Browsing by Subject "OLIGOSACCHARIDES"

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  • Razeq, Fakhria M.; Jurak, Edita; Stogios, Peter J.; Yan, Ruoyu; Tenkanen, Maija; Kabel, Mirjam A.; Wang, Weijun; Master, Emma R. (2018)
    Background: Acetylated 4-O-(methyl) glucuronoxylan (GX) is the main hemicellulose in deciduous hardwood, and comprises a beta-(1 -> 4)-linked xylopyranosyl (Xylp) backbone substituted by both acetyl groups and alpha-(1 -> 2)-linked 4-O-methylglucopyranosyluronic acid (MeGlcpA). Whereas enzymes that target singly acetylated Xylp or doubly 2,3-O-acetyl-Xylp have been well characterized, those targeting (2-O-MeGlcpA) 3-O-acetyl-Xylp structures in glucuronoxylan have remained elusive. Results: An unclassified carbohydrate esterase (FjoAcXE) was identified as a protein of unknown function from a polysaccharide utilization locus (PUL) otherwise comprising carbohydrate-active enzyme families known to target xylan. FjoAcXE was shown to efficiently release acetyl groups from internal (2-O-MeGlcpA) 3-O-acetyl-Xylp structures, an activity that has been sought after but lacking in known carbohydrate esterases. FjoAcXE action boosted the activity of alpha-glucuronidases from families GH67 and GH115 by five and nine times, respectively. Moreover, FjoAcXE activity was not only restricted to GX, but also deacetylated (3-O-Araf)2-O-acetyl-Xylp of feruloylated xylooligomers, confirming the broad substrate range of this new carbohydrate esterase. Conclusion: This study reports the discovery and characterization of the novel carbohydrate esterase, FjoAcXE. In addition to cleaving singly acetylated Xylp, and doubly 2,3-O-acetyl-Xylp, FjoAcXE efficiently cleaves internal 3-O-acetyl-Xylp linkages in (2-O-MeGlcpA)3-O-acetyl-Xylp residues along with densely substituted and branched xylooligomers; activities that until now were missing from the arsenal of enzymes required for xylan conversion.
  • Hautala, Laura C.; Pang, Poh-Choo; Antonopoulos, Aristotelis; Pasanen, Annukka; Lee, Cheuk-Lun; Chiu, Philip C. N.; Yeung, William S. B.; Loukovaara, Mikko; Bützow, Ralf; Haslam, Stuart M.; Dell, Anne; Koistinen, Hannu (2020)
    Glycodelin is a major glycoprotein expressed in reproductive tissues, like secretory and decidualized endometrium. It has several reproduction related functions that are dependent on specific glycosylation, but it has also been found to drive differentiation of endometrial carcinoma cells toward a less malignant phenotype. Here we aimed to elucidate whether the glycosylation and function of glycodelin is altered in endometrial carcinoma as compared with a normal endometrium. We carried out glycan structure analysis of glycodelin expressed in HEC-1B human endometrial carcinoma cells (HEC-1B Gd) by mass spectrometry glycomics strategies. Glycans of HEC-1B Gd were found to comprise a typical mixture of high-mannose, hybrid, and complex-type N-glycans, often containing undecorated LacNAc (Gal beta 1-4GlcNAc) antennae. However, several differences, as compared with previously reported glycan structures of normal human decidualized endometrium-derived glycodelin isoform, glycodelin-A (GdA), were also found. These included a lower level of sialylation and more abundant poly-LacNAc antennae, some of which are fucosylated. This allowed us to select lectins that showed different binding to these classes of glycodelin. Despite the differences in glycosylation between HEC-1B Gd and GdA, both showed similar inhibitory activity on trophoblast cell invasion and peripheral blood mononuclear cell proliferation. For the detection of cancer associated glycodelin, we established a novel in situ proximity-ligation based histochemical staining method using a specific glycodelin antibody and UEAI lectin. We found that the UEAI reactive glycodelin was abundant in endometrial carcinoma, but virtually absent in normal endometrial tissue even when glycodelin was strongly expressed. In conclusion, we established a histochemical staining method for the detection of endometrial carcinoma-associated glycodelin and showed that this specific glycodelin is exclusively expressed in cancer, not in normal endometrium. Similar methods can be used for studies of other glycoproteins. Glycodelin is a major endometrial glycoprotein. The authors analyzed glycan structures of endometrial carcinoma associated glycodelin and established a novel glycodelin-glycoform specific histochemical staining method. With this, they showed that glycodelin is differentially glycosylated in endometrial carcinoma tissue, as compared to normal endometrium, representing a neoantigen with potential clinical applications.
  • Gomez, Marta; Moles, Laura; Espinosa-Martos, Irene; Bustos, Gerardo; de Vos, Willem M.; Fernandez, Leonides; Rodriguez, Juan M.; Fuentes, Susana; Jimenez, Esther (2017)
    An abnormal colonization pattern of the preterm gut may affect immune maturation and exert a long-term influence on the intestinal bacterial composition and host health. However, follow-up studies assessing the evolution of the fecal microbiota of infants that were born preterm are very scarce. In this work, the bacterial compositions of fecal samples, obtained from sixteen 2-year-old infants were evaluated using a phylogenetic microarray; subsequently, the results were compared with those obtained in a previous study from samples of meconium and feces collected from the same infants while they stayed in the neonatal intensive care unit (NICU). In parallel, the concentration of a wide range of cytokines, chemokines, growth factors and immunoglobulins were determined in meconium and fecal samples. Globally, a higher bacterial diversity and a lower interindividual variability were observed in 2-year-olds' feces, when compared to the samples obtained during their first days of life. Hospital-associated fecal bacteria, that were dominant during the NICU stay, seemed to be replaced, two years later, by genera, which are usually predominant in the healthy adult microbiome. The immune profile of the meconium and fecal samples differed, depending on the sampling time, showing different immune maturation statuses of the gut.
  • Trouvelot, Sophie; Héloir, Marie-Claire; Poinssot, Benoît; Gauthier, Adrien; Paris, Franck; Guillier, Christelle; Combier, Maud; Tdra, Lucie; Daire, Xavier; Adrian, Marielle (2014)
    Increasing interest is devoted to carbohydrates for their roles in plant immunity. Some of them are elicitors of plant defenses whereas other ones act as signaling molecules in a manner similar to phytohormones. This review first describes the main classes of carbohydrates associated to plant immunity, their role and mode of action. More precisely, the state of the art about perception of “PAMP, MAMP and DAMP type” oligosaccharides is presented and examples of induced defense events are provided. A particular attention is paid to the structure / activity relationships of these compounds. The role of sugars as signaling molecules, especially in plant microbe interactions, is also presented. Secondly, the potentialities and limits of foliar sprays of carbohydrates to stimulate plant immunity for crop protection against diseases are discussed, with focus on the roles of the leaf cuticle and phyllosphere microflora.
  • Dilokpimol, Adiphol; Mäkelä, Miia R.; Mansouri, Sadegh; Belova, Olga; Waterstraat, Martin; Bunzel, Mirko; de Vries, Ronald P.; Hilden, Kristiina S. (2017)
    A feruloyl esterase (FAE) from Aspergillus niger N402, FaeC was heterologously produced in Pichia pastoris X-33 in a yield of 10 mg/L. FaeC was most active at pH 7.0 and 50 degrees C, and showed broad substrate specificity and catalyzed the hydrolysis of methyl 3,4-dimethoxycinnamate, ethyl ferulate, methyl ferulate, methyl p-coumarate, ethyl coumarate, methyl sinapate, and methyl caffeate. The enzyme released both ferulic acid and p-coumaric acid from wheat arabinoxylan and sugar beet pectin (up to 3 mg/g polysaccharide), and acted synergistically with a commercial xylanase increasing the release of ferulic acid up to six-fold. The expression of faeC increased over time in the presence of feruloylated polysaccharides. Cinnamic, syringic, caffeic, vanillic and ferulic acid induced the expression of faeC. Overall expression of faeC was very low in all tested conditions, compared to two other A. niger FAE encoding genes, faeA and faeB. Our data showed that the fae genes responded differently towards the feruloylated polysaccharides and tested monomeric phenolic compounds suggesting that the corresponding FAE isoenzymes may target different substrates in a complementary manner. This may increase the efficiency of the degradation of diverse plant biomass. (C) 2017 Elsevier B.V. All rights reserved.
  • Dilokpimol, Adiphol; Mäkelä, Miia Riitta; Varriale, Simona; Zhou, Miaomiao; Cerullo, Gabriella; Gidijala, Loknath; Hinkka, Harri Tapio; Brás, Joana L.A.; Jütten, Peter; Piechot, Alexander; Verhaert, Raymond; Hilden, Sari Kristiina; Faraco, Vincenza; de Vries, Ronald (2018)
    Feruloyl esterases (FAEs) are a diverse group of enzymes that specifically catalyze the hydrolysis of ester bonds between a hydroxycinnamic (e.g. ferulic) acid and plant poly- or oligosaccharides. FAEs as auxiliary enzymes significantly assist xylanolytic and pectinolytic enzymes in gaining access to their site of action during biomass saccharification for biofuel and biochemical production. A limited number of FAEs have been functionally characterized compared to over 1000 putative fungal FAEs that were recently predicted by similarity-based genome mining, which divided phylogenetically into different subfamilies (SFs). In this study, 27 putative and six characterized FAEs from both ascomycete and basidiomycete fungi were selected and heterologously expressed in Pichia pastoris and the recombinant proteins biochemically characterized to validate the previous genome mining and phylogenetical grouping and to expand the information on activity of fungal FAEs. As a result, 20 enzymes were shown to possess FAE activity, being active towards pNP-ferulate and/or methyl hydroxycinnamate substrates, and covering 11 subfamilies. Most of the new FAEs showed activities comparable to those of previously characterized fungal FAEs.
  • Antonopoulou, Io; Dilokpimol, Adiphol; Iancu, Laura; Mäkelä, Miia R.; Varriale, Simona; Cerullo, Gabriella; Huttner, Silvia; Uthoff, Stefan; Juetten, Peter; Piechot, Alexander; Steinbuechel, Alexander; Olsson, Lisbeth; Faraco, Vincenza; Hilden, Kristiina S.; de Vries, Ronald P.; Rova, Ulrika; Christakopoulos, Paul (2018)
    Twenty-eight fungal feruloyl esterases (FAEs) were evaluated for their synthetic abilities in a ternary system of n-hexane: t-butanol: 100 mM MOPS-NaOH pH 6.0 forming detergentless microemulsions. Five main derivatives were synthesized, namely prenyl ferulate, prenyl caffeate, butyl ferulate, glyceryl ferulate, and l-arabinose ferulate, offering, in general, higher yields when more hydrophilic alcohol substitutions were used. Acetyl xylan esterase-related FAEs belonging to phylogenetic subfamilies (SF) 5 and 6 showed increased synthetic yields among tested enzymes. In particular, it was shown that FAEs belonging to SF6 generally transesterified aliphatic alcohols more efficiently while SF5 members preferred bulkier l-arabinose. Predicted surface properties and structural characteristics were correlated with the synthetic potential of selected tannase-related, acetyl-xylan-related, and lipase-related FAEs (SF1-2, -6, -7 members) based on homology modeling and small molecular docking simulations.