| 1. |
- Aspeborg, Henrik, et al.
(författare)
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Carbohydrate-active enzymes involved in the secondary cell wall biogenesis in hybrid aspen
- 2005
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 0032-0889 .- 1532-2548. ; 137:3, s. 983-997
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Tidskriftsartikel (refereegranskat)abstract
- 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.
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| 2. |
- Henriksson, H., et al.
(författare)
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N-linked glycosylation of native and recombinant cauliflower xyloglucan endotransglycosylase 16A
- 2003
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Ingår i: Biochemical Journal. - : Portland Press Ltd.. - 0264-6021 .- 1470-8728. ; 375, s. 61-73
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Tidskriftsartikel (refereegranskat)abstract
- The gene encoding a XET (xyloglucan endotransglycosylase) from cauliflower (Brassica oleracea var. botrytis) florets has been cloned and sequenced. Sequence analysis indicated a high degree of similarity to other XET enzymes belonging to glycosyl hydrolase family 16 (GH16). In addition to the conserved GH16 catalytic sequence motif EIDFE, there exists one potential N-linked glycosylation site. which is also highly conserved in XET enzymes from this family. Purification of the corresponding protein from extracts of cauliflower florets allowed the fractionation of a single, pure glycoform. which was analysed by MS techniques. Accurate protein mass determination following the enzymic deglycosylation of this glycoform indicated the presence of a high-mannose-type glycan of the general structure GlcNAc(2)Man(6). LC/MS and MS/MS (tandem MS) analysis provided supporting evidence for this structure and confirmed that the glycosylation site (underlined) was situated close to the predicted catalytic residues in the conserved sequence YLSSTNNEHDEIDFEFLGNRTGQPVILQTNVFTGGK. Heterologous expression in Pichia pastoris produced a range of protein glycoforms, which were, on average, more highly mannosylated than the purified native enzyme. This difference in glycosylation did not influence the apparent enzymic activity of the enzyme significantly. However, the removal of high-mannose glycosylation in recombinant cauliflower XET by endoglycosidase H, quantified by electrospray-ionization MS, caused a 40 % decrease in the transglycosylation activity of the enzyme. No hydrolytic activity was detected in native or heterologously expressed BobXET16A, even when almost completely deglycosylated.
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| 3. |
- Wang, W. J., et al.
(författare)
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Biochemical and Structural Characterization of a Five-domain GH115-Glucuronidase from the Marine Bacterium Saccharophagus degradans 2-40(T)
- 2016
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 291:27, s. 14120-14133
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Tidskriftsartikel (refereegranskat)abstract
- Glucuronic acid (GlcAp) and/or methylglucuronic acid (MeGlcAp) decorate the major forms of xylan in hardwood and coniferous softwoods as well as many cereal grains. Accordingly, the complete utilization of glucuronoxylans or conversion to sugar precursors requires the action of main chain xylanases as well as -glucuronidases that release the - (12)-linked (Me)GlcAp side groups. Herein, a family GH115 enzymefrom the marine bacterium Saccharophagus degradans 2-40(T), SdeAgu115A, demonstrated activity toward glucuronoxylan and oligomers thereof with preference toward MeGlcAp linked to internal xylopyranosyl residues. Unique biochemical characteristics of NaCl activation were also observed. The crystal structure of SdeAgu115A revealed a five-domain architecture, with an additional insertion C+ domain that had significant impact on the domain arrangement of SdeAgu115A monomer and its dimerization. The participation of domain C+ in substrate binding was supported by reduced substrate inhibition upon introducing W773A, W689A, and F696A substitutions within this domain. In addition to Asp-335, the catalytic essentiality of Glu-216 was revealed by site-specific mutagenesis. A primary sequence analysis suggested that the SdeAgu115A architecture is shared by more than half of GH115 members, thus defining a distinct archetype for GH115 enzymes.
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| 4. |
- Wigren, Julia, et al.
(författare)
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At-home sampling to meet geographical challenges for serological assessment of SARS-CoV-2 exposure in a rural region of northern Sweden, March to May 2021 : a retrospective cohort study
- 2023
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Ingår i: Eurosurveillance. - : European Centre for Disease Control and Prevention (ECDC). - 1025-496X .- 1560-7917. ; 28:13
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Tidskriftsartikel (refereegranskat)abstract
- Background: The current SARS-CoV-2 pandemic has highlighted a need for easy and safe blood sampling in combination with accurate serological methodology. Venipuncture for testing is usually performed by trained staff at healthcare centres. Long travel distances to healthcare centres in rural regions may introduce a bias of testing towards relatively large communities with closer access. Rural regions are therefore often not represented in population-based data.Aim: The aim of this retrospective cohort study was to develop and implement a strategy for at-home testing in a rural region of Sweden during spring 2021, and to evaluate its role to provide equal health care for its inhabitants.Methods: We developed a sensitive method to measure antibodies to the S-protein of SARS-CoV-2 and optimised this assay for clinical use together with a strategy of at-home capillary blood sampling.Results: We demonstrated that our ELISA gave comparable results after analysis of capillary blood or serum from SARS-CoV-2-experienced individuals. We demonstrated stability of the assay under conditions that reflected temperature and humidity during winter or summer. By assessment of capillary blood samples from 4,122 individuals, we could show both feasibility of the strategy and that implementation shifted the geographical spread of testing in favour of rural areas.Conclusion: Implementation of at-home sampling enabled citizens living in remote rural areas access to centralised and sensitive laboratory antibody tests. The strategy for testing used here could therefore enable disease control authorities to get rapid access to information concerning immunity to infectious diseases, even across vast geographical distance.
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