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Publications (10 of 27) Show all publications
Reichenbach, T., Kalyani, D., Gandini, R., Svartström, O., Aspeborg, H. & Divne, C. (2018). Structural and biochemical characterization of the Cutibacterium acnes exo-β-1,4-mannosidase that targets the N-glycan core of host glycoproteins.. PLoS ONE, 13(9), Article ID e0204703.
Open this publication in new window or tab >>Structural and biochemical characterization of the Cutibacterium acnes exo-β-1,4-mannosidase that targets the N-glycan core of host glycoproteins.
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2018 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 13, no 9, article id e0204703Article in journal (Refereed) Published
Abstract [en]

Commensal and pathogenic bacteria have evolved efficient enzymatic pathways to feed on host carbohydrates, including protein-linked glycans. Most proteins of the human innate and adaptive immune system are glycoproteins where the glycan is critical for structural and functional integrity. Besides enabling nutrition, the degradation of host N-glycans serves as a means for bacteria to modulate the host's immune system by for instance removing N-glycans on immunoglobulin G. The commensal bacterium Cutibacterium acnes is a gram-positive natural bacterial species of the human skin microbiota. Under certain circumstances, C. acnes can cause pathogenic conditions, acne vulgaris, which typically affects 80% of adolescents, and can become critical for immunosuppressed transplant patients. Others have shown that C. acnes can degrade certain host O-glycans, however, no degradation pathway for host N-glycans has been proposed. To investigate this, we scanned the C. acnes genome and were able to identify a set of gene candidates consistent with a cytoplasmic N-glycan-degradation pathway of the canonical eukaryotic N-glycan core. We also found additional gene sequences containing secretion signals that are possible candidates for initial trimming on the extracellular side. Furthermore, one of the identified gene products of the cytoplasmic pathway, AEE72695, was produced and characterized, and found to be a functional, dimeric exo-β-1,4-mannosidase with activity on the β-1,4 glycosidic bond between the second N-acetylglucosamine and the first mannose residue in the canonical eukaryotic N-glycan core. These findings corroborate our model of the cytoplasmic part of a C. acnes N-glycan degradation pathway.

National Category
Biological Sciences
Identifiers
urn:nbn:se:kth:diva-235707 (URN)10.1371/journal.pone.0204703 (DOI)000445907400093 ()30261037 (PubMedID)2-s2.0-85053860322 (Scopus ID)
Funder
Swedish Research Council, 20135717Swedish Research Council Formas, 20131741; 2012-1513; 2014-176
Note

QC 20181030

Available from: 2018-10-02 Created: 2018-10-02 Last updated: 2019-01-07Bibliographically approved
Wang, Y., Azhar, S., Gandini, R., Divne, C., Ezcurra, I. & Aspeborg, H. (2015). Biochemical characterization of the novel endo-β-mannanase AtMan5-2 from Arabidopsis thaliana. Plant Science, 241, 151-163
Open this publication in new window or tab >>Biochemical characterization of the novel endo-β-mannanase AtMan5-2 from Arabidopsis thaliana
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2015 (English)In: Plant Science, ISSN 0168-9452, E-ISSN 1873-2259, Vol. 241, p. 151-163Article in journal (Refereed) Published
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.

Place, publisher, year, edition, pages
Elsevier, 2015
Keywords
Glycoside hydrolase, GH5, endo-β-1, 4-Mannan hydrolase, Cell wall, Mannan polysaccharides/oligosaccharides
National Category
Biological Sciences
Research subject
Biotechnology
Identifiers
urn:nbn:se:kth:diva-178198 (URN)10.1016/j.plantsci.2015.10.002 (DOI)000367487500015 ()2-s2.0-84945291912 (Scopus ID)
Funder
Swedish Foundation for Strategic Research VINNOVASwedish Research Council Formas
Note

QC 20160104. QC 20160201

Available from: 2015-12-07 Created: 2015-12-07 Last updated: 2017-12-01Bibliographically approved
Wang, Y., Vilaplana, F., Brumer, H. & Aspeborg, H. (2014). Enzymatic characterization of a glycoside hydrolase family 5 subfamily 7 (GH5_7) mannanase from Arabidopsis thaliana. Planta, 239(3), 653-665
Open this publication in new window or tab >>Enzymatic characterization of a glycoside hydrolase family 5 subfamily 7 (GH5_7) mannanase from Arabidopsis thaliana
2014 (English)In: Planta, ISSN 0032-0935, E-ISSN 1432-2048, Vol. 239, no 3, p. 653-665Article in journal (Refereed) Published
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.

Keywords
GH5_7, beta-Mannanase, Glycoside hydrolase, Mannan, Plant cell wall, Carbohydrates
National Category
Plant Biotechnology
Identifiers
urn:nbn:se:kth:diva-143702 (URN)10.1007/s00425-013-2005-y (DOI)000331648500010 ()2-s2.0-84916929268 (Scopus ID)
Funder
Swedish Foundation for Strategic Research FormasSwedish Research Council
Note

QC 20140331

Available from: 2014-03-31 Created: 2014-03-27 Last updated: 2017-12-05Bibliographically approved
Herlemann, D. P. R., Lundin, D., Labrenz, M., Jürgens, K., Zheng, Z., Aspeborg, H. & Andersson, A. F. (2013). Metagenomic De Novo Assembly of an Aquatic Representative of the Verrucomicrobial Class Spartobacteria. mBio, 4(3), e00569-12
Open this publication in new window or tab >>Metagenomic De Novo Assembly of an Aquatic Representative of the Verrucomicrobial Class Spartobacteria
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2013 (English)In: mBio, ISSN 2161-2129, E-ISSN 2150-7511, Vol. 4, no 3, p. e00569-12-Article in journal (Refereed) Published
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.

Keywords
Multiple sequence alignment, central baltic sea, genome sequence, gen. Nov., bacterioplankton communities, phylum verrucomicrobia, phylogenetic diversity, substrate-specificity, bacterial communities, microbial communities
National Category
Microbiology
Identifiers
urn:nbn:se:kth:diva-125770 (URN)10.1128/mBio.00569-12 (DOI)000321187400041 ()2-s2.0-84879566132 (Scopus ID)
Funder
Swedish Research CouncilFormasScience for Life Laboratory - a national resource center for high-throughput molecular bioscience
Note

QC 20130814

Available from: 2013-08-14 Created: 2013-08-13 Last updated: 2017-12-06Bibliographically approved
Aspeborg, H., Coutinho, P. M., Wang, Y., Brumer, H. & Henrissat, B. (2012). Evolution, substrate specificity and subfamily classification of glycoside hydrolase family 5 (GH5). BMC Evolutionary Biology, 12(1), 186
Open this publication in new window or tab >>Evolution, substrate specificity and subfamily classification of glycoside hydrolase family 5 (GH5)
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2012 (English)In: BMC Evolutionary Biology, ISSN 1471-2148, E-ISSN 1471-2148, Vol. 12, no 1, p. 186-Article in journal (Refereed) Published
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.

Keywords
Protein evolution, Enzyme evolution, Functional prediction, Glycogenomics, Glycoside hydrolase family 5, Phylogenetic analysis, Subfamily classification
National Category
Bioinformatics and Systems Biology
Identifiers
urn:nbn:se:kth:diva-109742 (URN)10.1186/1471-2148-12-186 (DOI)000312732100001 ()22992189 (PubMedID)2-s2.0-84866500048 (Scopus ID)
Funder
Swedish Research Council FormasSwedish Foundation for Strategic Research
Note

QC 20130201

Available from: 2013-01-08 Created: 2013-01-08 Last updated: 2017-12-06Bibliographically approved
Ezcurra, I., Johansson, C., Tamizhselvan, P., Winzell, A. & Aspeborg, H. (2011). An AC-type element mediates transactivation of secondary cell wall carbohydrate-active enzymes by PttMYB021, the Populus MYB46 orthologue. BMC Proceedings, 5
Open this publication in new window or tab >>An AC-type element mediates transactivation of secondary cell wall carbohydrate-active enzymes by PttMYB021, the Populus MYB46 orthologue
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2011 (English)In: BMC Proceedings, ISSN 1753-6561, E-ISSN 1753-6561, Vol. 5Article in journal (Refereed) Published
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:kth:diva-51358 (URN)10.1186/1753-6561-5-S7-O40 (DOI)
Note

QC 20111227

Available from: 2011-12-12 Created: 2011-12-12 Last updated: 2017-12-08Bibliographically approved
Winzell, A., Guerriero, G., Aspeborg, H., Wang, Y., Rajangam, A. S., Teeri, T. T. & Ezcurra, I. (2010). Biochemical characterization of family 43 glycosyltransferases in the Populus xylem: challenges and prospects. Paper presented at Colloquium on Green Chemistry in Plants and Microorganisms Japanese Soc Promot Sci, Stockholm, SWEDEN, MAY 25, 2009. Plant Biotechnology, 27(3), 283-288
Open this publication in new window or tab >>Biochemical characterization of family 43 glycosyltransferases in the Populus xylem: challenges and prospects
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2010 (English)In: Plant Biotechnology, ISSN 1342-4580, Vol. 27, no 3, p. 283-288Article in journal (Refereed) Published
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.

Keywords
GT43 glycosyltransferase, IRX9, populus xylem, xylan, xylosyltransferase.
National Category
Botany
Identifiers
urn:nbn:se:kth:diva-14265 (URN)000280085800010 ()2-s2.0-77955322528 (Scopus ID)
Conference
Colloquium on Green Chemistry in Plants and Microorganisms Japanese Soc Promot Sci, Stockholm, SWEDEN, MAY 25, 2009
Funder
Swedish Foundation for Strategic Research
Note

QC 20110404

Available from: 2010-07-29 Created: 2010-07-29 Last updated: 2016-04-18Bibliographically approved
Winzell, A., Aspeborg, H., Wang, Y. & Ezcurra, I. (2010). Conserved CA-rich motifs in gene promoters of Pt x tMYB021-responsive secondary cell wall carbohydrate-active enzymes in Populus. Biochemical and Biophysical Research Communications - BBRC, 394(3), 848-853
Open this publication in new window or tab >>Conserved CA-rich motifs in gene promoters of Pt x tMYB021-responsive secondary cell wall carbohydrate-active enzymes in Populus
2010 (English)In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 394, no 3, p. 848-853Article in journal (Refereed) Published
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.

Keywords
Populus, Xylem, Xylan, Transcription factor, MYB46, Regulatory elements
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:kth:diva-28291 (URN)10.1016/j.bbrc.2010.03.101 (DOI)000276785900070 ()2-s2.0-77950492955 (Scopus ID)
Funder
Formas
Note
QC 20110113Available from: 2011-01-13 Created: 2011-01-12 Last updated: 2017-12-11Bibliographically approved
Rajangam, A. S., Kumar, M., Aspeborg, H., Guerriero, G., Arvestad, L., Pansri, P., . . . Teeri, T. T. (2008). MAP20, a Microtubule-Associated Protein in the Secondary Cell Walls of Hybrid Aspen, Is a Target of the Cellulose Synthesis Inhibitor 2,6-Dichlorobenzonitrile. Plant Physiology, 148(3), 1283-1294
Open this publication in new window or tab >>MAP20, a Microtubule-Associated Protein in the Secondary Cell Walls of Hybrid Aspen, Is a Target of the Cellulose Synthesis Inhibitor 2,6-Dichlorobenzonitrile
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2008 (English)In: Plant Physiology, ISSN 0032-0889, E-ISSN 1532-2548, Vol. 148, no 3, p. 1283-1294Article in journal (Refereed) Published
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.

Keywords
carbohydrate-active enzymes, plant functional genomics, kinesin-like, protein, cortical microtubules, plasma-membrane, wood formation, arabidopsis-thaliana, conifer tracheids, angiosperm trees, urea solution
National Category
Natural Sciences
Identifiers
urn:nbn:se:kth:diva-17951 (URN)10.1104/pp.108.121913 (DOI)000260719500010 ()2-s2.0-57749101097 (Scopus ID)
Note

QC 20100525

Available from: 2010-08-05 Created: 2010-08-05 Last updated: 2017-12-12Bibliographically approved
Aspeborg, H., Bhalerao, R., Hertzberg, M., Johansson, K., Johnsson, P., Karlsson, A., . . . Wallbäcks, L. (2008). Vegetabile material, plants and a method of producing a plant having altered lignin properties. WO 2010062240A1.
Open this publication in new window or tab >>Vegetabile material, plants and a method of producing a plant having altered lignin properties
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2008 (English)Patent (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.

National Category
Plant Biotechnology
Identifiers
urn:nbn:se:kth:diva-70451 (URN)
Patent
WO 2010062240A1
Note

QS 2015

Available from: 2012-01-30 Created: 2012-01-30 Last updated: 2015-01-28Bibliographically approved
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0002-8576-4370

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