstract [en]

    Renewable fibers produced by forest trees provide excellentraw material of high economic value for industrialapplications. Despite this, the genes and corresponding enzymesinvolved in wood fiber biosynthesis in trees are poorlycharacterized. This thesis describes a functional genomicsapproach for the identification of carbohydrate-active enzymesinvolved in secondary cell wall (wood) formation in hybridaspen.

    First, a 3' target amplification method was developed toenable microarray-based gene expression analysis on minuteamounts of RNA. The amplification method was evaluated usingboth a smaller microarray containing 192 cDNA clones and alarger microarray containing 2995 cDNA clones that werehybridized with targets isolated from xylem and phloem.Moreover, a gene expression study of phloem differentiation wasperformed to show the usefulness of the amplificationmethod.

    A microarray containing 2995 cDNA clones representing aunigene set of a cambial region EST library was used to studygene expression during wood formation. Transcript populationsfrom thin tissue sections representing different stages ofxylem development were hybridized onto the microarrays. It wasdemonstrated that genes encoding lignin and cellulosebiosynthetic enzymes, as well as a number of genes withoutassigned function, were differentially expressed across thedevelopmental gradient.

    Microarrays were also used to track changes in geneexpression in the developing xylem of transgenic, GA-20 oxidaseoverexpressing hybrid aspens that had increased secondarygrowth. The study revealed that a number of genes encoding cellwall related enzymes were upregulated in the transgenic trees.Moreover, most genes with high transcript changes could beassigned a role in the early events of xylogenesis.

    Ten genes encoding putative cellulose synthases (CesAs) wereidentified in our ownPopulusESTdatabase. Full length cDNA sequences wereobtained for five of them. Expression analyses performed withreal-time PCR and microarrays in normal wood undergoingxylogenesis and in tension wood revealed xylem specificexpression of four putative CesA isoenzymes.

    Finally, an approach combining expressionprofiling,bioinformatics as well as EST and full length sequencing wasadopted to identify secondary cell wall related genes encodingcarbohydrate-active enzymes, such as glycosyltransferases andglycoside hydrolases. As expected, glycosyltransferasesinvolved in the carbohydrate biosynthesis dominated thecollection of the secondary cell wall related enzymes that wereidentified.

    Key words:Populus, xylogenesis, secondary cell wall,cellulose, hemicellulose, microarrays, transcript profiling,carbohydrate-active enzyme, glycosyltransferase, glycosidehydrolase

  • 3.
    Aspeborg, Henrik
    et al.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Bhalerao, Rishikeshi
    Hertzberg, Magnus
    Johansson, Karin
    Johnsson, P.
    Karlsson, Ann
    Sandberg, Göran
    Schrader, Jarmo
    Sundberg, Björn
    Teeri, Tuula
    Trygg, Johan
    Wallbäcks, Lars
    Vegetabile material, plants and a method of producing a plant having altered lignin properties2008Patent (Other (popular science, discussion, etc.))
    Abstract [en]

    The present invention is related to a set of genes, which when modified in plants gives altered lignin properties. The invention provides DNA construct such as a vector useful in the method of the invention. Further, the invention relates to a plant cell or plant progeny of the plants and wood produced by the plants according to the invention Lower lignin levels will result in improved saccharification for bio-refining and ethanol production and improved pulp and paper. Increased lignin levels will utilise lignin properties for energy production. The genes and DNA constructs may be used for the identification of plants having altered lignin characteristics as compared to the wild-type. According to the invention genes and DNA constructs may also be used as candidate genes in marker assisted breeding.

  • 4.
    Aspeborg, Henrik
    et al.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Coutinho, Pedro M.
    Wang, Yang
    KTH, School of Biotechnology (BIO), Glycoscience.
    Brumer, Harry
    KTH, School of Biotechnology (BIO), Glycoscience.
    Henrissat, Bernard
    Evolution, substrate specificity and subfamily classification of glycoside hydrolase family 5 (GH5)2012In: BMC Evolutionary Biology, ISSN 1471-2148, E-ISSN 1471-2148, Vol. 12, no 1, p. 186-Article in journal (Refereed)
    Abstract [en]

    Background: The large Glycoside Hydrolase family 5 (GH5) groups together a wide range of enzymes acting on beta-linked oligo- and polysaccharides, and glycoconjugates from a large spectrum of organisms. The long and complex evolution of this family of enzymes and its broad sequence diversity limits functional prediction. With the objective of improving the differentiation of enzyme specificities in a knowledge-based context, and to obtain new evolutionary insights, we present here a new, robust subfamily classification of family GH5. Results: About 80% of the current sequences were assigned into 51 subfamilies in a global analysis of all publicly available GH5 sequences and associated biochemical data. Examination of subfamilies with catalytically-active members revealed that one third are monospecific (containing a single enzyme activity), although new functions may be discovered with biochemical characterization in the future. Furthermore, twenty subfamilies presently have no characterization whatsoever and many others have only limited structural and biochemical data. Mapping of functional knowledge onto the GH5 phylogenetic tree revealed that the sequence space of this historical and industrially important family is far from well dispersed, highlighting targets in need of further study. The analysis also uncovered a number of GH5 proteins which have lost their catalytic machinery, indicating evolution towards novel functions. Conclusion: Overall, the subfamily division of GH5 provides an actively curated resource for large-scale protein sequence annotation for glycogenomics; the subfamily assignments are openly accessible via the Carbohydrate-Active Enzyme database at http://www.cazy.org/GH5.html.

  • 5.
    Aspeborg, Henrik
    et al.
    KTH, School of Biotechnology (BIO).
    Schrader, J.
    Coutinho, P. M.
    Stam, M.
    Kallas, A.
    Djerbi, S.
    Nilsson, Peter
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Denman, S.
    Amini, B.
    Sterky, Fredrik
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Master, E.
    Sandberg, G.
    Mellerowicz, E.
    Sundberg, B.
    Henrissat, B.
    Teeri, Tuula T.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Carbohydrate-active enzymes involved in the secondary cell wall biogenesis in hybrid aspen2005In: Plant Physiology, ISSN 0032-0889, E-ISSN 1532-2548, Vol. 137, no 3, p. 983-997Article in journal (Refereed)
    Abstract [en]

    Wood formation is a fundamental biological process with significant economic interest. While lignin biosynthesis is currently relatively well understood, the pathways leading to the synthesis of the key structural carbohydrates in wood fibers remain obscure. We have used a functional genomics approach to identify enzymes involved in carbohydrate biosynthesis and remodeling during xylem development in the hybrid aspen Populus tremula x tremuloides. Microarrays containing cDNA clones from different tissue-specific libraries were hybridized with probes obtained from narrow tissue sections prepared by cryosectioning of the developing xylem. Bioinformatic analyses using the sensitive tools developed for carbohydrate-active enzymes allowed the identification of 25 xylem-specific glycosyltransferases belonging to the Carbohydrate-Active EnZYme families GT2, GT8, GT14, GT31, GT43, GT47, and GT61 and nine glycosidases (or transglycosidases) belonging to the Carbohydrate-Active EnZYme families GH9, GH10, GH16, GH17, GH19, GH28, GH35, and GH51. While no genes encoding either polysaccharide lyases or carbohydrate esterases were found among the secondary wall-specific genes, one putative O-acetyltransferase was identified. These wood-specific enzyme genes constitute a valuable resource for future development of engineered fibers with improved performance in different applications.

  • 6.
    Blomqvist, Kristina
    et al.
    KTH, School of Biotechnology (BIO), Wood Biotechnology.
    Djerbi, Soraya
    KTH, School of Biotechnology (BIO), Wood Biotechnology.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Wood Biotechnology.
    Teeri, Tuula
    KTH, School of Biotechnology (BIO), Wood Biotechnology.
    Cellulose Biosynthesis in Forest Trees2007In: Cellulose: Molecular and Structural Biology: Selected Articles on the Synthesis, Structure, and Applications of Cellulose / [ed] R. Malcolm Brown and Inder M. Saxena, Dordrecht: Springer Netherlands, 2007, p. 85-106Chapter in book (Other academic)
    Abstract [en]

    Wood formation is a fundamental biological process of significant economic andcommercial interest. During wood formation, most glucose from the carbohydratemetabolism is channeled to cellulose in the secondary cell walls. The cellulose microfibrils associate with hemicellulose, proteins, and lignin to form the strong and flexiblebiocomposite known as wood. As the main wood component, cellulose is essential forthe survival of trees and for their exploitation by man.In spite of this, the molecular details of cellulose biosynthesis have remained obscure in all plants. In particular, the toughness of wood cells makes it hard to isolateactive enzymes and study cellulose synthesis in trees. Functional genomics providespowerful new tools to study complex metabolic processes. In this way, 18 CesA geneshave been recently identified in the genome sequence of Populus trichocarpa.Expression profiling during wood formation has shown that four of these genesare specifically upregulated during xylogenesis and/or tension wood formation. Othergenes that follow the same expression pattern as the wood-related CesA genes encodethe putative Korrigan ortholog PttCel9A and a novel microtubule associated proteinPttMAP20. Cell suspension cultures of hybrid aspen with elevated expression of thesecondary cell wall specific PttCesA genes have been used for efficient in vitro synthesisof cellulose, which will facilitate future studies of this challenging process in trees.

  • 7.
    Djerbi, Soraya
    et al.
    KTH, Superseded Departments, Biotechnology.
    Aspeborg, Henrik
    KTH, Superseded Departments, Biotechnology.
    Nilsson, Peter
    KTH, Superseded Departments, Biotechnology.
    Blomqvist, Kristina
    KTH, Superseded Departments, Biotechnology.
    Teeri, Tuula
    KTH, Superseded Departments, Biotechnology.
    Identification and expression analysis of genes encoding putative cellulose synthases (CesA) in the hybrid aspen, Populus tremula (L.) × P. tremuloides (Michx.)2004In: Cellulose (London), ISSN 0969-0239, E-ISSN 1572-882X, Vol. 11, no 3-4, p. 301-312Article in journal (Refereed)
    Abstract [en]

    Cellulose is synthesized in plant cell walls by large membrane-bound protein complexes proposed to contain several copies of the catalytic subunit of the cellulose synthase, CesA. Here we report identification of 10 distinct CesA genes within a database of 100,000 ESTs of the hybrid aspen, Populus tremula (L.) x P. tremuloides (Michx.). Expression analyses in normal wood undergoing xylogenesis and in tension wood indicate xylem specific expression of four putative CesA isoenzymes, PttCesA1, PttCesA3-1, PttCesA3-2 and PttCesA9. Both the protein sequences and the expression profiles of PttCesA3-1 and PttCesA3-2 are very similar, and they may thus represent redundant copies of an enzyme with essentially the same function. Further, one of the generally more constitutively expressed CesA genes, PttCesA2, seems to be activated on the opposite side of a tension wood induced stem, while PttCesA6 appears to be more specific for leaf tissues. The rest of the hybrid aspen CesA genes were found to be relatively evenly expressed over the poplar tissues hereby studied.

  • 8.
    Ezcurra, Ines
    et al.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Johansson, Camilla
    KTH, School of Biotechnology (BIO), Glycoscience.
    Tamizhselvan, Prashanth
    KTH, School of Biotechnology (BIO), Glycoscience.
    Winzell, Anders
    KTH, School of Biotechnology (BIO), Glycoscience.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Glycoscience.
    An AC-type element mediates transactivation of secondary cell wall carbohydrate-active enzymes by PttMYB021, the Populus MYB46 orthologue2011In: BMC Proceedings, ISSN 1753-6561, E-ISSN 1753-6561, Vol. 5Article in journal (Refereed)
  • 9. Geisler-Lee, J.
    et al.
    Geisler, M.
    Coutinho, P. M.
    Segerman, B.
    Nishikubo, N.
    Takahashi, J.
    Aspeborg, H.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Djerbi, S.
    Master, E.
    Andersson-Gunneras, S.
    Sundberg, B.
    Karpinski, S.
    Teeri, Tuula T.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Kleczkowski, L. A.
    Henrissat, B.
    Mellerowicz, E. J.
    Poplar carbohydrate-active enzymes. Gene identification and expression analyses2006In: Plant Physiology, ISSN 0032-0889, E-ISSN 1532-2548, Vol. 140, no 3, p. 946-962Article in journal (Refereed)
    Abstract [en]

    Over 1,600 genes encoding carbohydrate-active enzymes (CAZymes) in the Populus trichocarpa (Torr.&Gray) genome were identified based on sequence homology, annotated, and grouped into families of glycosyltransferases, glycoside hydrolases, carbohydrate esterases, polysaccharide lyases, and expansins. Poplar ( Populus spp.) had approximately 1.6 times more CAZyme genes than Arabidopsis ( Arabidopsis thaliana). Whereas most families were proportionally increased, xylan and pectin-related families were underrepresented and the GT1 family of secondary metabolite-glycosylating enzymes was overrepresented in poplar. CAZyme gene expression in poplar was analyzed using a collection of 100,000 expressed sequence tags from 17 different tissues and compared to microarray data for poplar and Arabidopsis. Expression of genes involved in pectin and hemicellulose metabolism was detected in all tissues, indicating a constant maintenance of transcripts encoding enzymes remodeling the cell wall matrix. The most abundant transcripts encoded sucrose synthases that were specifically expressed in wood-forming tissues along with cellulose synthase and homologs of KORRIGAN and ELP1. Woody tissues were the richest source of various other CAZyme transcripts, demonstrating the importance of this group of enzymes for xylogenesis. In contrast, there was little expression of genes related to starch metabolism during wood formation, consistent with the preferential flux of carbon to cell wall biosynthesis. Seasonally dormant meristems of poplar showed a high prevalence of transcripts related to starch metabolism and surprisingly retained transcripts of some cell wall synthesis enzymes. The data showed profound changes in CAZyme transcriptomes in different poplar tissues and pointed to some key differences in CAZyme genes and their regulation between herbaceous and woody plants.

  • 10. Herlemann, Daniel P. R.
    et al.
    Lundin, Daniel
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Labrenz, Matthias
    Jürgens, Klaus
    Zheng, Zongli
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Glycoscience.
    Andersson, Anders F.
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Metagenomic De Novo Assembly of an Aquatic Representative of the Verrucomicrobial Class Spartobacteria2013In: mBio, ISSN 2161-2129, E-ISSN 2150-7511, Vol. 4, no 3, p. e00569-12-Article in journal (Refereed)
    Abstract [en]

    The verrucomicrobial subdivision 2 class Spartobacteria is one of the most abundant bacterial lineages in soil and has recently also been found to be ubiquitous in aquatic environments. A 16S rRNA gene study from samples spanning the entire salinity range of the Baltic Sea indicated that, in the pelagic brackish water, a phylotype of the Spartobacteria is one of the dominating bacteria during summer. Phylogenetic analyses of related 16S rRNA genes indicate that a purely aquatic lineage within the Spartobacteria exists. Since no aquatic representative from the Spartobacteria has been cultured or sequenced, the metabolic capacity and ecological role of this lineage are yet unknown. In this study, we reconstructed the genome and metabolic potential of the abundant Baltic Sea Spartobacteria phylotype by metagenomics. Binning of genome fragments by nucleotide composition and a self-organizing map recovered the near-complete genome of the organism, the gene content of which suggests an aerobic heterotrophic metabolism. Notably, we found 23 glycoside hydrolases that likely allow the use of a variety of carbohydrates, like cellulose, mannan, xylan, chitin, and starch, as carbon sources. In addition, a complete pathway for sulfate utilization was found, indicating catabolic processing of sulfated polysaccharides, commonly found in aquatic phytoplankton. The high frequency of glycoside hydrolase genes implies an important role of this organism in the aquatic carbon cycle. Spatiotemporal data of the phylotype's distribution within the Baltic Sea indicate a connection to Cyanobacteria that may be the main source of the polysaccharide substrates. IMPORTANCE The ecosystem roles of many phylogenetic lineages are not yet well understood. One such lineage is the class Spartobacteria within the Verrucomicrobia that, despite being abundant in soil and aquatic systems, is relatively poorly studied. Here we circumvented the difficulties of growing aquatic Verrucomicrobia by applying shotgun metagenomic sequencing on a water sample from the Baltic Sea. By using a method based on sequence signatures, we were able to in silico isolate genome fragments belonging to a phylotype of the Spartobacteria. The genome, which represents the first aquatic representative of this clade, encodes a diversity of glycoside hydrolases that likely allow degradation of various complex carbohydrates. Since the phylotype cooccurs with Cyanobacteria, these may be the primary producers of the carbohydrate substrates. The phylotype, which is highly abundant in the Baltic Sea during summer, may thus play an important role in the carbon cycle of this ecosystem.

  • 11. Hertzberg, M.
    et al.
    Aspeborg, H.
    Schrader, J.
    Andersson, A.
    Erlandsson, R.
    Blomqvist, K.
    Bhalerao, R.
    Uhlén, Mathias
    KTH, Superseded Departments, Biotechnology.
    Teeri, Tuula T.
    KTH, Superseded Departments, Biotechnology.
    Lundeberg, Joakim
    KTH, Superseded Departments, Biotechnology.
    Sundberg, B.
    Nilsson, Peter
    KTH, Superseded Departments, Biotechnology.
    Sandberg, G.
    A transcriptional roadmap to wood formation2001In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 98, no 25, p. 14732-14737Article in journal (Refereed)
    Abstract [en]

    The large vascular meristem of poplar trees with its highly organized secondary xylem enables the boundaries between different developmental zones to be easily distinguished. This property of wood-forming tissues allowed us to determine a unique tissue-specific transcript profile for a well defined developmental gradient. RNA was prepared from different developmental stages of xylogenesis for DNA microarray analysis by using a hybrid aspen unigene set consisting of 2,995 expressed sequence tags. The analysis revealed that the genes encoding lignin and cellulose biosynthetic enzymes, as well as a number of transcription factors and other potential regulators of xylogenesis, are under strict developmental stage-specific transcriptional regulation.

  • 12. Hertzberg, Magnus
    et al.
    Aspeborg, Henrik
    KTH, Superseded Departments, Biotechnology.
    Sievertzon, Maria
    KTH, Superseded Departments, Biotechnology.
    Nilsson, Peter
    KTH, Superseded Departments, Biotechnology.
    Sandberg, G.
    Lundeberg, Joakim
    KTH, Superseded Departments, Biotechnology.
    cDNA microarray analysis of small tissue samples using a cDNA tag target amplification protocol2001In: The Plant Journal, ISSN 0960-7412, E-ISSN 1365-313X, Vol. 25, no 5, p. 585-591Article in journal (Refereed)
    Abstract [en]

    Microarray technology is becoming an important comprehensive tool to study gene expression in plants. However, the use of this technology is limited by the large amount of sample tissue needed for microarray analysis. Generally, 50-200 mug of total RNA and 1-2 mug of mRNA is required for each hybridisation, which is equivalent to 50-100 mg of plant tissue. This requirement for large amounts of starting material severely constrains the use of microarrays for transcript profiling in specific tissues and cell types during plant development. Here we report on a robust and reliable target amplification method that enables transcript profiling from sub-mg amounts of plant tissue. Using 0.1 mug of total RNA we show that twofold expression differences are possible to distinguish with 99% confidence. We also demonstrate the application of this method in an analysis of secondary phloem development in hybrid aspen using defined tissue sections, corresponding to 2-4 cell layers with a fresh weight of similar to 0.5 mg.

  • 13. Israelsson, M.
    et al.
    Eriksson, M. E.
    Hertzberg, M.
    Aspeborg, H.
    Nilsson, Peter
    KTH, Superseded Departments, Biotechnology.
    Moritz, T.
    Changes in gene expression in the wood-forming tissue of transgenic hybrid aspen with increased secondary growth2003In: Plant Molecular Biology, ISSN 0167-4412, E-ISSN 1573-5028, Vol. 52, no 4, p. 893-903Article in journal (Refereed)
    Abstract [en]

    Transgenic lines of hybrid aspen with elevated levels of gibberellin (GA) show greatly increased numbers of xylem fibres and increases in xylem fibre length. These plants therefore provide excellent models for studying secondary growth. We have used cDNA microarry analysis to investigate how gene transcription in the developing xylem is affected by GA-induced growth. A recent investigation has shown that genes encoding lignin and cellulose biosynthetic enzymes, as well as a number of transcription factors and other potential regulators of xylogenesis, are under developmental-stage-specific transcriptional control. The present study shows that the highest transcript changes in our transgenic trees occurs in genes generally restricted to the early stages of xylogenesis, including cell division, early expansion and late expansion. The results reveal genes among those arrayed that are up-regulated with an increased xylem production, thus indicating key components in the production of wood.

  • 14.
    Kallas, Åsa M.
    et al.
    KTH, School of Biotechnology (BIO).
    Baumann, Martin J.
    KTH, School of Biotechnology (BIO).
    Fäldt, Jenny
    KTH.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO).
    Denman, Stuart
    KTH.
    Mellerowicz, Ewa J.
    Nishikubo, Nobuyushi
    Brumer, Harry
    KTH, School of Biotechnology (BIO), Glycoscience.
    Teeri, Tuula T.
    KTH, School of Biotechnology (BIO).
    Enzymatic characterization of a recombinant xyloglucan endotransglycosylase PttXET16-35 from Populus tremula x tremuloidesManuscript (Other academic)
  • 15.
    Rajangam, Alex S.
    et al.
    KTH, School of Biotechnology (BIO), Centres, Swedish Center for Biomimetic Fiber Engineering, BioMime.
    Kumar, Manoj
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Centres, Swedish Center for Biomimetic Fiber Engineering, BioMime.
    Guerriero, Gea
    KTH, School of Biotechnology (BIO), Centres, Swedish Center for Biomimetic Fiber Engineering, BioMime.
    Arvestad, Lars
    KTH, School of Computer Science and Communication (CSC), Computational Biology, CB.
    Pansri, Podjamas
    KTH, School of Biotechnology (BIO), Centres, Swedish Center for Biomimetic Fiber Engineering, BioMime.
    Brown, Christian J. L.
    KTH, School of Biotechnology (BIO), Centres, Swedish Center for Biomimetic Fiber Engineering, BioMime.
    Hober, Sophia
    KTH, School of Biotechnology (BIO), Proteomics (closed 20130101).
    Blomqvist, Kristina
    KTH, School of Biotechnology (BIO), Centres, Swedish Center for Biomimetic Fiber Engineering, BioMime.
    Divne, Christina
    KTH, School of Biotechnology (BIO), Glycoscience. KTH, School of Biotechnology (BIO), Centres, Swedish Center for Biomimetic Fiber Engineering, BioMime.
    Ezcurra, Inés
    KTH, School of Biotechnology (BIO), Glycoscience. KTH, School of Biotechnology (BIO), Centres, Swedish Center for Biomimetic Fiber Engineering, BioMime.
    Mellerowicz, Ewa
    Sundberg, Bjorn
    Bulone, Vincent
    KTH, School of Biotechnology (BIO), Glycoscience. KTH, School of Biotechnology (BIO), Centres, Swedish Center for Biomimetic Fiber Engineering, BioMime.
    Teeri, Tuula T.
    KTH, School of Biotechnology (BIO), Glycoscience. KTH, School of Biotechnology (BIO), Centres, Swedish Center for Biomimetic Fiber Engineering, BioMime.
    MAP20, a Microtubule-Associated Protein in the Secondary Cell Walls of Hybrid Aspen, Is a Target of the Cellulose Synthesis Inhibitor 2,6-Dichlorobenzonitrile2008In: Plant Physiology, ISSN 0032-0889, E-ISSN 1532-2548, Vol. 148, no 3, p. 1283-1294Article in journal (Refereed)
    Abstract [en]

    We have identified a gene, denoted PttMAP20, which is strongly up-regulated during secondary cell wall synthesis and tightly coregulated with the secondary wall-associated CESA genes in hybrid aspen (Populus tremula x tremuloides). Immunolocalization studies with affinity-purified antibodies specific for PttMAP20 revealed that the protein is found in all cell types in developing xylem and that it is most abundant in cells forming secondary cell walls. This PttMAP20 protein sequence contains a highly conserved TPX2 domain first identified in a microtubule-associated protein (MAP) in Xenopus laevis. Overexpression of PttMAP20 in Arabidopsis (Arabidopsis thaliana) leads to helical twisting of epidermal cells, frequently associated with MAPs. In addition, a PttMAP20-yellow fluorescent protein fusion protein expressed in tobacco (Nicotiana tabacum) leaves localizes to microtubules in leaf epidermal pavement cells. Recombinant PttMAP20 expressed in Escherichia coli also binds specifically to in vitro-assembled, taxol-stabilized bovine microtubules. Finally, the herbicide 2,6-dichlorobenzonitrile, which inhibits cellulose synthesis in plants, was found to bind specifically to PttMAP20. Together with the known function of cortical microtubules in orienting cellulose microfibrils, these observations suggest that PttMAP20 has a role in cellulose biosynthesis.

  • 16. Ubeda-Tomas, Susana
    et al.
    Edvardsson, Ellinor
    Eland, Cathlene
    Singh, Sunil Kumar
    Zadik, Daniel
    Aspeborg, Henrik
    Gorzsas, Andras
    Teeri, Tuula T.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Sundberg, Bjorn
    Persson, Per
    Bennett, Malcolm
    Marchant, Alan
    Genomic-assisted identification of genes involved in secondary growth in Arabidopsis utilising transcript profiling of poplar wood-forming tissues2007In: Physiologia Plantarum: An International Journal for Plant Biology, ISSN 0031-9317, E-ISSN 1399-3054, Vol. 129, no 2, p. 415-428Article in journal (Refereed)
    Abstract [en]

    Despite the importance of secondary growth in plants, relatively few genes regulating this process have been identified to date. By using data from detailed transcript profiling of the poplar wood-forming tissues, 150 genes that are differentially expressed within the zone of secondary growth were identified. In order to determine the possible function of these poplar genes, potential Arabidopsis thaliana orthologs were identified and gene knockout lines analysed. Three selection filters were used to identify the most likely orthologous genes using poplar and Arabidopsis sequence comparisons, expression profiling in secondary thickened Arabidopsis hypocotyls and global expression analysis of Arabidopsis tissues. Three genes encoding AtCSLA2 (At5g22740), the AtGUT1 GT47 glycosyltransferase (At1g27440) and a protein with no proposed function AtUNKA (At4g27435) were selected for further detailed analysis of their role in secondary growth in Arabidopsis. The presented genome-based approach using both poplar and Arabidopsis systems provides powerful means towards assigning biological functions to enzymes with poorly understood biochemical activity, such as AtCSLA2 and AtGUT1, as well as for proteins with no known function.

  • 17.
    Wang, Yang
    et al.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Azhar, Shoaib
    KTH, School of Chemical Science and Engineering (CHE), Fibre and Polymer Technology.
    Gandini, Rosaria
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Divne, Christina
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Ezcurra, Ines
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Biochemical characterization of the novel endo-β-mannanase AtMan5-2 from Arabidopsis thalianaManuscript (preprint) (Other academic)
  • 18.
    Wang, Yang
    et al.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Azhar, Shoaib
    KTH, School of Chemical Science and Engineering (CHE), Centres, Wallenberg Wood Science Center. KTH, School of Chemical Science and Engineering (CHE), Fibre and Polymer Technology.
    Gandini, Rosaria
    KTH, School of Biotechnology (BIO), Industrial Biotechnology. Karolinska Institutet, Stockholm.
    Divne, Christina
    KTH, School of Biotechnology (BIO), Industrial Biotechnology. Karolinska Institutet, Stockholm.
    Ezcurra, Ines
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Biochemical characterization of the novel endo-β-mannanase AtMan5-2 from Arabidopsis thaliana2015In: Plant Science, ISSN 0168-9452, E-ISSN 1873-2259, Vol. 241, p. 151-163Article in journal (Refereed)
    Abstract [en]

    Plant mannanases are enzymes that carry out fundamentally important functions in cell wall metabolism during plant growth and development by digesting manno-polysaccharides. In this work, the Arabidopsis mannanase 5-2 (AtMan5-2) from a previously uncharacterized subclade of glycoside hydrolase family 5 subfamily 7 (GH5_7) has been heterologously produced in Pichia pastoris. Purified recombinant AtMan5-2 is a glycosylated protein with an apparent molecular mass of 50 kDa, a pH optimum of 5.5-6.0 and a temperature optimum of 25 degrees C. The enzyme exhibits high substrate affinity and catalytic efficiency on mannan substrates with main chains containing both glucose and mannose units such as konjac glucomannan and spruce galactoglucomannan. Product analysis of manno-oligosaccharide hydrolysis shows that AtMan5-2 requires at least six substrate-binding subsites. No transglycosylation activity for the recombinant enzyme was detected in the present study. Our results demonstrate diversification of catalytic function among members in the Arabidopsis GH5_7 subfamily.

  • 19.
    Wang, Yang
    et al.
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Azhar, Shoaib
    KTH, School of Chemical Science and Engineering (CHE), Fibre and Polymer Technology.
    Gandini, Rosaria
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Divne, Christina
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Ezcurra, Ines
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Investigating the function and biochemical properties of Arabidopsis mannanase 5-6Manuscript (preprint) (Other academic)
  • 20.
    Wang, Yang
    et al.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Vilaplana, Francisco
    KTH, School of Biotechnology (BIO), Glycoscience.
    Brumer, Harry
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Glycoscience.
    Enzymatic characterization of a GH5_7 mannanase from Arabidopsis thalianaManuscript (preprint) (Other academic)
  • 21.
    Wang, Yang
    et al.
    KTH, School of Biotechnology (BIO), Glycoscience. KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Vilaplana, Francisco
    KTH, School of Biotechnology (BIO), Glycoscience. KTH, School of Chemical Science and Engineering (CHE), Centres, Wallenberg Wood Science Center.
    Brumer, Harry
    KTH, School of Biotechnology (BIO), Glycoscience.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Glycoscience. KTH, School of Biotechnology (BIO), Industrial Biotechnology.
    Enzymatic characterization of a glycoside hydrolase family 5 subfamily 7 (GH5_7) mannanase from Arabidopsis thaliana2014In: Planta, ISSN 0032-0935, E-ISSN 1432-2048, Vol. 239, no 3, p. 653-665Article in journal (Refereed)
    Abstract [en]

    Each plant genome contains a repertoire of beta-mannanase genes belonging to glycoside hydrolase family 5 subfamily 7 (GH5_7), putatively involved in the degradation and modification of various plant mannan polysaccharides, but very few have been characterized at the gene product level. The current study presents recombinant production and in vitro characterization of AtMan5-1 as a first step towards the exploration of the catalytic capacity of Arabidopsis thaliana beta-mannanase. The target enzyme was expressed in both E. coli (AtMan5-1e) and P. pastoris (AtMan5-1p). The main difference between the two forms was a higher observed thermal stability for AtMan5-1p, presumably due to glycosylation of that particular variant. AtMan5-1 displayed optimal activity at pH 5 and 35 A degrees C and hydrolyzed polymeric carob galactomannan, konjac glucomannan, and spruce galactoglucomannan as well as oligomeric mannopentaose and mannohexaose. However, the galactose-rich and highly branched guar gum was not as efficiently degraded. AtMan5-1 activity was enhanced by Co2+ and inhibited by Mn2+. The catalytic efficiency values for carob galactomannan were 426.8 and 368.1 min(-1) mg(-1) mL for AtMan5-1e and AtMan5-1p, respectively. Product analysis of AtMan5-1p suggested that at least five substrate-binding sites were required for manno-oligosaccharide hydrolysis, and that the enzyme also can act as a transglycosylase.

  • 22.
    Winzell, Anders
    et al.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Glycoscience.
    Wang, Yucheng
    KTH, School of Biotechnology (BIO), Glycoscience.
    Ezcurra, Ines
    KTH, School of Biotechnology (BIO), Glycoscience.
    Conserved CA-rich motifs in gene promoters of Pt x tMYB021-responsive secondary cell wall carbohydrate-active enzymes in Populus2010In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 394, no 3, p. 848-853Article in journal (Refereed)
    Abstract [en]

    In order to understand gene regulation during wood formation, we cloned a MYB46-like gene in hybrid aspen. Populus tremula x tremuloides, called PtxtMYB021 Phylogenetic and paired identity analysis of MYB46-like genes in Populus and Arabidopsis reveals relationships between paralogous pairs of Populus MYB46-like proteins and their Arabidopsis counterparts MYB46 and MYB83, and suggest that PtxtMYB021 is the ortholog of MYB46 Ptxt-MYB021 is expressed mainly in xylem tissues, and transiently expressed PtxtMYB46 transactivates gene promoters of xylan-active CAZymes GT43A, GT43B and Xyn10A Analysis of conserved motifs within these promoters identify the sequence CCACCAAC, called ACTYP, which is similar to the AC elements mediating transactivation by MYB transcription factors during lignin biosynthesis Further analysis by Motif Finder identifies four 6 bp CA-rich motifs overlapping ACTYP, and we show that these motifs are enriched in xylem-specific promoters We propose that AC-type regulatory elements mediate xylem-specific MYB46-dependent expression of secondary cell wall carbohydrate-active enzymes (CAZymes), besides activating gene expression of lignin biosynthesis enzymes.

  • 23.
    Winzell, Anders
    et al.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Glycoscience.
    Wang, Yucheng
    KTH, School of Biotechnology (BIO), Glycoscience.
    Ezcurra, Ines
    KTH, School of Biotechnology (BIO), Glycoscience.
    Molecular cloning and functional characterization of Pt×tMYB021, a MYB46-like transcription factor in PopulusManuscript (preprint) (Other academic)
  • 24.
    Winzell, Anders
    et al.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Guerriero, Gea
    KTH, School of Biotechnology (BIO), Glycoscience.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Glycoscience.
    Wang, Yiqiang
    KTH, School of Biotechnology (BIO), Glycoscience.
    Rajangam, Alex S.
    Teeri, Tuula T.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Ezcurra, Inés
    KTH, School of Biotechnology (BIO), Glycoscience.
    Biochemical characterization of family 43 glycosyltransferases in the Populus xylem: challenges and prospects2010In: Plant Biotechnology, ISSN 1342-4580, Vol. 27, no 3, p. 283-288Article in journal (Refereed)
    Abstract [en]

    Wood formation is a biological process of great economical importance. Genes active during the secondary cellwall formation of wood fibers from Populus tremulatremuloides were previously identified by expression profilingthrough microarray analyses. A number of these genes encode glycosyltransferases (GTs) with unknown substratespecificities. Here we report heterologous expression of one of these enzymes, PttGT43A, a putative IRREGULARXYLEM9 (IRX9) homologue. Expression trials in Pichia pastoris and insect cells revealed very low levels of accumulationof immunoreactive PttGT43A, whereas transient expression in Nicotiana benthamiana leaves by Agrobacterium infiltration(agroinfiltration) using a viral vector produced substantial amounts of protein that mostly precipitated in the crude pellet.Agroinfiltration induced weak endogenous xylosyltransferase activity in microsomal extracts, and transient PttGT43Aexpression further increased this activity, albeit only to low levels. PttGT43A may be inactive as an individual subunit,requiring complex formation with unknown partners to display enzymatic activity. Our results suggest that transient coexpressionin leaves of candidate subunit GTs may provide a viable approach for formation of an active xylanxylosyltransferase enzymatic complex.

  • 25.
    Winzell, Anders
    et al.
    KTH, School of Biotechnology (BIO).
    Rajangam, Alex
    KTH, School of Biotechnology (BIO).
    Arvestad, Lars
    KTH, School of Computer Science and Communication (CSC).
    Filling, Charlotta
    KTH, School of Biotechnology (BIO).
    Divine, Christina
    KTH, School of Biotechnology (BIO).
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO).
    Master, Emma R.
    KTH, School of Biotechnology (BIO).
    Teeri, Tuula T.
    KTH, School of Biotechnology (BIO).
    Sequence Analysis and Recombinant Expression of Family 43 GlycosyltransferasesManuscript (preprint) (Other academic)
  • 26.
    Winzell, Anders
    et al.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Ratke, Christine
    Naumann, Marcel
    Wang, Yucheng
    KTH, School of Biotechnology (BIO), Glycoscience.
    Aspeborg, Henrik
    KTH, School of Biotechnology (BIO), Glycoscience.
    Teeri, Tuula T.
    KTH, School of Biotechnology (BIO), Glycoscience.
    Mellerowicz, Ewa
    Ezcurra, Ines
    KTH, School of Biotechnology (BIO), Glycoscience.
    Family 43 glycosyltransferases in Populus and Arabidopsis: phylogeny and expression analysisManuscript (preprint) (Other academic)
1 - 26 of 26
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