279 results found
Peswani AR, Narkpuk J, Krueger A, et al., 2022, Novel constructs and 1-step chromatography protocols for the production of Porcine Circovirus 2d (PCV2d) and Circovirus 3 (PCV3) subunit vaccine candidates, Food and Bioproducts Processing, Vol: 131, Pages: 125-135, ISSN: 0960-3085
Porcine circovirus type 2 (PCV2) has been a major problem for the pig production industry worldwide for decades. While the majority of commercially available vaccines are based on the original PCV2a genotype, the current dominant genotype is PCV2d. The notable differences between genotypes could lead to incomplete cross-protection. Moreover, most current subunit PCV2 vaccines are generated from expensive insect cell culture technology. In this work, we present a new workflow for production of an updated and relatively inexpensive PCV2d vaccine candidate. After expression in fed-batch Escherichia coli fermentation systems with a simple one-step ion-exchange chromatography purification protocol, the yield of purified PCV2d-based antigen reached over 1 g per litre bacterial culture. Using similar procedures, we also demonstrated even higher PCV2d-based antigen yields from a chimeric PCV2d-PCV3 capsid construct, which is cleaved during fermentation to release PCV2d- and PCV3-related polypeptides. Although the PCV2d-based recombinant protein from this protocol did not form viral-like particles as analysed by size-exclusion chromatography, it could effectively induce capsid-specific and PCV2d-neutralising antibodies in immunised animals, indicating significant potential as a new vaccine candidate that can be easily manufactured at commercial scale.
Kotidis P, Marbiah M, Donini R, et al., 2022, Rapid Antibody Glycoengineering in CHO Cells Via RNA Interference and CGE-LIF N-Glycomics., Methods Mol Biol, Vol: 2370, Pages: 147-167
The impact of the glycan distribution on the in vivo function and half-life of monoclonal antibodies has long motivated the genetic engineering of producer cells to achieve structures that enhance efficacy, safety and stability. To facilitate glycoengineering of IgG-producing Chinese hamster ovary cells, we present a rapid protocol that involves the use of RNA interference for the knockdown of genes of interest coupled with capillary gel electrophoresis and laser-induced fluorescence detection (CGE-LIF) for fast, high-throughput glycan analysis. We apply this methodology to the Fut8 gene, responsible for the addition of core fucose, which is a typical target for increasing antibody-dependent cellular cytotoxicity.
Kawahara R, Chernykh A, Alagesan K, et al., 2021, Community evaluation of glycoproteomics informatics solutions reveals high-performance search strategies for serum glycopeptide analysis, NATURE METHODS, Vol: 18, Pages: 1304-+, ISSN: 1548-7091
Baksmeier C, Blundell P, Steckel J, et al., 2021, Modified recombinant human IgG1-Fc is superior to natural IVIG at inhibiting immune-mediated demyelination., Immunology, Vol: 464, Pages: 90-105, ISSN: 0019-2805
Intravenous immunoglobulin (IVIG) is an established treatment for numerous autoimmune conditions. Although Fc fragments derived from IVIG have shown efficacy in controlling immune thrombocytopenia (ITP) in children, the mechanisms of action are unclear and controversial. The aim of this study is to dissect IVIG effector mechanisms using further adapted Fc fragments on demyelination in an ex vivo model of the central nervous system (CNS)-immune interface. Using organotypic cerebellar slice cultures (OSC) from transgenic mice we induced extensive immune-mediated demyelination and oligodendrocyte loss with an antibody specific for myelin oligodendrocyte glycoprotein (MOG) and complement. Protective effects of adapted Fc fragments were assessed by live imaging of GFP expression, immunohistochemistry and confocal microscopy. Cysteine and glycan adapted Fc fragments protected OSC from demyelination in a dose-dependent manner where equimolar concentrations of either IVIG or control Fc were ineffective. The protective effects of the adapted Fc fragments are partly attributed to interference with complement-mediated oligodendroglia damage. Transcriptome analysis ruled out signatures associated with inflammatory or innate immune responses. Taken together our findings show that recombinant biomimetics can be made that are at least two hundred-fold more effective than IVIG in controlling demyelination by anti-MOG antibodies.
Ezeabikwa B, Mondal N, Antonopoulos A, et al., 2021, Major differences in glycosylation and Fucosyltransferase expression in low-grade versus high-grade bladder cancer cell lines., Glycobiology, ISSN: 0959-6658
Bladder cancer is the ninth most frequently diagnosed cancer worldwide, and there is a need to develop new biomarkers for staging and prognosis of this disease. Here we report that cell lines derived from low-grade and high-grade bladder cancers exhibit major differences in expression of glycans in surface glycoproteins. We analyzed protein glycosylation in three low-grade bladder cancer cell lines RT4 (grade-1-2), 5637 (grade-2), and SW780 (grade-1), and three high-grade bladder cancer cell lines J82COT (grade-3), T24 (grade-3), and TCCSUP (grade-4), with primary bladder epithelial cells, A/T/N, serving as a normal bladder cell control. Using a variety of approaches including flow cytometry, immunofluorescence, glycomics, and gene expression analysis, we observed that the low-grade bladder cancer cell lines RT4, 5637, and SW780 express high levels of the fucosylated Lewis x (Lex) antigen (CD15) (Galβ1-4(Fucα1-3) GlcNAcβ1-R), while normal bladder epithelial A/T/N cells lack Lex expression. T24 and TCCSUP cells also lack Lex, whereas J82COT cells express low levels of Lex. Glycomics analyses revealed other major differences in fucosylation and sialylation of N-glycans between these cell types. O-glycans are highly differentiated, as RT4 cells synthesize core 2-based O-glycans that are lacking in the T24 cells. These differences in glycan expression correlated with differences in RNA expression levels of their cognate glycosyltransferases, including α1-3/4-fucosyltransferase genes. These major differences in glycan structures and gene expression profiles between low- and high-grade bladder cancer cells suggest that glycans and glycosyltransferases are candidate biomarkers for grading bladder cancers.
Chakraborty A, Perez M, Mohammed NBB, et al., 2021, Hypoxia-mediated downregulation of GCNT2/I-antigen in metastatic melanoma accelerates disease progression and mortality., Publisher: AMER ASSOC CANCER RESEARCH, ISSN: 0008-5472
Bonnardel F, Haslam SM, Dell A, et al., 2021, Proteome-wide prediction of bacterial carbohydrate-binding proteins as a tool for understanding commensal and pathogen colonisation of the vaginal microbiome, npj Biofilms and Microbiomes, Vol: 7, Pages: 1-10, ISSN: 2055-5008
Bacteria use carbohydrate-binding proteins (CBPs), such as lectins and carbohydrate-binding modules (CBMs), to anchor to specific sugars on host surfaces. CBPs in the gut microbiome are well studied, but their roles in the vagina microbiome and involvement in sexually transmitted infections, cervical cancer and preterm birth are largely unknown. We established a classification system for lectins and designed Hidden Markov Model (HMM) profiles for data mining of bacterial genomes, resulting in identification of >100,000 predicted bacterial lectins available at unilectin.eu/bacteria. Genome screening of 90 isolates from 21 vaginal bacterial species shows that those associated with infection and inflammation produce a larger CBPs repertoire, thus enabling them to potentially bind a wider array of glycans in the vagina. Both the number of predicted bacterial CBPs and their specificities correlated with pathogenicity. This study provides new insights into potential mechanisms of colonisation by commensals and potential pathogens of the reproductive tract that underpin health and disease states.
Ng BG, Sosicka P, Fenaille F, et al., 2021, A mutation in SLC37A4 causes a dominantly inherited congenital disorder of glycosylation characterized by liver dysfunction, AMERICAN JOURNAL OF HUMAN GENETICS, Vol: 108, Pages: 1040-1052, ISSN: 0002-9297
Wang S-S, Solar VD, Yu X, et al., 2021, Efficient inhibition of O-glycan biosynthesis using the hexosamine analog Ac(5)GalNTGc, CELL CHEMICAL BIOLOGY, Vol: 28, Pages: 699-+, ISSN: 2451-9448
Wang Y, Khan A, Antonopoulos A, et al., 2021, Loss of alpha 2-6 sialylation promotes the transformation of synovial fibroblasts into a pro-inflammatory phenotype in arthritis, NATURE COMMUNICATIONS, Vol: 12, ISSN: 2041-1723
Donini R, Haslam S, Kontoravdi K, 2021, Glycoengineering Chinese hamster ovary cells: a short history, Biochemical Society Transactions, Vol: 49, Pages: 915-931, ISSN: 0300-5127
Biotherapeutic glycoproteins have revolutionised the field of pharmaceuticals, with new discoveries and continuous improvements underpinning the rapid growth of this industry. N-glycosylation is a critical quality attribute of biotherapeutic glycoproteins that influences the efficacy, half-life and immunogenicity of these drugs. This review will focus on the advances and future directions of remodelling N-glycosylation in Chinese hamster ovary (CHO) cells, which are the workhorse of recombinant biotherapeutic production, with particular emphasis on antibody products, using strategies such as cell line and protein backbone engineering.
Cao H, Antonopoulos A, Henderson S, et al., 2021, Red blood cell mannoses as phagocytic ligands mediating both sickle cell anaemia and malaria resistance, Nature Communications, Vol: 12, Pages: 1-13, ISSN: 2041-1723
In both sickle cell disease and malaria, red blood cells (RBCs) are phagocytosed in the spleen, but receptor-ligand pairs mediating uptake have not been identified. Here, we report that patches of high mannose N-glycans (Man5-9GlcNAc2), expressed on diseased or oxidized RBC surfaces, bind the mannose receptor (CD206) on phagocytes to mediate clearance. We find that extravascular hemolysis in sickle cell disease correlates with high mannose glycan levels on RBCs. Furthermore, Plasmodium falciparum-infected RBCs expose surface mannose N-glycans, which occur at significantly higher levels on infected RBCs from sickle cell trait subjects compared to those lacking hemoglobin S. The glycans are associated with high molecular weight complexes and protease-resistant, lower molecular weight fragments containing spectrin. Recognition of surface N-linked high mannose glycans as a response to cellular stress is a molecular mechanism common to both the pathogenesis of sickle cell disease and resistance to severe malaria in sickle cell trait.
Antonopoulos A, Broome S, Sharov V, et al., 2021, Site-specific characterisation of SARS-CoV-2 spike glycoprotein receptor binding domain, Glycobiology, Vol: 31, Pages: 181-187, ISSN: 0959-6658
The novel coronavirus SARS-CoV-2, the infective agent causing COVID-19, is having a global impact both in terms of human disease as well as socially and economically. Its heavily glycosylated spike glycoprotein is fundamental for the infection process, via its receptor binding domains interaction with the glycoprotein angiotensin converting enzyme 2 on human cell surfaces. We therefore utilized an integrated glycomic and glycoproteomic analytical strategy to characterise both N- and O- glycan site specific glycosylation within the receptor binding domain. We demonstrate the presence of complex type N-glycans with unusual fucosylated LacdiNAc at both sites N331 and N343 and a single site of O-glycosylation on T323.
Cao H, Bagchi A, Tampakis D, et al., 2021, Human erythrocyte surface fucose expression increases with age and hyperglycemia, Wellcome Open Research, Vol: 6, Pages: 1-14, ISSN: 2398-502X
Background: Reactive oxygen species and other free radicals, together with glucose and its metabolites are believed to play important roles in the aging process. The carbohydrate components of glycosylated proteins are important in mediating cell-cell interactions and a role has been suggested for them in the aging process. Erythrocytes are critical cells in the human body, heavily glycosylated and relatively easily available and so are good candidates to yield insights into how patterns of glycosylation change with age and disease. It has been claimed, based on a periodic acid Schiff assay, that human aging is associated with a decline of erythrocyte surface sialic acids. Plant lectins allow for more specific assays for glycans, including determining the linkage of sialic acids and analysis of single cells by flow cytometry.Methods: Plant lectins, including Maackia amurensis lectin II (MAL), binding to α-2,3 linked sialic acids and Sambucus nigra (SNA), α-2,6 sialic acids, were used in flow cytometry and western blot of erythrocyte surface membrane. N-glycomics mass spectrometry determines glycan structures. Donors varying in age and hyperglycemia, as indicated by HbA1c were analysed.Results: Erythrocyte surface sialic acids have no significant associations with donor age. A combination of storage and cellular aging produces a specific loss of α-2,6 sialic acids. By contrast, erythrocyte surface terminal fucoses increase significantly with donor age. In order to determine which aspects of aging are important in determining this change, we investigated whether this novel human aging biomarker is associated with higher plasma glucose values, assessed by glycated hemoglobin (HbA1c) and reactive oxygen species (ROS) generation. Fucose levels were associated with HbA1c levels, but not ROS generation.Conclusion: Our study identifies novel glycan-based biomarkers for human aging and disease. The simplicity of lectin-based assays provide an attractive cel
Mendoza M, Lu D, Ballesteros A, et al., 2020, Glycan characterization of pregnancy-specific glycoprotein 1 and its identification as a novel Galectin-1 ligand, Glycobiology, Vol: 30, Pages: 895-909, ISSN: 0959-6658
Pregnancy-specific beta 1 glycoprotein (PSG1) is secreted from trophoblast cells of the human placenta in increasing concentrations as pregnancy progresses, becoming one of the most abundant proteins in maternal serum in the third trimester. PSG1 has seven potential N-linked glycosylation sites across its four domains. We carried out glycomic and glycoproteomic studies to characterize the glycan composition of PSG1 purified from serum of pregnant women and identified the presence of complex N-glycans containing poly LacNAc epitopes with α2,3 sialyation at four sites. Using different techniques, we explored whether PSG1 can bind to galectin-1 (Gal-1) as these two proteins were previously shown to participate in processes required for a successful pregnancy. We confirmed that PSG1 binds to Gal-1 in a carbohydrate-dependent manner with an affinity of the interaction of 0.13 μM. In addition, we determined that out of the three N-glycosylation-carrying domains, only the N and A2 domains of recombinant PSG1 interact with Gal-1. Lastly, we observed that the interaction between PSG1 and Gal-1 protects this lectin from oxidative inactivation and that PSG1 competes the ability of Gal-1 to bind to some but not all of its glycoprotein ligands.
Loxley GM, Hooks DO, Antonopoulos A, et al., 2020, Vulpeculin: a novel and abundant lipocalin in the urine of the common brushtail possum,Trichosurus vulpecula, OPEN BIOLOGY, Vol: 10
Murphy N, Rooney B, Bhattacharyya T, et al., 2020, Glycosylation of trypanosoma cruzi TcI antigen reveals recognition by chagasic sera, Scientific Reports, Vol: 10, ISSN: 2045-2322
Chagas disease is considered the most important parasitic disease in Latin America. The protozoan agent, Trypanosoma cruzi, comprises six genetic lineages, TcI-TcVI. Genotyping to link lineage(s) to severity of cardiomyopathy and gastrointestinal pathology is impeded by the sequestration and replication of T. cruzi in host tissues. We describe serology specific for TcI, the predominant lineage north of the Amazon, based on expression of recombinant trypomastigote small surface antigen (gTSSA-I) in the eukaryote Leishmania tarentolae, to allow realistic glycosylation and structure of the antigen. Sera from TcI-endemic regions recognised gTSSA-I (74/146; 50.7%), with no cross reaction with common components of gTSSA-II/V/VI recombinant antigen. Antigenicity was abolished by chemical (periodate) oxidation of gTSSA-I glycosylation but retained after heat-denaturation of conformation. Conversely, non-specific recognition of gTSSA-I by non-endemic malaria sera was abolished by heat-denaturation. TcI-specific serology facilitates investigation between lineage and diverse clinical presentations. Glycosylation cannot be ignored in the search for immunogenic antigens.
Debets MF, Tastan OY, Wisnovsky SP, et al., 2020, Metabolic precision labeling enables selective probing of O-linked N-acetylgalactosamine glycosylation, Proceedings of the National Academy of Sciences of USA, Vol: 117, Pages: 25293-25301, ISSN: 0027-8424
Protein glycosylation events that happen early in the secretory pathway are often dysregulated during tumorigenesis. These events can be probed, in principle, by monosaccharides with bioorthogonal tags that would ideally be specific for distinct glycan subtypes. However, metabolic interconversion into other monosaccharides drastically reduces such specificity in the living cell. Here, we use a structure-based design process to develop the monosaccharide probe GalNAzMe that is specific for cancer-relevant Ser/Thr-N-acetylgalactosamine (O-GalNAc) glycosylation. By virtue of a branched N-acylamide side chain, GalNAzMe is not interconverted by epimerization to the corresponding N-acetylglucosamine analog by the epimerase GALE like conventional GalNAc-based probes. GalNAzMe enters O-GalNAc glycosylation but does not enter other major cell surface glycan types including Asn(N)-linked glycans. We transfect cells with the engineered pyrophosphorylase mut-AGX1 to biosynthesize the nucleotide-sugar donor UDP-GalNAzMe from a sugar-1-phosphate precursor. Tagged with a bioorthogonal azide group, GalNAzMe serves as an O-glycan specific reporter in superresolution microscopy, chemical glycoproteomics, a genome-wide CRISPR knock-out (KO) screen, and imaging of intestinal organoids. Additional ectopic expression of an engineered glycosyltransferase, BH-GalNAc-T2, boosts labeling in a programmable fashion by increasing incorporation of GalNAzMe into the cell surface glycoproteome. Alleviating the need for GALE-KO cells in metabolic labeling experiments, GalNAzMe is a precision tool that allows a detailed view into the biology of a major type of cancer-relevant protein glycosylation.
Li H, Marceau M, Yang T, et al., 2020, East-Asian Helicobacter pylori strains synthesize heptan-deficient lipopolysaccharide, Publisher: WILEY, Pages: 75-75, ISSN: 1083-4389
Sela I, Goss V, Becker-Cohen M, et al., 2020, The glycomic sialylation profile of GNE Myopathy muscle cells does not point to consistent hyposialylation of individual glycoconjugates, NEUROMUSCULAR DISORDERS, Vol: 30, Pages: 621-630, ISSN: 0960-8966
Hautala LC, Pang P-C, Antonopoulos A, et al., 2020, Altered glycosylation of glycodelin in endometrial carcinoma, Laboratory Investigation, Vol: 100, Pages: 1014-1025, ISSN: 0023-6837
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 (Galβ1–4GlcNAc) 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
Blois SM, Verlohren S, Wu G, et al., 2020, Role of galectin-glycan circuits in reproduction: from healthy pregnancy to preterm birth (PTB), Seminars in Immunopathology, Vol: 42, Pages: 469-486, ISSN: 1863-2297
Growing evidence suggests that galectins, an evolutionarily conserved family of glycan-binding proteins, fulfill key roles in pregnancy including blastocyst implantation, maternal-fetal immune tolerance, placental development, and maternal vascular expansion, thereby establishing a healthy environment for the growing fetus. In this review, we comprehensively present the function of galectins in shaping cellular circuits that characterize a healthy pregnancy. We describe the current understanding of galectins in term and preterm labor and discuss how the galectin-glycan circuits contribute to key immunological pathways sustaining maternal tolerance and preventing microbial infections. A deeper understanding of the glycoimmune pathways regulating early events in preterm birth could offer the broader translational potential for the treatment of this devastating syndrome.
Osimanjiang W, Santos Roballo KC, Houck BD, et al., 2020, Analysis of N- and O-Linked Glycosylation: Differential Glycosylation after Rat Spinal Cord Injury, JOURNAL OF NEUROTRAUMA, Vol: 37, Pages: 1954-1962, ISSN: 0897-7151
Blundell PA, Lu D, Dell A, et al., 2020, Choice of Host Cell Line Is Essential for the Functional Glycosylation of the Fc Region of Human IgG1 Inhibitors of Influenza B Viruses, JOURNAL OF IMMUNOLOGY, Vol: 204, Pages: 1022-1034, ISSN: 0022-1767
Silver ZA, Antonopoulos A, Haslam SM, et al., 2020, Discovery of O-linked carbohydrate on HIV-1 envelope and its role in shielding against one category of broadly neutralizing antibodies, Cell Reports, Vol: 30, Pages: 1862-1869.e4, ISSN: 2211-1247
Approximately 50% of the mass of the Envelope (Env) glycoprotein surface subunit (gp120) of human immunodeficiency virus type 1 (HIV-1) is composed of N-linked carbohydrate. Until now, the dogma has been that HIV-1 lacks O-linked carbohydrate on Env. Here we show that a subset of patient-derived HIV-1 isolates contain O-linked carbohydrate on the variable 1 (V1) domain of Env gp120. We demonstrate the presence of this O-glycosylation both on virions and on gp120 expressed as a secreted protein. Further, we establish that these O-linked glycans can confer a more than 1,000-fold decrease in neutralization sensitivity (IC50) to V3-glycan broadly neutralizing antibodies. These findings uncover a structural modification to the HIV-1 Env and suggest a functional role in promoting viral escape from one category of broadly neutralizing antibodies.
Ibeto L, Antonopoulos A, Grassi P, et al., 2020, Insights into the hyperglycosylation of human chorionic gonadotropin revealed by glycomics analysis, PLoS One, Vol: 15, ISSN: 1932-6203
Human chorionic gonadotropin (hCG) is a glycoprotein hormone that is essential for the maintenance of pregnancy. Glycosylation of hCG is known to be essential for its biological activity. "Hyperglycosylated" variants secreted during early pregnancy have been proposed to be involved in initial implantation of the embryo and as a potential diagnostic marker for gestational diseases. However, what constitutes "hyperglycosylation" is not yet fully understood. In this study, we perform comparative N-glycomic analysis of hCG expressed in the same individuals during early and late pregnancy to help provide new insights into hCG function, reveal new targets for diagnostics and clarify the identity of hyperglycosylated hCG. hCG was isolated in urine collected from women at 7 weeks and 20 weeks' gestation. hCG was also isolated in urine from women diagnosed with gestational trophoblastic disease (GTD). We used glycomics methodologies including matrix assisted laser desorption/ionisation-time of flight (MALDI-TOF) mass spectrometry (MS) and MS/MS methods to characterise the N-glycans associated with hCG purified from the individual samples. The structures identified on the early pregnancy (EP-hCG) and late pregnancy (LP-hCG) samples corresponded to mono-, bi-, tri-, and tetra-antennary N-glycans. A novel finding was the presence of substantial amounts of bisected type N-glycans in pregnancy hCG samples, which were present at much lower levels in GTD samples. A second novel observation was the presence of abundant LewisX antigens on the bisected N-glycans. GTD-hCG had fewer glycoforms which constituted a subset of those found in normal pregnancy. When compared to EP-hCG, GTD-hCG samples had decreased signals for tri- and tetra-antennary N-glycans. In terms of terminal epitopes, GTD-hCG had increased signals for sialylated structures, while LewisX antigens were of very minor abundance. hCG carries the same N-glycans throughout pregnancy but in different propo
Schedin-Weiss S, Gaunitz S, Sui P, et al., 2020, Glycan biomarkers for Alzheimer disease correlate with T-tau and P-tau in cerebrospinal fluid in subjective cognitive impairment, FEBS JOURNAL, Vol: 287, Pages: 3221-3234, ISSN: 1742-464X
Li H, Marceau M, Yang T, et al., 2019, East-Asian Helicobacter pylori Strains Synthesize Heptan-deficient Lipopolysaccharide, PLOS GENETICS, Vol: 15, ISSN: 1553-7404
Monzon Manzano E, Justo Sanz R, Fernandez Bello I, et al., 2019, CHANGES IN THE GLYCOSYLATION PATTERN OF THE PLATELET MEMBRANE MAY BE THE CHANGES IN THE GLYCOSYLATION PATTERN OF THE PLATELET MEMBRANE MAY BE INVOLVED IN THE PATOGENESIS OF THE PRIMARY IMMUNE THROMBOCYTOPENIA, 61st National Congress of the Spanish-Society-of-Hematology-and-Hemotherapy, Publisher: FERRATA STORTI FOUNDATION, Pages: 349-349, ISSN: 0390-6078
North SJ, Botchway K, Doonan J, et al., 2019, Site-specific glycoproteomic characterization of ES-62: The major secreted product of the parasitic worm Acanthocheilonema viteae, Glycobiology, Vol: 29, Pages: 562-571, ISSN: 0959-6658
ES-62 is the major secreted product of the parasitic filarial nematode Acanthocheilonema viteae and has potent anti-inflammatory activities as a consequence of post-translational decoration by phosphorylcholine. Previously we showed that ES-62's phosphorylcholine was attached to N-linked glycans and using fast atom bombardment mass spectrometry, we characterised the structure of the glycans. However, it was unknown at this time which of ES-62's four potential N-glycosylation sites carries the phosphorylcholine-modified glycans. In the present study, we now employ more advanced analytical tools - nano-flow liquid chromatography with high definition electrospray mass spectrometry - to show that phosphorylcholine-modified glycans are found at all four potential N-glycosylation sites. Also, our earlier studies showed up to two phosphorylcholine groups were detected per glycan and we are now able to characterise N-glycans with up to five phosphorylcholine groups. The number per glycan varies in three of the four glycosylation sites and in addition, for the first time, we have detected phosphorylcholine on the N-glycan chitobiose core in addition to terminal GlcNAc. Nevertheless, the majority of phosphorylcholine is detected on terminal GlcNAc, enabling it to interact with the cells and molecules of the immune system. Such expression may explain the potent immunomodulatory effects of a molecule that is considered to have significant therapeutic potential in the treatment of certain human allergic and autoimmune conditions.
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