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Cornelis H Hokke - One of the best experts on this subject based on the ideXlab platform.
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novel o linked methylated Glycan antigens decorate secreted immunodominant glycoproteins from the intestinal nematode heligmosomoides polygyrus
International Journal for Parasitology, 2016Co-Authors: James P Hewitson, Linh D Nguyen, Angela Van Diepen, Cornelis H Smit, Carolien A M Koeleman, Henry J Mcsorley, Janice Murray, Rick M Maizels, Cornelis H HokkeAbstract:Glycan molecules from helminth parasites have been associated with diverse biological functions ranging from interactions with neighbouring host cell populations to down-modulation of specific host immunity. Glycoproteins secreted by the intestinal nematode Heligmosomoides polygyrus are of particular interest as the excretory–secretory products (termed HES) of this parasite contain both heat-labile and heat-stable components with immunomodulatory effects. We used MALDI-TOF-MS and LC–MS/MS to analyse the repertoire of N- and O-linked Glycans released from Heligmosomoides polygyrus excretory–secretory products by PNGase A and F, β-elimination and hydrazinolysis revealing a broad range of structures including novel methylhexose- and methylfucose-containing Glycans. Monoclonal antibodies to two immunodominant Glycans of H. polygyrus, previously designated Glycans A and B, were found to react by Glycan array analysis to a methyl-hexose-rich fraction and to a sulphated LacDiNAc (LDN; GalNAcβ1–4GlcNAc) structure, respectively. We also analysed the Glycan repertoire of a major glycoprotein in Heligmosomoides polygyrus excretory–secretory products, VAL-2, which contains many Glycan structures present in Heligmosomoides polygyrus excretory–secretory products including Glycan A. However, it was found that this set of Glycans is not responsible for the heat-stable immunomodulatory properties of Heligmosomoides polygyrus excretory–secretory products, as revealed by the inability of VAL-2 to inhibit allergic lung inflammation. Taken together, these studies reveal that H. polygyrus secretes a diverse range of antigenic glycoconjugates, and provides a framework to explore the biological and immunomodulatory roles they may play within the mammalian host.
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surface expression patterns of defined Glycan antigens change during schistosoma mansoni cercarial transformation and development of schistosomula
Glycobiology, 2015Co-Authors: Cornelis H Smit, Angela Van Diepen, Arne Homann, Vincent P Van Hensbergen, Gabriele Schramm, Helmut L Haas, Cornelis H HokkeAbstract:During the complex lifecycle of Schistosoma mansoni, a large variety of Glycans is expressed. To many of these Glycans, antibodies are induced by the infected host and some might be targets for vaccines or diagnostic tests. Spatial changes in Glycan expression during schistosome development are largely unexplored. To study the surface-exposed Glycans during the important initial stages of infection, we analyzed the binding of a panel of anti-Glycan monoclonal antibodies (mAbs) to cercariae and schistosomula up to 72 h after transformation by immunofluorescence microscopy. The mAb specificity toward their natural targets was studied using a microarray containing a wide range of schistosomal N-Glycans, O-Glycans and glycosphingolipid Glycans. With the exception of GalNAcβ1-4(Fucα1-3)GlcNAc (LDN-F), mono- and multifucosylated GalNAcβ1-4GlcNAc (LDN)-motifs were exposed at the surface of all developmental stages studied. Multifucosylated LDN-motifs were present on cercarial glycocalyx-derived O-Glycans as well as cercarial glycolipids. In contrast, the Galβ1-4(Fucα1-3)GlcNAc (Lewis X) and LDN-F-motifs, also expressed on cercarial glycolipids, and in addition on a range of cercarial N- and O-Glycans, became surface expressed only after transformation of cercariae to schistosomula. In line with the documented shedding of the O-Glycan-rich cercarial glycocalyx after transformation these observations suggest that surface accessible multifucosylated LDN-motifs are mostly expressed by O-Glycans in cercariae, but principally by glycosphingolipids in schistosomula. We hypothesize that these temporal changes in surface exposure of Glycan antigens are relevant to the interaction with the host during the initial stages of infection with schistosomes and discuss the potential of these Glycan antigens as intervention targets.
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serum antibody screening by surface plasmon resonance using a natural Glycan microarray
Glycoconjugate Journal, 2008Co-Authors: Arjen R De Boer, Andre M Deelder, Cornelis H Hokke, Manfred WuhrerAbstract:A surface plasmon resonance (SPR) based natural Glycan microarray was developed for screening of interactions between Glycans and carbohydrate-binding proteins (CBPs). The microarray contained 144 Glycan samples and allowed the real-time and simultaneous screening for recognition by CBPs without the need of fluorescent labeling. Glycans were released from their natural source and coupled by reductive amination with the fluorescent labels 2-aminobenzamide (2AB) or anthranilic acid (AA) followed by high-performance liquid chromatography (HPLC) fractionation making use of the fluorescent tag. The released and labeled Glycans, in addition to fluorescently labeled synthetic Glycans and (neo)glycoproteins, were printed on an epoxide-activated chip at fmol amounts. This resulted in covalent immobilization, with the epoxide groups forming covalent bonds to the secondary amine groups present on the fluorescent glycoconjugates. The generated SPR Glycan array presented a subset of the Glycan repertoire of the human parasite Schistosoma mansoni. In order to demonstrate the usefulness of the array in the simultaneous detection of Glycan-specific serum antibodies, the anti-Glycan antibody profiles from sera of S. mansoni-infected individuals as well as from non-endemic uninfected controls were recorded. The SPR screening was sensitive for differences between infection sera and control sera, and revealed antibody titers and antibody classes (IgG or IgM). All SPR analyses were performed with a single SPR array chip, which required regeneration and blocking of the chip before the application of a serum sample. Our results indicate that SPR-based arrays constructed from Glycans of natural or synthetic origin, pure or as mixture, can be used for determining serum antibody profiles as possible markers for the infection status of an individual.
Richard D. Cummings - One of the best experts on this subject based on the ideXlab platform.
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Fluorescent Glycosylamides Produced by Microscale Derivatization of Free Glycans for Natural Glycan Microarrays
2015Co-Authors: Xuezheng Song, Yi Lasanajak, David F Smith, Baoyun Xia, Richard D. CummingsAbstract:A novel strategy for creating naturally derived Glycan microarrays has been developed. Glycosylamines are prepared from free reducing Glycans and stabilized by reaction with acryloyl chloride to generate a glycosylamide in which the reducing monosaccharide has a closed-ring structure. Ozonolysis of the protected Glycan yields an active aldehyde, to which a bifunctional fluorescent linker is coupled by reductive amination. The fluorescent derivatives are easily coupled through a residual primary alkylamine to generate Glycan microarrays. This strategy preserves structural features of Glycans required for antibody recognition and allows development of natural arrays of fluorescent Glycans in which the cyclic pyranose structure of the reducing-end sugar residue is retained
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structural characterization by multistage mass spectrometry msn of human milk Glycans recognized by human rotaviruses
Molecular & Cellular Proteomics, 2014Co-Authors: David J Ashline, Yi Lasanajak, Liya Hu, Sasirekha Ramani, B Venkataram V Prasad, Xuezheng Song, Richard D. Cummings, Ying Yu, Mary K. Estes, David F SmithAbstract:We have shown that recombinant forms of VP8* domains of the human rotavirus outer capsid spike protein VP4 from human neonatal strains (N155(G10P[11]) and RV3(G3P[6]) and a bovine strain (B223) recognize unique Glycans within the repertoire of human milk Glycans. The accompanying study by Yu et al.2, describes a human milk Glycan shotgun Glycan microarray that led to the identification of 32 specific Glycans in the human milk tagged Glycan library that were recognized by these human rotaviruses. These microarray analyses also provided a variety of metadata about the recognized Glycan structures compiled from anti-Glycan antibody and lectin binding before and after specific glycosidase digestions, along with compositional information from mass analysis by matrix-assisted laser desorption ionization-mass spectrometry. To deduce Glycan sequence and utilize information predicted by analyses of metadata from each Glycan, 28 of the Glycan targets were retrieved from the tagged Glycan library for detailed sequencing using sequential disassembly of Glycans by ion-trap mass spectrometry. Our aim is to obtain a deeper structural understanding of these key Glycans using an orthogonal approach for structural confirmation in a single ion trap mass spectrometer. This sequential ion disassembly strategy details the complexities of linkage and branching in multiple compositions, several of which contained isomeric mixtures including several novel structures. The application of this approach exploits both library matching with standard materials and de novo approaches. This combination together with the metadata generated from lectin and antibody-binding data before and after glycosidase digestions provide a heretofore-unavailable level of analytical detail to Glycan structure analysis. The results of these studies showed that, among the 28 Glycan targets analyzed, 27 unique structures were identified, and 23 of the human milk Glycans recognized by human rotaviruses represent novel structures not previously described as Glycans in human milk. The functional glycomics analysis of human milk Glycans provides significant insight into the repertoire of Glycans comprising the human milk metaglycome.
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a sialylated Glycan microarray reveals novel interactions of modified sialic acids with proteins and viruses
Journal of Biological Chemistry, 2011Co-Authors: Xuezheng Song, Yi Lasanajak, Mary M. Tappert, Gillian M. Air, Xi Chen, Hongzhi Cao, Vinod K Tiwari, Harshal A Chokhawala, Haojie Zheng, Richard D. CummingsAbstract:Many Glycan-binding proteins in animals and pathogens recognize sialic acid or its modified forms, but their molecular recognition is poorly understood. Here we describe studies on sialic acid recognition using a novel sialylated Glycan microarray containing modified sialic acids presented on different Glycan backbones. Glycans terminating in β-linked galactose at the non-reducing end and with an alkylamine-containing fluorophore at the reducing end were sialylated by a one-pot three-enzyme system to generate α2–3- and α2–6-linked sialyl Glycans with 16 modified sialic acids. The resulting 77 sialyl Glycans were purified and quantified, characterized by mass spectrometry, covalently printed on activated slides, and interrogated with a number of key sialic acid-binding proteins and viruses. Sialic acid recognition by the sialic acid-binding lectins Sambucus nigra agglutinin and Maackia amurensis lectin-I, which are routinely used for detecting α2–6- and α2–3-linked sialic acids, are affected by sialic acid modifications, and both lectins bind Glycans terminating with 2-keto-3-deoxy-d-glycero-d-galactonononic acid (Kdn) and Kdn derivatives stronger than the derivatives of more common N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc). Three human parainfluenza viruses bind to Glycans terminating with Neu5Ac or Neu5Gc and some of their derivatives but not to Kdn and its derivatives. Influenza A virus also does not bind Glycans terminating in Kdn or Kdn derivatives. An especially novel aspect of human influenza A virus binding is its ability to equivalently recognize Glycans terminated with either α2–6-linked Neu5Ac9Lt or α2–6-linked Neu5Ac. Our results demonstrate the utility of this sialylated Glycan microarray to investigate the biological importance of modified sialic acids in protein-Glycan interactions.
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Glycan gimmickry by parasitic helminths a strategy for modulating the host immune response
Glycobiology, 2010Co-Authors: I Van Die, Richard D. CummingsAbstract:Parasitic helminths (worms) co-evolved with vertebrate immune systems to enable long-term survival of worms in infected hosts. Among their survival strategies, worms use their Glycans within glycoproteins and glycolipids, which are abundant on helminth surfaces and in their excretory/ secretory products, to regulate and suppress host immune responses. Many helminths express unusual and antigenic (nonhost-like) Glycans, including those containing polyfucose, tyvelose, terminal GalNAc, phosphorylcholine, methyl groups, and sugars in unusual linkages. In addition, some Glycan antigens are expressed that share structural features with those in their intermediate and vertebrate hosts (hostlike Glycans), including Le X (Galβ1-4[Fucα1-3]GlcNAc-), LDNF (GalNAcβ1-4[Fucα1-3]GlcNAc-), LDN (GalNAcβ14GlcNAc-), and Tn (GalNAcα1-O-Thr/Ser) antigens. The expression of host-like Glycan determinants is remarkable and suggests that helminths may gain advantages by synthesizing such Glycans. The expression of host-like Glycans by parasites previously led to the concept of “molecular mimicry,” in which molecules are either derived from the pathogen or acquired from the host to evade recognition by the host immune system. However, recent discoveries into the potential of host Glycan-binding proteins (GBPs), such as C-type lectin receptors and galectins, to functionally interact with various host-like helminth Glycans provide new insights. Host GBPs through their interactions with wormderived Glycans participate in shaping innate and adaptive immune responses upon infection. We thus propose an alternative concept termed “Glycan gimmickry,” which is defined as an active strategy of parasites to use their Glycans to target GBPs within the host to promote their survival.
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Glycan gimmickry by parasitic helminths a strategy for modulating the host immune response
Glycobiology, 2010Co-Authors: I Van Die, Richard D. CummingsAbstract:Parasitic helminths (worms) co-evolved with vertebrate immune systems to enable long-term survival of worms in infected hosts. Among their survival strategies, worms use their Glycans within glycoproteins and glycolipids, which are abundant on helminth surfaces and in their excretory/ secretory products, to regulate and suppress host immune responses. Many helminths express unusual and antigenic (nonhost-like) Glycans, including those containing polyfucose, tyvelose, terminal GalNAc, phosphorylcholine, methyl groups, and sugars in unusual linkages. In addition, some Glycan antigens are expressed that share structural features with those in their intermediate and vertebrate hosts (host-like Glycans), including Le(X) (Galbeta1-4[Fucalpha1-3]GlcNAc-), LDNF (GalNAcbeta1-4[Fucalpha1-3]GlcNAc-), LDN (GalNAcbeta1-4GlcNAc-), and Tn (GalNAcalpha1-O-Thr/Ser) antigens. The expression of host-like Glycan determinants is remarkable and suggests that helminths may gain advantages by synthesizing such Glycans. The expression of host-like Glycans by parasites previously led to the concept of "molecular mimicry," in which molecules are either derived from the pathogen or acquired from the host to evade recognition by the host immune system. However, recent discoveries into the potential of host Glycan-binding proteins (GBPs), such as C-type lectin receptors and galectins, to functionally interact with various host-like helminth Glycans provide new insights. Host GBPs through their interactions with worm-derived Glycans participate in shaping innate and adaptive immune responses upon infection. We thus propose an alternative concept termed "Glycan gimmickry," which is defined as an active strategy of parasites to use their Glycans to target GBPs within the host to promote their survival.
Yusuf Vohra - One of the best experts on this subject based on the ideXlab platform.
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vaccine elicitation of high mannose dependent neutralizing antibodies against the v3 Glycan broadly neutralizing epitope in nonhuman primates
Cell Reports, 2017Co-Authors: Kevin O Saunders, Nathan I Nicely, Kevin Wiehe, Mattia Bonsignori, Ryan R Meyerhoff, Robert Parks, William E Walkowicz, Baptiste Aussedat, Fangping Cai, Yusuf VohraAbstract:Summary Induction of broadly neutralizing antibodies (bnAbs) that target HIV-1 envelope (Env) is a goal of HIV-1 vaccine development. A bnAb target is the Env third variable loop (V3)-Glycan site. To determine whether immunization could induce antibodies to the V3-Glycan bnAb binding site, we repetitively immunized macaques over a 4-year period with an Env expressing V3-high mannose Glycans. Env immunizations elicited plasma antibodies that neutralized HIV-1 expressing only high-mannose Glycans—a characteristic shared by early bnAb B cell lineage members. A rhesus recombinant monoclonal antibody from a vaccinated macaque bound to the V3-Glycan site at the same amino acids as broadly neutralizing antibodies. A structure of the antibody bound to Glycan revealed that the three variable heavy-chain complementarity-determining regions formed a cavity into which Glycan could insert and neutralized multiple HIV-1 isolates with high-mannose Glycans. Thus, HIV-1 Env vaccination induced mannose-dependent antibodies with characteristics of V3-Glycan bnAb precursors.
Alsteens David - One of the best experts on this subject based on the ideXlab platform.
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Initial Step of Virus Entry: Virion Binding to Cell-Surface Glycans
'Annual Reviews', 2021Co-Authors: Koehler Melanie, Delguste Martin, Sieben Christian, Gillet Laurent, Alsteens DavidAbstract:Virus infection is an intricate process that requires the concerted action of both viral and host cell components. Entry of viruses into cells is initiated by interactions between viral proteins and cell-surface receptors. Various cell-surface Glycans function as initial, usually low-affinity attachment factors, providing a first anchor of the virus to the cell surface, and further facilitate high-affinity binding to virus-specific cell-surface receptors, while other Glycans function as specific entry receptors themselves. It is now possible to rapidly identify specific Glycan receptors using different techniques, define atomic-level structures of virus-Glycan complexes, and study these interactions at the single-virion level. This review provides a detailed overview of the role of Glycans in viral infection and highlights experimental approaches to study virus-Glycan binding along with specific examples. In particular, we highlight the development of the atomic force microscope to investigate interactions with Glycans at the single-virion level directly on living mammalian cells, which offers new perspectives to better understand virus-Glycan interactions in physiologically relevant conditions
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Initial step of virus entry: virion binding to cell surface Glycans
'Annual Reviews', 2020Co-Authors: Köhler Melanie, Delguste Martin, Sieben Christian, Gillet Laurent, Alsteens DavidAbstract:Virus infection is an intricate process that requires the concerted action of both viral and host cell components. Entry of viruses into cells is initiated by interactions between viral proteins and cell surface receptors. Various cell surface Glycans function as initial, usually low-affinity attachment factors, providing a first anchor of the viruses to the cell surface and further facilitate high-affinity binding to virus-specific cell surface receptors, while other Glycans function as specific entry receptors themselves. It is now possible to rapidly identify specific Glycan receptors using different techniques, to define atomic-level structures of virus-Glycan complexes and to study these interactions at the single-virion level. This review gives a detailed overview of the role of Glycans in viral infection and highlights experimental approaches to study virus-Glycan binding along with specific examples. In particular, we highlight the development of the atomic force microscope to investigate interactions with Glycans at the single-virion level directly on living mammalian cells offering new perspectives to better understand virus-Glycan interactions in physiologically relevant conditions
Kevin O Saunders - One of the best experts on this subject based on the ideXlab platform.
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vaccine elicitation of high mannose dependent neutralizing antibodies against the v3 Glycan broadly neutralizing epitope in nonhuman primates
Cell Reports, 2017Co-Authors: Kevin O Saunders, Nathan I Nicely, Kevin Wiehe, Mattia Bonsignori, Ryan R Meyerhoff, Robert Parks, William E Walkowicz, Baptiste Aussedat, Fangping Cai, Yusuf VohraAbstract:Summary Induction of broadly neutralizing antibodies (bnAbs) that target HIV-1 envelope (Env) is a goal of HIV-1 vaccine development. A bnAb target is the Env third variable loop (V3)-Glycan site. To determine whether immunization could induce antibodies to the V3-Glycan bnAb binding site, we repetitively immunized macaques over a 4-year period with an Env expressing V3-high mannose Glycans. Env immunizations elicited plasma antibodies that neutralized HIV-1 expressing only high-mannose Glycans—a characteristic shared by early bnAb B cell lineage members. A rhesus recombinant monoclonal antibody from a vaccinated macaque bound to the V3-Glycan site at the same amino acids as broadly neutralizing antibodies. A structure of the antibody bound to Glycan revealed that the three variable heavy-chain complementarity-determining regions formed a cavity into which Glycan could insert and neutralized multiple HIV-1 isolates with high-mannose Glycans. Thus, HIV-1 Env vaccination induced mannose-dependent antibodies with characteristics of V3-Glycan bnAb precursors.
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Vaccine Elicitation of High Mannose-Dependent Neutralizing Antibodies against the V3-Glycan Broadly Neutralizing Epitope in Nonhuman Primates
Elsevier, 2017Co-Authors: Kevin O Saunders, Nathan I Nicely, Kevin Wiehe, Mattia Bonsignori, Ryan R Meyerhoff, Robert Parks, William E Walkowicz, Baptiste Aussedat, Fangping CaiAbstract:Summary: Induction of broadly neutralizing antibodies (bnAbs) that target HIV-1 envelope (Env) is a goal of HIV-1 vaccine development. A bnAb target is the Env third variable loop (V3)-Glycan site. To determine whether immunization could induce antibodies to the V3-Glycan bnAb binding site, we repetitively immunized macaques over a 4-year period with an Env expressing V3-high mannose Glycans. Env immunizations elicited plasma antibodies that neutralized HIV-1 expressing only high-mannose Glycans—a characteristic shared by early bnAb B cell lineage members. A rhesus recombinant monoclonal antibody from a vaccinated macaque bound to the V3-Glycan site at the same amino acids as broadly neutralizing antibodies. A structure of the antibody bound to Glycan revealed that the three variable heavy-chain complementarity-determining regions formed a cavity into which Glycan could insert and neutralized multiple HIV-1 isolates with high-mannose Glycans. Thus, HIV-1 Env vaccination induced mannose-dependent antibodies with characteristics of V3-Glycan bnAb precursors. : Most bnAb epitopes on HIV-1 Envelope include host Glycans, but previous Env vaccines have not induced Glycan-dependent antibodies. Saunders et al. describe here the ontogeny, crystal structure with Glycan, and virion Man9GlcNAc2-dependent neutralization for Glycan-reactive antibodies induced by envelope vaccination. Keywords: HIV, V3 Glycan, vaccination, Glycan, long-term immunizatio