| 1. |
- Guo, Betty P, et al.
(författare)
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Relapsing fever Borrelia binds to neolacto glycans and mediates rosetting of human erythrocytes.
- 2009
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 1091-6490 .- 0027-8424. ; 106:46, s. 19280-5
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Tidskriftsartikel (refereegranskat)abstract
- A hallmark of acute relapsing fever borreliosis is severe bacteremia. Some Borrelia species, such as B. duttonii and B. crocidurae, associate with erythrocytes and induce aggregation recognized as erythrocyte rosetting. Erythrocyte rosettes contribute to disease severity by increased tissue invasiveness (such as invasion of CNS and encephalitis), hemorrhaging, and reduced blood flow in affected microcapillaries. Here we report that relapsing fever Borrelia binds to neolacto (Galbeta4GlcNAcbeta3Galbeta4Glcbeta1)-carrying glycoconjugates that are present on human erythrocytes. This interaction is of low affinity but is compensated for by the multivalency of neo-lacto-oligosaccharides on the erythrocyte cell surface. Hence, the protein-carbohydrate interaction is dependent on multivalent neolacto-glycans to mediate binding.
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| 2. |
- Johansson, Petra, 1974, et al.
(författare)
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Interaction of Helicobacter pylori with sialylated carbohydrates: the dependence on different parts of the binding trisaccharide Neu5Ac{alpha}3Gal{beta}4GlcNAc.
- 2005
-
Ingår i: Glycobiology. - : Oxford University Press (OUP). - 0959-6658 .- 1460-2423. ; 15:6, s. 625-36
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Tidskriftsartikel (refereegranskat)abstract
- We have recently shown that binding of Helicobacter pylori to sialylated carbohydrates is dependent on the presence of the carboxyl group and the glycerol chain of Neu5Ac. In this work we investigated the importance of GlcNAc in the binding trisaccharide Neu5Acalpha3Galbeta4GlcNAc and the role of the N-acetamido groups of both Neu5Ac and GlcNAc. An important part of the project was epitope dissection, that is chemical derivatizations of the active carbohydrate followed by binding studies. In addition we used a panel of various unmodified carbohydrate structures in the form of free oligosaccharides or glycolipids. These were tested for binding by hemagglutination inhibition assay, TLC overlay tests, and a new quantitative approach using radiolabeled neoglycoproteins. The studies showed that the N-acetamido group of Neu5Ac is important for binding by H. pylori, whereas the same group of GlcNAc is not. In addition, Fuc attached to GlcNAc, as tested with sialyl-Lewis x, did not affect the binding. Free Neu5Ac was inactive as inhibitor, and Neu5Acalpha3Gal turned out to be active. The binding preference for neolacto structures was confirmed, although one strain also was inhibited by lacto chains. The combined results revealed that an intact Neu5Ac is crucial for the interactions with H. pylori. Parts of Gal also seem to be necessary, whereas the role of the GlcNAc is secondary. GlcNAc does influence binding, however, primarily serving as a guiding carrier for the binding epitope rather than being a part of the binding structure.
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| 3. |
- Barone, Angela, et al.
(författare)
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Structural Complexity of Non-acid Glycosphingolipids in Human Embryonic Stem Cells Grown under Feeder-free Conditions.
- 2013
-
Ingår i: The Journal of biological chemistry. - 1083-351X. ; 288:14, s. 10035-50
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Tidskriftsartikel (refereegranskat)abstract
- Due to their pluripotency and growth capability, there are great expectations for human embryonic stem cells, both as a resource for functional studies of early human development and as a renewable source of cells for use in regenerative medicine and transplantation. However, to bring human embryonic stem cells into clinical applications, their cell surface antigen expression and its chemical structural complexity have to be defined. In the present study, total non-acid glycosphingolipid fractions were isolated from two human embryonic stem cell lines (SA121 and SA181) originating from leftover in vitro fertilized human embryos, using large amounts of starting material (1 × 10(9) cells/cell line). The total non-acid glycosphingolipid fractions were characterized by antibody and lectin binding, mass spectrometry, and proton NMR. In addition to the globo-series and type 1 core chain glycosphingolipids previously described in human embryonic stem cells, a number of type 2 core chain glycosphingolipids (neo-lactotetraosylceramide, the H type 2 pentaosylceramide, the Le(x) pentaosylceramide, and the Le(y) hexaosylceramide) were identified as well as the blood group A type 1 hexaosylceramide. Finally, the mono-, di-, and triglycosylceramides were characterized as galactosylceramide, glucosylceramide, lactosylceramide, galabiaosylceramide, globotriaosylceramide, and lactotriaosylceramide. Thus, the glycan diversity of human embryonic stem cells, including cell surface immune determinants, is more complex than previously appreciated.
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| 4. |
- Benktander, John, et al.
(författare)
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Redefinition of the carbohydrate binding specificity of Helicobacter pylori BabA adhesin.
- 2012
-
Ingår i: The Journal of biological chemistry. - 1083-351X. ; 287:38, s. 31712-31724
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Tidskriftsartikel (refereegranskat)abstract
- Certain Helicobacter pylori strains adhere to the human gastric epithelium using the BabA adhesin (blood group antigen binding adhesin). All BabA-expressing H. pylori strains bind to the blood group O determinants on type 1 core chains, i.e. to the Lewis b antigen (Fucα2Galβ3(Fucα4)GlcNAc; Leb) and the H type 1 determinant (Fucα2Galβ3GlcNAc). Recently BabA strains have been categorized into those recognizing only Leb and H type 1 determinants (designated specialist strains), and those that also bind to A and B type 1 determinants (designated generalist strain). Here, the structural requirements for carbohydrate recognition by generalist and specialist BabA were further explored by binding of these types of strains to a panel of different glycosphingolipids. Three glycosphingolipids recognized by both specialist and generalist BabA were isolated from the small intestine of a blood group O pig, and characterized by mass spectrometry and proton NMR as H type 1 pentaglycosylceramide (Fucα2Galβ3GlcNAcβ3Galβ4Glcβ1Cer), Globo H hexaglycosylceramide (Fucα2Galβ3GalNAcβ3Galα4Galβ4Glcβ1Cer), and a mixture of three complex glycosphingolipids (Fucα2Galβ4GlcNAcβ6(Fucα2Galβ3GlcNAcβ3)Galβ3GlcNAcβ3Galβ4Glcβ1Cer, Fucα2Galβ3GlcNAcβ6(Fucα2Galβ3GlcNAcβ3)Galβ3GlcNAcβ3Galβ4Glcβ1Cer and Fucα2Galβ4(Fucα3)GlcNAcβ6(Fucα2Galβ3GlcNAcβ3)Galβ3GlcNAcβ3Galβ4Glcβ1Cer). In addition to the binding of both strains to the Globo H hexaglycosylceramide, i.e. a blood group O determinant on a type 4 core chain, the generalist strain bound to the Globo A heptaglycosylceramide (GalNAcα3(Fucα2)Galβ3GalNAcβ3Galα4Galβ4Glcβ1Cer), i.e. a blood group A determinant on a type 4 core chain. The binding of BabA to the two sets of isoreceptors is due to onformational similarities of the terminal disaccharides of H type 1 and Globo H, and of the terminal trisaccharides of A type 1 and Globo A.
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| 5. |
- Benktander, John, et al.
(författare)
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The Repertoire of Glycosphingolipids Recognized by Vibrio cholerae
- 2013
-
Ingår i: Plos One. - : Public Library of Science (PLoS). - 1932-6203. ; 8:1
-
Tidskriftsartikel (refereegranskat)abstract
- The binding of cholera toxin to the ganglioside GM1 as the initial step in the process leading to diarrhea is nowadays textbook knowledge. In contrast, the knowledge about the mechanisms for attachment of Vibrio cholerae bacterial cells to the intestinal epithelium is limited. In order to clarify this issue, a large number of glycosphingolipid mixtures were screened for binding of El Tor V. cholerae. Several specific interactions with minor complex non-acid glycosphingolipids were thereby detected. After isolation of binding-active glycosphingolipids, characterization by mass spectrometry and proton NMR, and comparative binding studies, three distinct glycosphingolipid binding patterns were defined. Firstly, V. cholerae bound to complex lacto/neolacto glycosphingolipids with the GlcNAcβ3Galβ4GlcNAc sequence as the minimal binding epitope. Secondly, glycosphingolipids with a terminal Galα3Galα3Gal moiety were recognized, and the third specificity was the binding to lactosylceramide and related compounds. V. cholerae binding to lacto/neolacto glycosphingolipids, and to the other classes of binding-active compounds, remained after deletion of the chitin binding protein GbpA. Thus, the binding of V. cholerae to chitin and to lacto/neolacto containing glycosphingolipids represents two separate binding specificities.
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| 6. |
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| 7. |
- Ciopraga, J, et al.
(författare)
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Isolectins from Solanum tuberosum with different detailed carbohydrate binding specificities: unexpected recognition of lactosylceramide by N-acetyllactosamine-binding lectins.
- 2000
-
Ingår i: Journal of biochemistry. - 0021-924X. ; 128:5, s. 855-67
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Tidskriftsartikel (refereegranskat)abstract
- Glycosphingolipid recognition by two isolectins from Solanum tuberosum was compared by the chromatogram binding assay. One lectin (PL-I) was isolated from potato tubers by affinity chromatography, and identified by MALDI-TOF mass spectrometry as a homodimer with a subunit molecular mass of 63,000. The other (PL-II) was a commercial lectin, characterized as two homodimeric isolectins with subunit molecular masses of 52,000 and 55,000, respectively. Both lectins recognized N-acetyllactosamine-containing glycosphingolipids, but the fine details of their carbohydrate binding specificities differed. PL-II preferentially bound to glycosphingolipids with N-acetyllactosamine branches, as Galbeta4GlcNAcbeta6(Galbeta4GlcNAcbeta3)Galbeta4Glcbeta1C er. PL-I also recognized this glycosphingolipid, but bound equally well to the linear glycosphingolipid Galbeta4GlcNAcbeta3Galbeta4GlcNAcbeta3Galbeta4Glcbeta1Cer. Neolactotetraosylceramide and the B5 pentaglycosylceramide were also bound by PL-I, while other glycosphingolipids with only one N-acetyllactosamine unit were non-binding. Surprisingly, both lectins also bound to lactosylceramide, with an absolute requirement for sphingosine and non-hydroxy fatty acids. The inhibition of binding to both lactosylceramide and N-acetyllactosamine-containing glycosphingolipids by N-acetylchitotetraose suggests that lactosylceramide is also accomodated within the N-acetylchitotetraose/N-acetyllactosamine-binding sites of the lectins. Through docking of glycosphingolipids onto a three-dimensional model of the PL-I hevein binding domain, a Galbeta4GlcNAcbeta3Galbeta4 binding epitope was defined. Furthermore, direct involvement of the ceramide in the binding of lactosylceramide was suggested.
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| 8. |
- Coddens, Annelies, et al.
(författare)
-
Erythrocyte and Porcine Intestinal Glycosphingolipids Recognized by F4 Fimbriae of Enterotoxigenic Escherichia coli.
- 2011
-
Ingår i: PloS One. - : Public Library of Science (PLoS). - 1932-6203. ; 6:9
-
Tidskriftsartikel (refereegranskat)abstract
- Enterotoxigenic F4-fimbriated Escherichia coli is associated with diarrheal disease in neonatal and postweaning pigs. The F4 fimbriae mediate attachment of the bacteria to the pig intestinal epithelium, enabling an efficient delivery of diarrhea-inducing enterotoxins to the target epithelial cells. There are three variants of F4 fimbriae designated F4ab, F4ac and F4ad, respectively, having different antigenic and adhesive properties. In the present study, the binding of isolated F4ab, F4ac and F4ad fimbriae, and F4ab/ac/ad-fimbriated E. coli, to glycosphingolipids from erythrocytes and from porcine small intestinal epithelium was examined, in order to get a comprehensive view of the F4-binding glycosphingolipids involved in F4-mediated hemagglutination and adhesion to the epithelial cells of porcine intestine. Specific interactions between the F4ab, F4ac and F4ad fimbriae and both acid and non-acid glycosphingolipids were obtained, and after isolation of binding-active glycosphingolipids and characterization by mass spectrometry and proton NMR, distinct carbohydrate binding patterns were defined for each fimbrial subtype. Two novel glycosphingolipids were isolated from chicken erythrocytes, and characterized as GalNAcα3GalNAcß3Galß4Glcß1Cer and GalNAcα3GalNAcß3Galß4GlcNAcß3Galß4Glcß1Cer. These two compounds, and lactosylceramide (Galß4Glcß1Cer) with phytosphingosine and hydroxy fatty acid, were recognized by all three variants of F4 fimbriae. No binding of the F4ad fimbriae or F4ad-fimbriated E. coli to the porcine intestinal glycosphingolipids occurred. However, for F4ab and F4ac two distinct binding patterns were observed. The F4ac fimbriae and the F4ac-expressing E. coli selectively bound to galactosylceramide (Galß1Cer) with sphingosine and hydroxy 24:0 fatty acid, while the porcine intestinal glycosphingolipids recognized by F4ab fimbriae and the F4ab-fimbriated bacteria were characterized as galactosylceramide, sulfatide (SO(3)-3Galß1Cer), sulf-lactosylceramide (SO(3)-3Galß4Glcß1Cer), and globotriaosylceramide (Galα4Galß4Glcß1Cer) with phytosphingosine and hydroxy 24:0 fatty acid. Finally, the F4ad fimbriae and the F4ad-fimbriated E. coli, but not the F4ab or F4ac subtypes, bound to reference gangliotriaosylceramide (GalNAcß4Galß4Glcß1Cer), gangliotetraosylceramide (Galß3GalNAcß4Galß4Glcß1Cer), isoglobotriaosylceramide (Galα3Galß4Glcß1Cer), and neolactotetraosylceramide (Galß4GlcNAcß3Galß4Glcß1Cer).
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| 9. |
- Coddens, Annelies, et al.
(författare)
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Recognition of blood group ABH type 1 determinants by the FedF adhesin of F18-fimbriated Escherichia coli.
- 2009
-
Ingår i: The Journal of biological chemistry. - 0021-9258. ; 284:15, s. 9713-26
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Tidskriftsartikel (refereegranskat)abstract
- F18-fimbriated Escherichia coli are associated with porcine postweaning diarrhea and edema disease. Adhesion of F18-fimbriated bacteria to the small intestine of susceptible pigs is mediated by the minor fimbrial subunit FedF. However, the target cell receptor for FedF has remained unidentified. Here we report that F18-fimbriated E. coli selectively interact with glycosphingolipids having blood group ABH determinants on type 1 core, and blood group A type 4 heptaglycosylceramide. The minimal binding epitope was identified as the blood group H type 1 determinant (Fucalpha2Galbeta3GlcNAc), while an optimal binding epitope was created by addition of the terminal alpha3-linked galactose or N-acetylgalactosamine of the blood group B type 1 determinant (Galalpha3(Fucalpha2)Galbeta3GlcNAc) and the blood group A type 1 determinant (GalNAcalpha3(Fucalpha2)-Galbeta3GlcNAc). To assess the role of glycosphingolipid recognition by F18-fimbriated E. coli in target tissue adherence, F18-binding glycosphingolipids were isolated from the small intestinal epithelium of blood group O and A pigs and characterized by mass spectrometry and proton NMR. The only glycosphingolipid with F18-binding activity of the blood group O pig was an H type 1 pentaglycosylceramide (Fucalpha2Galbeta3GlcNAc-beta3Galbeta4Glcbeta1Cer). In contrast, the blood group A pig had a number of F18-binding glycosphingolipids, characterized as A type 1 hexaglycosylceramide (GalNAcalpha3(Fucalpha2)Galbeta3GlcNAcbeta3Galbeta4Glcbeta1Cer), A type 4 heptaglycosylceramide (GalNAcalpha3(Fucalpha2)Galbeta3GalNAcbeta3Galalpha4Galbeta4Glcbeta1Cer), A type 1 octaglycosylceramide (GalNAcalpha3(Fucalpha2)Galbeta3GlcNAcbeta3Galbeta3GlcNAcbeta3Galbeta4Glcbeta1Cer), and repetitive A type 1 nonaglycosylceramide (GalNAcalpha3(Fucalpha2)Galbeta3GalNAcalpha3-(Fucalpha2)Galbeta3GlcNAcbeta3Galbeta4Glcbeta1Cer). No blood group antigen-carrying glycosphingolipids were recognized by a mutant E. coli strain with deletion of the FedF adhesin, demonstrating that FedF is the structural element mediating binding of F18-fimbriated bacteria to blood group ABH determinants.
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| 10. |
- Diswall, Mette, 1979, et al.
(författare)
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Glycolipid studies in small intestine and pancreas of alpha1,3-galactosyltransferase knockout miniature swine: alpha1,3GALT-KO animals lack alphaGAL antigens and contain novel blood group H compounds.
- 2008
-
Ingår i: Transplantation proceedings. - : Elsevier BV. - 0041-1345. ; 40:2, s. 543-6
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: To avoid hyperacute rejection of xeno-organs, alpha1,3-galactosyltransferase knock-out (GalT-KO) pigs have been produced. However, Galalpha1,3Gal (Gal) determinant elimination may expose cryptic carbohydrate antigens and/or generate new antigens that might interfere with the human immune response. METHODS: Glycolipids isolated from small intestine and pancreas of two GalT-KO and one wild-type (WT) pig were tested for immune reactivity with antibodies on thin-layer chromatograms after separation by high-performance liquid chromatography, and selected fractions were analysed by proton NMR spectroscopy. RESULTS: Immunostaining using purified human anti-Gal Abs revealed that tissues from WT animals express large amounts of Gal-antigens whereas GalT-KO tissues lacked these antigens. Proton NMR spectroscopy on small intestine fractions revealed both linear and branched nona- and decaglycosylceramides, respectively, with terminal Gal-epitopes. In corresponding GalT-KO fractions, Gal-epitopes seemed to be replaced by terminal alpha1,2fucoses. Two novel branched blood group H compounds was found in the GalT-KO intestine. CONCLUSIONS: The structural complexity of alphaGal-terminating antigens in the WT organs is very high. Knockout of alpha1,3GalT by gene-targeting results in elimination of Gal-determinants. In addition structurally novel alpha1,2fucose-terminated blood group H compounds were identified in the GalT-KO tissue. These compounds are not expected to be recognized by the human immune system.
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| 11. |
- Diswall, Mette, 1979, et al.
(författare)
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Structural characterization of alpha1,3-galactosyltransferase knockout pig heart and kidney glycolipids and their reactivity with human and baboon antibodies.
- 2010
-
Ingår i: Xenotransplantation. - : Wiley. - 1399-3089 .- 0908-665X. ; 17:1, s. 48-60
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: alpha1,3-galactosyltranferase knockout (GalT-KO) pigs have been established to avoid hyperacute rejection in GalT-KO pig-to-human xenotransplantation. GalT-KO pig heart and kidney glycolipids were studied focusing on elimination of Gal-antigens and whether novel antigens would appear. Non-human primates are used as pre-clinical transplantation experimental models. Therefore, sera from baboons transplanted with GalT-KO hearts were compared with human serum regarding reactivity with pig glycolipids. METHODS: Neutral and acidic glycolipids were isolated from GalT-KO and WT pig hearts and kidneys. Glycolipid immune reactivity was tested on TLC plates using human affinity-purified anti-Gal Ig, anti-blood group monoclonal antibodies, lectins, and human serum as well as baboon serum collected before and after GalT-KO pig heart transplantations. Selected glycolipid fractions, isolated by HPLC, were structurally characterized by mass spectrometry and proton NMR spectroscopy. RESULTS: GalT-KO heart and kidney lacked alpha3Gal-terminated glycolipids completely. Levels of uncapped N-acetyllactosamine precursor compounds, blood group H type 2 core chain compounds, the P1 antigen and the x(2) antigen were increased. Human serum antibodies reacted with Gal-antigens and N-glycolylneuraminic acid (NeuGc) in WT organs of which only the NeuGc reactivity remained in the GalT-KO tissues. A clear difference in reactivity between baboon and human antibodies with pig glycolipids was found. This was most pronounced for acidic, not yet identified, compounds in GalT-KO organs which were less abundant or lacking in the corresponding WT tissues. CONCLUSIONS: GalT-KO pig heart and kidney completely lacked Gal glycolipid antigens whilst glycolipids synthesized by competing pathways were increased. Baboon and human serum antibodies showed a different reactivity pattern to pig glycolipid antigens indicating that non-human primates have limitations as a human pre-clinical model for immune rejection studies.
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| 12. |
- Diswall, Mette, 1979, et al.
(författare)
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Studies on glycolipid antigens in small intestine and pancreas from alpha1,3-galactosyltransferase knockout miniature swine.
- 2007
-
Ingår i: Transplantation. - : Ovid Technologies (Wolters Kluwer Health). - 0041-1337. ; 84:10, s. 1348-56
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: To avoid hyperacute rejection of xeno-organs, alpha1,3-galactosyltransferase knockout (GalT-KO) pigs have been produced. Galalpha1,3Gal determinant elimination may expose cryptic carbohydrate antigens and/or generate new antigens. This is the first biochemical study of carbohydrate antigens in GalT-KO pig organs. METHODS: Neutral and acidic glycolipids were isolated from small intestine and pancreas of two GalT-KO and one wild-type (WT) pig. Glycolipid immune reactivity was tested on thin-layer chromatograms. Small intestine neutral glycolipids were separated by high-performance liquid chromatography and selected fractions were analyzed by proton nuclear magnetic resonance spectroscopy. Total gangliosides were quantified on thin-layer chromatograms and in microtiter wells. RESULTS: Using Galalpha1,3nLc4 glycolipid reference, total Galalpha1,3Gal glycolipid antigens in the WT animal was estimated at about 30 microg (small intestine) and 3 microg (pancreas) per gram of dry tissue. Galalpha1,3Gal determinants were not detected in GalT-KO tissues at a detection limit of less than 0.25% (small intestine) and 0.5% (pancreas) of the WT tissues. Isoglobotriaosylceramide (iGb3) was absent but trace amounts of Fuc-iGb3 was found in both GalT-KO and WT pig small intestine. Blood group H type 2 core saccharide compounds were increased in GalT-KO pancreas. Total amount of gangliosides was decreased in GalT-KO tissues. The alpha1,3-galactosyltransferase acceptor, N-acetyllactosamine determinant, was not increased in GalT-KO tissues. Human serum antibodies reacted with WT organ Galalpha1,3Gal antigens and gangliosides, of which the ganglioside reactivity remained in GalT-KO tissues. CONCLUSIONS: Knockout of porcine alpha1,3-galactosyltransferase gene results in elimination of Galalpha1,3Gal-terminated glycolipid compounds. GalT-KO genetic modification did not produce new compensatory glycolipid compounds reactive with human serum antibodies.
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| 13. |
- Diswall, Mette, 1979, et al.
(författare)
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The alpha1,3GalT knockout/alpha1,2FucT transgenic pig does not appear to have an advantage over the alpha1,3GalT knockout pig with respect to glycolipid reactivity with human serum antibodies.
- 2014
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Ingår i: Xenotransplantation. - : Wiley. - 1399-3089 .- 0908-665X. ; 21:1, s. 57-71
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Tidskriftsartikel (refereegranskat)abstract
- The human H-transferase (α2FucT) was introduced in Gal-negative pigs to produce pig organs not only free from Gal-antigens, but also in which the uncapped N-acetyllactosamine precursor had been transformed into non-xenogenic blood group H type 2 compounds. This work is the first descriptive analysis of glycolipids from the GalT-KO/FucT-TG pig. The aim was to investigate the cell membrane antigens in GalT-KO/FucT-TG tissues to explore its efficacy as an organ donor. Also, detailed knowledge on the correlation between the cellular glycosyltransferase configuration and the resulting carbohydrate phenotype expression is valuable from a basic glycobiological perspective.
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| 14. |
- Fagerberg, David, et al.
(författare)
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Novel Leb-like Helicobacter pylori-binding glycosphingolipid created by the expression of human alpha-1,3/4-fucosyltransferase in FVB/N mouse stomach.
- 2009
-
Ingår i: Glycobiology. - : Oxford University Press (OUP). - 1460-2423 .- 0959-6658. ; 19:2, s. 182-91
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Tidskriftsartikel (refereegranskat)abstract
- The "Le(b) mouse" was established as a model for investigations of the molecular events following Le(b)-mediated adhesion of Helicobacter pylori to the gastric epithelium. By the expression of a human alpha-1,3/4-fucosyltransferase in the gastric pit cell lineage of FVB/N transgenic mice, a production of Le(b) glycoproteins in gastric pit and surface mucous cells was obtained in this "Le(b) mouse," as demonstrated by binding of monoclonal anti-Le(b) antibodies. To explore the effects of the human alpha-1,3/4-fucosyltransferase on glycosphingolipid structures, neutral glycosphingolipids were isolated from stomachs of transgenic alpha-1,3/4-fucosyltransferase-expressing mice. A glycosphingolipid recognized by BabA-expressing H. pylori was isolated and characterized by mass spectrometry and proton NMR as Fuc alpha 2Gal beta 3(Fuc alpha 4)GalNAc beta 4 Gal beta 4 Glc beta 1Cer, i.e., a novel Le(b)-like glycosphingolipid on a ganglio core. In addition, two other novel glycosphingolipids were isolated from the mouse stomach epithelium that were found to be nonbinding with regard to H. pylori. The first was a pentaglycosylceramide, GalNAc beta 3 Gal alpha 3(Fuc alpha 2)Gal beta 4 Glc beta 1Cer, in which the isoglobotetrasaccharide has been combined with Fuc alpha 2 to yield an isoglobotetraosylceramide with an internal blood group B determinant. The second one was an elongated fucosyl-gangliotetraosylceramide, GalNAc beta 3(Fuc alpha 2)Gal beta 3GalNAc beta 4Gal beta 4 Glc beta 1Cer.
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| 15. |
- Gustafsson, Anki, et al.
(författare)
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Pichia pastoris-produced mucin-type fusion proteins with multivalent O-glycan substitution as targeting molecules for mannose-specific receptors of the immune system
- 2011
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Ingår i: Glycobiology. - : Oxford University Press (OUP). - 0959-6658 .- 1460-2423. ; 21:8, s. 1071-1086
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Tidskriftsartikel (refereegranskat)abstract
- Mannose-binding proteins like the macrophage mannose receptor (MR), the dendritic cell-specific intercellular adhesion molecule-3 grabbing non-integrin (DC-SIGN) and mannose-binding lectin (MBL) play crucial roles in both innate and adaptive immune responses. Immunoglobulin fusion proteins of the P-selectin glycoprotein ligand-1 (PSGL-1/mIgG2b) carrying mostly O-glycans and, as a control, the a1-acid glycoprotein (AGP/mIgG2b) carrying mainly N-linked glycans were stably expressed in the yeast Pichia pastoris. P. pastoris-produced PSGL-1/mIgG2b was shown to carry O-glycans that mediated strong binding to mannose-specific lectins in a lectin array and were susceptible to cleavage by a-mannosidases including an a1,2- but not an a1,6-mannosidase. Electrospray ionization - ion trap mass spectrometry (ESI-MS) confirmed the presence of O-glycans containing up to 9 hexoses with the penta- and hexasaccharides being the predominant ones. a1,2- and a1,3-linked, but not a1,6-linked, mannose residues were detected by 1H-nuclear magnetic resonance (1H-NMR) spectroscopy confirming the results of the mannosidase cleavage. The apparent equilibrium dissociation constants for binding of PNGase F-treated mannosylated PSGL-1/mIgG2b to MR, DC-SIGN and MBL were shown by surface plasmon resonance to be 126, 56 and 16 nM, respectively. In conclusion, PSGL-1/mIgG2b expressed in P. pastoris carried O-glycans mainly comprised of a-linked mannoses and with up to nine residues. It bound mannose-specific receptors with high apparent affinity and may become a potent targeting molecule for these receptors in vivo.
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| 16. |
- Hallén, Ulrika, et al.
(författare)
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Glycolipid binding epitopes involved in adherence of the periodontitis-associated bacterium Porphyromonas gingivalis.
- 2008
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Ingår i: Glycoconjugate journal. - : Springer Science and Business Media LLC. - 0282-0080 .- 1573-4986. ; 25:6, s. 561-572
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Tidskriftsartikel (refereegranskat)abstract
- The ability of the periodontal pathogen Porphyromonas gingivalis to use different glycolipid structures as receptors has previously been demonstrated. The bacterium adhered to acid and nonacid glycolipids originating from human organs and to nonacid glycolipids of porcine origin. The aim of the present study was to analyze these binding epitopes by structural characterization. Glycolipid fractions with positive bacterial binding from e.g. human and porcine origin, were purified by the high performance liquid chromatography technique and thereafter used in bacterial overlay assays with (35)S-labeled P. gingivalis. Purified fractions with positive binding were structurally characterized by proton nuclear magnetic resonance spectroscopy. Complementing thin-layer chromatograms and bacterial overlay assays with pure reference glycolipid fractions and competition experiments with lactose were performed to define potential receptors. The P. gingivalis binding epitopes, including cerebrosides with nonhydroxy fatty acids, lactosylceramide with hydroxy fatty acids, sulfatides, lacto-, neolacto- and gangliotetraosylceramides, are in several instances similar to those found for other bacteria, e.g. H. pylori, H. influenzae and N. meningitidis. In addition P. gingivalis also bound to the Galalpha4Gal epitope of the globo series of glycolipids. In the future these results may be valuable for development of new treatment strategies, such as anti-adhesion therapies and vaccines specifically directed against P. gingivalis infection.
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| 17. |
- Holmner, Åsa, et al.
(författare)
-
Novel binding site identified in a hybrid between cholera toxin and heat-labile enterotoxin: 1.9 A crystal structure reveals the details.
- 2004
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Ingår i: Structure (London, England : 1993). - : Elsevier BV. - 0969-2126 .- 1878-4186. ; 12:9, s. 1655-67
-
Tidskriftsartikel (refereegranskat)abstract
- A hybrid between the B subunits of cholera toxin and Escherichia coli heat-labile enterotoxin has been described, which exhibits a novel binding specificity to blood group A and B type 2 determinants. In the present investigation, we have determined the crystal structure of this protein hybrid, termed LCTBK, in complex with the blood group A pentasaccharide GalNAcalpha3(Fucalpha2)Galbeta4(Fucalpha3)GlcNAcbeta, confirming not only the novel binding specificity but also a distinct new oligosaccharide binding site. Binding studies revealed that the new specificity can be ascribed to a single mutation (S4N) introduced into the sequence of Escherichia coli heat-labile enterotoxin. At a resolution of 1.9 A, the new binding site is resolved in excellent detail. Main features include a complex network of water molecules, which is well preserved by the parent toxins, and an unexpectedly modest contribution to binding by the critical residue Asn4, which interacts with the ligand only via a single water molecule.
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| 18. |
- Hugosson, S, et al.
(författare)
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Glycosphingolipid binding specificities of Neisseria meningitidis and Haemophilus influenzae: detection, isolation, and characterization of a binding-active glycosphingolipid from human oropharyngeal epithelium.
- 1998
-
Ingår i: Journal of biochemistry. - 0021-924X. ; 124:6, s. 1138-52
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Tidskriftsartikel (refereegranskat)abstract
- The glycosphingolipid binding specificities of Haemophilus influenzae and Neisseria meningitidis were investigated as to the binding of radiolabeled bacteria to glycosphingolipids on thin-layer chromatograms. Thereby, similar binding profiles, for the binding of the two bacteria to lactosylceramide, isoglobotriaosylceramide, gangliotriaosylceramide, gangliotetraosylceramide, lactotetraosylceramide, neolactotetraosylceramide, and sialylneolactohexaosylceramide, were obtained. On a closer view the binding preferences of the bacteria could be differentiated into three groups. The first specificity is recognition of lactosylceramide. The second specificity is binding to gangliotriaosylceramide and gangliotetraosylceramide, since conversion of the acetamido group of the N-acetylgalactosamine of gangliotriaosylceramide and gangliotetraosylceramide to an amine prevented the binding of the bacteria, and thus the binding to these two glycosphingolipids represents a separate specificity from lactosylceramide recognition. Preincubation of H. influenzae with neolactotetraose inhibited the binding to neolactotetraosylceramide, while the binding to lactosylceramide, gangliotetraosylceramide, or lactotetraosylceramide was unaffected. Thus, the third binding specificity is represented by neolactotetraosylceramide, and involves recognition of other neolacto series glycosphingolipids with linear N-acetyllactosamine chains, such as sialyl-neolactohexaosylceramide. The relevance of the detected binding specificities for adhesion to target cells was addressed as to the binding of the bacteria to glycosphingolipids from human granulocytes, epithelial cells of human nasopharyngeal tonsils and human plexus choroideus. Binding-active neolactotetraosylceramide was thereby detected in human granulocytes and the oropharyngeal epithelium.
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| 19. |
- Jansson, Lena, 1979, et al.
(författare)
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No direct binding of the heat-labile enterotoxin of Escherichia coli to E. coli lipopolysaccharides.
- 2009
-
Ingår i: Glycoconjugate journal. - : Springer Science and Business Media LLC. - 1573-4986 .- 0282-0080.
-
Tidskriftsartikel (refereegranskat)abstract
- A novel carbohydrate binding site recognizing blood group A and B determinants in a hybrid of cholera toxin and Escherichia coli heat-labile enterotoxin B-subunits (termed LCTBK) has previously been described, and also the native heat-labile enterotoxin bind to some extent to blood group A/B terminated glycoconjugates. The blood group antigen binding site is located at the interface of the B-subunits. Interestingly, the same area of the B-subunits has been proposed to be involved in binding of the heat-labile enterotoxin to lipopolysaccharides on the bacterial cell surface. Binding of the toxin to lipopolysaccharides does not affect the GM1 binding capacity. The present study aimed at characterizing the relationship between the blood group A/B antigen binding site and the lipopolysaccharide binding site. However, no binding of the B-subunits to E. coli lipopolysaccharides in microtiter wells or on thin-layer chromatograms was obtained. Incubation with lipopolysaccharides did not affect the binding of the B-subunits of heat-labile enterotoxin of human isolates to blood group A-carrying glycosphingolipids, indicating that the blood group antigen site is not involved in LPS binding. However, the saccharide competition experiments showed that GM1 binding reduced the affinity for blood group A determinants and vice versa, suggesting that a concurrent occupancy of the two binding sites does not occur. The latter finding is related to a connection between the blood group antigen binding site and the GM1 binding site through residues interacting with both ligands.
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| 20. |
- Karlsson, Karl-Anders, 1935, et al.
(författare)
-
Microbial interaction with animal cell surface carbohydrates.
- 1992
-
Ingår i: APMIS. Supplementum. - 0903-465X. ; 27, s. 71-83
-
Tidskriftsartikel (refereegranskat)abstract
- Microbes have selected primarily carbohydrates for attachment to host animal cells. Recent studies have revealed essential characteristics in the recognition of receptor carbohydrates. Of importance is the property of recognizing also sequences placed inside an oligosaccharide chain, which differs from most animal antibodies. This is the basis for series of isoreceptors with the minimum receptor sequence in common but with separate neighbouring groups. There are families of microbial ligands that show different preferences for members within one series of isoreceptors, indicating only slight differences in the complementary binding sites of the proteins. Such differences may explain shifts in the selectivity of separate host tissues for infection. A second characteristic is the low affinity interaction often found where simple receptor-containing saccharides are unable to inhibit attachment. Technical possibilities are rapidly developing for the design of synthetic receptor analogues to be used in the therapy of clinical infections. This is urgently needed in cases where no rational therapy exists today.
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| 21. |
- Karlsson, Karl-Anders, 1935, et al.
(författare)
-
Unexpected carbohydrate cross-binding by Escherichia coli heat-labile enterotoxin. Recognition of human and rabbit target cell glycoconjugates in comparison with cholera toxin.
- 1996
-
Ingår i: Bioorganic & medicinal chemistry. - 0968-0896. ; 4:11, s. 1919-28
-
Tidskriftsartikel (refereegranskat)abstract
- The bacterial protein enterotoxins, cholera toxin (CT) of Vibrio cholerae and heat-labile toxin (LT) of Escherichia coli, induce diarrhea by enhancing the secretory activity of the small intestine of man and rabbit (animal model). This physiological effect is mediated by toxin binding to a glycolipid receptor, the ganglioside GM1, Gal beta 3GalNAc beta 4(NeuAc alpha 3)GAl beta 4Glc beta 1Cer. However, LT, but not CT, was recently shown by us to bind also to paragloboside, Gal beta 4GlcNAc beta 3Gal beta 4Glc beta 1Cer, identified in the target cells. By molecular modeling of this tetrasaccharide in the known binding site of LT, the saccharide-peptide interaction was shown to be limited to the terminal disaccharide (N-acetyllactosamine). This sequence is expressed in many glycoconjugates, and we have therefore assayed glycolipids and glycoproteins prepared from the target tissues. In addition to paragloboside, receptor activity for LT was detected in glycoproteins of human origin and in polyglycosylceramides of rabbit. However, CT bound only to GM1. Two variants of LT with slightly different sequences, human (hLT) and porcine (pLT), were identical in their binding to target glycoproteins and polyglycosylceramides, but different regarding paragloboside, which was positive for pLT but negative for hLT. This difference is discussed on basis of modeling, taking in view the difference at position 13, with Arg in pLT and His in hLT. Although N-acetyllactosamine is differently recognized in form of paragloboside by the two toxin variants, we speculate that this sequence in human glycoproteins and rabbit polyglycosylceramides is the basis for the common binding. Much work remains, however, to clear up up this unexpected sophistication in target recognition.
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| 22. |
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| 23. |
- Lanne, B, et al.
(författare)
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Glycoconjugate receptors for P-fimbriated Escherichia coli in the mouse. An animal model of urinary tract infection.
- 1995
-
Ingår i: The Journal of biological chemistry. - 0021-9258. ; 270:15, s. 9017-25
-
Tidskriftsartikel (refereegranskat)abstract
- Glycosphingolipids were isolated from kidneys, urethers, and bladders (including urethrae) of C3H/HeN mice. Binding was studied of a clinical isolate and recombinant strains of uropathogenic P-fimbriated Escherichia coli to these glycolipids. A series of receptor-active glycolipids with Gal alpha 4Gal in common, previously shown to be recognized by these bacteria, was identified by use of specific monoclonal antibodies, fast-atom bombardment and electron-impact mass spectrometry, and proton nuclear magnetic resonance spectroscopy: galabiosylceramide (Gal alpha 4Gal beta Cer), globotriaosylceramide (Gal alpha 4Gal beta 4Glc beta Cer), globoside (GalNAc beta 3Gal alpha 4Gal beta 4Glc beta Cer), the Forssman glycolipid (GalNAc alpha 3GalNAc beta 3Gal alpha 4Gal beta 4Glc beta Cer), Gal beta 4GlcNAc beta 6(Gal beta 3)GalNAc beta 3Gal alpha 4Gal beta 4Glc beta Cer, and Gal beta 4(Fuc alpha 3)GlcNAc beta 6(Gal beta 3)GalNAc beta 3Gal alpha 4Gal beta 4Glc beta Cer. The binding pattern for mouse kidney glycolipids differed from that for kidney glycolipids of man and monkey. In particular, the dominant 8-sugar glycolipid in the mouse was not detected in the primates. A second difference was found in the binding of E. coli to kidney glycoproteins on blots after electrophoresis; the mouse showed distinct receptor-active bands while human and monkey did not. These differences may be of relevance when using the mouse as a model for clinical urinary tract infection of man.
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| 24. |
- Miller-Podraza, Halina, 1948, et al.
(författare)
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Novel binding epitope for Helicobacter pylori found in neolacto carbohydrate chains: structure and cross-binding properties.
- 2005
-
Ingår i: The Journal of biological chemistry. - 0021-9258. ; 280:20, s. 19695-703
-
Tidskriftsartikel (refereegranskat)abstract
- Helicobacter pylori is a bacterium that colonizes the stomach of a majority of the global human population causing common gastric diseases like ulcers and cancer. It has an unusually complex pattern of binding to various host glycoconjugates including interaction with sialylated, sulfated, and fucosylated sequences. The present study describes an additional binding epitope comprising the neolacto internal sequence of GlcNAcbeta3-Galbeta4GlcNAcbeta. The binding was detected on TLC plates as an interaction with a seven-sugar ganglioside of rabbit thymus. The glycolipid was purified and characterized as Neu5Gcalpha3Galbeta4GlcNAcbeta3Galbeta4GlcNAcbeta3-Galbeta4Glcbeta1Cer with less than 10% of the fraction carrying a repeated lacto (type-1) core chain, Galbeta3Glc-NAcbeta3Galbeta3GlcNAcbeta. After stepwise chemical and enzymatic degradation and structural analysis of products the strongest binder was found to be the pentaglycosylceramide GlcNAcbeta3Galbeta4GlcNAcbeta3Galbeta4Glcbeta1-Cer, whereas the hexa- and tetraglycosylceramides were less active, and the trihexosylceramide was inactive. Further studies revealed that the terminal GlcNAcbeta of the pentaglycosylceramide may be exchanged for either GalNAcbeta3, GalNAcalpha3, or Galalpha3 without loss of the activity. Calculated minimum energy conformers of these four isoreceptors show a substantial topographical similarity suggesting that this binding is a result of a molecular mimicry. Although the glycoconjugate composition of human gastric epithelial cells is not known in detail it is proposed that repeating N-acetyllactosamine units of glycoconjugates may serve as bacterial attachment sites in the stomach.
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| 25. |
- Miller-Podraza, Halina, 1948, et al.
(författare)
-
Studies on gangliosides with affinity for Helicobacter pylori: binding to natural and chemically modified structures.
- 2004
-
Ingår i: Glycobiology. - : Oxford University Press (OUP). - 0959-6658 .- 1460-2423. ; 14:3, s. 205-17
-
Tidskriftsartikel (refereegranskat)abstract
- Helicobacter pylori, like many other microbes, has the ability to bind to carbohydrate epitopes. Several sugar sequences have been reported as active for the bacterium, including some neutral, sulfated, and sialylated structures. We investigated structural requirements for the sialic acid-dependent binding using a number of natural and chemically modified gangliosides. We have chosen for derivatization studies two kinds of binding-active glycolipids, the simple ganglioside S-3PG (Neu5Ac alpha 3Gal beta 4GlcNAc beta 3Gal beta 4Glc beta 1Cer, sialylparagloboside) and branched polyglycosylceramides (PGCs) of human origin. The modifications included oxidation of the sialic acid glycerol chain, reduction of the carboxyl group, amidation of the carboxyl group, and lactonization. Binding experiments confirmed a preference of H. pylori for 3-linked sialic acid and penultimate 4-linked galactose. As expected, neolacto gangliosides (with Gal beta 4GlcNAc in the core structure) were active in our assays, whereas gangliosides with lacto (Gal beta 3GlcNAc) and ganglio (Gal beta 3GalNAc) carbohydrate chains were not. Negative binding results were also obtained for disialylparagloboside (with terminal NeuAc alpha 8NeuAc) and NeuAc alpha 6-containing glycolipids. Chemical studies revealed dependence of the binding on Neu5Ac and its glycerol and carboxyl side chains. Most of the derivatizations performed on these groups abolished the binding; however, some of the amide forms turned out to be active, and one of them (octadecylamide) was found to be an excellent binder. The combined data from molecular dynamics simulations indicate that the binding-active configuration of the terminal disaccharide of S-3PG is with the sialic acid in the anticlinal conformation, whereas in branched PGCs the same structural element most likely assumes the synclinal presentation.
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| 26. |
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| 27. |
- Roche, Niamh, 1969, et al.
(författare)
-
Helicobacter pylori and complex gangliosides.
- 2004
-
Ingår i: Infection and immunity. - 0019-9567. ; 72:3, s. 1519-29
-
Tidskriftsartikel (refereegranskat)abstract
- Recognition of sialic acid-containing glycoconjugates by the human gastric pathogen Helicobacter pylori has been repeatedly demonstrated. To investigate the structural requirements for H. pylori binding to complex gangliosides, a large number of gangliosides were isolated and characterized by mass spectrometry and proton nuclear magnetic resonance. Ganglioside binding of sialic acid-recognizing H. pylori strains (strains J99 and CCUG 17874) and knockout mutant strains with the sialic acid binding adhesin SabA or the NeuAcalpha3Galbeta4GlcNAcbeta3Galbeta4GlcNAcbeta-binding neutrophil-activating protein HPNAP deleted was investigated using the thin-layer chromatogram binding assay. The wild-type bacteria bound to N-acetyllactosamine-based gangliosides with terminal alpha3-linked NeuAc, while gangliosides with terminal NeuGcalpha3, NeuAcalpha6, or NeuAcalpha8NeuAcalpha3 were not recognized. The factors affecting binding affinity were identified as (i) the length of the N-acetyllactosamine carbohydrate chain, (ii) the branches of the carbohydrate chain, and (iii) fucose substitution of the N-acetyllactosamine core chain. While the J99/NAP(-) mutant strain displayed a ganglioside binding pattern identical to that of the parent J99 wild-type strain, no ganglioside binding was obtained with the J99/SabA(-) mutant strain, demonstrating that the SabA adhesin is the sole factor responsible for the binding of H. pylori bacterial cells to gangliosides.
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| 28. |
- Roche, Niamh, 1969, et al.
(författare)
-
Human gastric glycosphingolipids recognized by Helicobacter pylori vacuolating cytotoxin VacA.
- 2007
-
Ingår i: Microbes and infection / Institut Pasteur. - : Elsevier BV. - 1286-4579. ; 9:5, s. 605-14
-
Tidskriftsartikel (refereegranskat)abstract
- Many bacterial toxins utilize cell surface glycoconjugate receptors for attachment to target cells. In the present study the potential carbohydrate binding of Helicobacter pylori vacuolating cytotoxin VacA was investigated by binding to human gastric glycosphingolipids on thin-layer chromatograms. Thereby a distinct binding of the toxin to two compounds in the non-acid glycosphingolipid fraction was detected. The VacA-binding glycosphingolipids were isolated and characterized by mass spectrometry and proton NMR as galactosylceramide (Galbeta1Cer) and galabiosylceramide (Galalpha4Galbeta1Cer). Comparison of the binding preferences of the protein to reference glycosphingolipids from other sources showed an additional recognition of glucosylceramide (Glcbeta1Cer), lactosylceramide (Galbeta4Glcbeta1Cer) and globotriaosylceramide (Galalpha4Galbeta4Glcbeta1Cer). No binding to the glycosphingolipids recognized by the VacA holotoxin was obtained with a mutant toxin with deletion of the 37 kDa fragment of VacA (P58 molecule). Collectively our data show that the VacA cytotoxin is a glycosphingolipid binding protein, where the 37 kDa moiety is required for carbohydrate recognition. The ability to bind to short chain glycosphingolipids will position the toxin close to the cell membrane, which may facilitate toxin internalization.
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| 29. |
- Rojas, Gertrudis, et al.
(författare)
-
Light-chain shuffling results in successful phage display selection of functional prokaryotic - expressed antibody fragments to N-glycolyl GM3 ganglioside
- 2004
-
Ingår i: Journal of Immunological Methods. - : Elsevier BV. - 0022-1759. ; 293:1-2, s. 71-83
-
Tidskriftsartikel (refereegranskat)abstract
- Phage display technology makes it possible to introduce and rapidly screen diversity in antibody binding sites. Chain shuffling has been successfully used to humanize murine antibody fragments and also to obtain affinity matured variants. Here we report a different application of this method: the use of chain shuffling to overcome improper prokaryotic expression behavior of a hybridoma-derived single-chain antibody fragment. Construction and expression of such recombinant antibody fragments remain as empirical entities, hampered by the inability to express some antibody genes coming from eukaryotic cells in bacterial expression systems. Such problems are different for each combination of variable regions and can be serious enough to preclude the use of some hybridomas as sources of V regions to obtain recombinant antibody fragments. The particular binding properties and potential usefulness of some monoclonal antibodies make it highly desirable to bypass these technical limitations in order to develop smaller size therapeutic agents in the form of antibody fragments. The 14F7 mouse monoclonal antibody is one such attractive candidate due to its high specificity for the N-glycolyl GM3 ganglioside overexpressed in tumor cells and its ability to distinguish this antigen from closely related gangliosides like N-acetyl GM3. Our goal was to construct a phage-di splayed single-chain Fv antibody fragment derived from 14F7. After cloning the original variable regions from the 14F7 hybridoma in a phagemid vector, we were unable to detect either binding activity or even expression of antibody fragments in bacteria, despite repetitive efforts. We constructed light-chain shuffling libraries, from which functional antibody fragments were readily selected. These combined the original 14F7 heavy chain variable region with a wide variety of unrelated murine and human light-chain variable regions. New antibody fragments retained the valuable properties of the monoclonal antibody in terms of fine specificity, affinity and tumor recognition. They were readily produced by bacteria, either in phage-displayed form or as soluble molecules, and provided a panel of potentially useful variants for cancer diagnosis and immunotherapy. Chain shuffling and phage display were found to be useful strategies for selecting antibody fragments on the basis of both prokaryotic expression and antigen binding criteria. © 2004 Elsevier B.V. All rights reserved.
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| 30. |
- Satomaa, Tero, et al.
(författare)
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Analysis of the human cancer glycome identifies a novel group of tumor-associated N-acetylglucosamine glycan antigens.
- 2009
-
Ingår i: Cancer research. - 1538-7445. ; 69:14, s. 5811-9
-
Tidskriftsartikel (refereegranskat)abstract
- The cell surface is covered by a dense layer of protein- and lipid-linked glycans. Although it has been known that distinct glycan structures are associated with cancer, the whole spectrum of cancer-associated glycans has remained undiscovered. In the present study, we analyzed the protein-linked cancer glycome by matrix-assisted laser desorption/ionization time-of-flight mass spectrometric glycan profiling of cancer patient tissue samples. In lung cancer, we detected accumulation of a novel group of tumor-associated glycans. These protein-linked glycans carried abnormal nonreducing terminal beta-N-acetyl-D-glucosamine (GlcNAc) residues. A similar phenomenon was also detected in structural analyses of tumor-derived glycosphingolipids. This showed that glycan biosynthesis may dramatically change in cancer and that direct glycome analysis can detect the resulting marker glycans. Based on the structural knowledge, we further devised a covalent labeling technique for the detection of GlcNAc-expressing tumors with a specific transferase enzyme. In normal tissues, terminal GlcNAc antigens are capped by galactosylation. Similarly to common cancer-associated glycan antigens T, Tn, and sialyl-Tn, the newly discovered GlcNAc antigens result from incomplete glycosylation. In conclusion, the identified terminal GlcNAc glycans should be recognized as a novel class of tumor markers.
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| 31. |
- Svensson, Lola, 1948, et al.
(författare)
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Forssman expression on human erythrocytes: biochemical and genetic evidence of a new histo-blood group system
- 2013
-
Ingår i: Blood. - : American Society of Hematology. - 0006-4971 .- 1528-0020. ; 121:8, s. 1459-68
-
Tidskriftsartikel (refereegranskat)abstract
- Abstract In analogy with histo-blood group A antigen, Forssman (Fs) antigen terminates with α3-N-acetylgalactosamine and can be utilized by pathogens as a host receptor in many mammals. However, primates including humans lack Fs synthase activity and have naturally-occurring Fs antibodies in plasma. We investigated individuals with the enigmatic ABO subgroup Apae and found them to be homozygous for common O alleles. Their erythrocytes had no A antigens but instead expressed Fs glycolipids. The unexpected Fs antigen was confirmed in structural, serological and flowcytometric studies. The Fs synthase gene, GBGT1, in Apae individuals encoded an arginine to glutamine change at residue 296. Gln296 is present in lower mammals whereas Arg296 was found in six other primates, >250 blood donors and Apae family relatives without the Apae phenotype. Transfection experiments and molecular modelling showed that 296Gln reactivates the human Fs synthase. Uropathogenic E.coli containing prsG-adhesin-encoding plasmids agglutinated Apae but not group O cells, suggesting biological implications. Predictive tests for intravascular hemolysis with crossmatch-incompatible sera indicated complement-mediated destruction of Fspositive erythrocytes. Taken together, we provide the first conclusive description of Fs expression in normal human hematopoietic tissue and the basis of a new histo-blood group system in man, FORS.
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| 32. |
- Svensson, Lola, 1948, et al.
(författare)
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The structural basis of blood group A-related glycolipids in an A3 red cell phenotype and potential explanation to a serological phenomenon
- 2011
-
Ingår i: Glycobiology. - : Oxford University Press (OUP). - 1460-2423 .- 0959-6658. ; 21:2, s. 162-174
-
Tidskriftsartikel (refereegranskat)abstract
- Glycolipids from the red cells of a rare blood group A subgroup individual, expressing the blood group A(3) phenotype with the classical mixed-field agglutination phenomenon, A(2(539G>A))/O(1) genotype, and an unusual blood group A glycolipid profile, were submitted to a comprehensive biochemical and structural analysis. To determine the nature of blood group A glycolipids in this A(3) phenotype, structural determination was carried out with complementary techniques including proton nuclear magnetic resonance (1D and 2D), mass spectrometry (MS) (nano-electrospray ionization/quadrupole time-of-flight and tandem mass spectrometry) and thin layer chromatography with immunostaining detection. As expected, total blood group A structures were of low abundance, but contrary to expectations extended-A type 2 and A type 3 glycolipids were more dominant than A hexaglycosylceramides based on type 2 chain (A-6-2 glycolipids), which normally is the major A glycolipid. Several para-Forssman (GalNAcbeta3GbO(4)) structures, including extended forms, were identified but surmised not to contribute to the classic mixed-field agglutination of the A(3) phenotype. It is proposed that the low level of A antigen combined with an absence of extended branched glycolipids may be the factor determining the mixed-field agglutination phenomenon in this individual.
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| 33. |
- Teneberg, Susann, 1955, et al.
(författare)
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Carbohydrate recognition by enterohemorrhagic Escherichia coli: characterization of a novel glycosphingolipid from cat small intestine.
- 2004
-
Ingår i: Glycobiology. - : Oxford University Press (OUP). - 0959-6658 .- 1460-2423. ; 14:2, s. 187-96
-
Tidskriftsartikel (refereegranskat)abstract
- A key virulence trait of pathogenic bacteria is the ability to bind to receptors on mucosal cells. Here the potential glycosphingolipid receptors of enterohemorrhagic Escherichia coli were examined by binding of 35S-labeled bacteria to glycosphingolipids on thin-layer chromatograms. Thereby a selective interaction with two nonacid glycosphingolipids of cat small intestinal epithelium was found. The binding-active glycosphingolipids were isolated and, on the basis of mass spectrometry, proton NMR spectroscopy, and degradation studies, identified as Galalpha3Galbeta4Glcbeta1Cer (isoglobotriaosylceramide) and Galalpha3Galalpha3Galbeta4Glcbeta1Cer. The latter glycosphingolipid has not been described before. The interaction was not based on terminal Galalpha3 because the bacteria did not recognize the structurally related glycosphingolipids Galalpha3Galalpha4Galbeta4Glcbeta1Cer and Galalpha3Galbeta4GlcNAcbeta3Galbeta4Glcbeta1Cer (B5 glycosphingolipid). However, further binding assays using reference glycosphingolipids showed that the enterohemorrhagic E. coli also bound to lactosylceramide with phytosphingosine and/or hydroxy fatty acids, suggesting that the minimal structural element recognized is a correctly presented lactosyl unit. Further binding of neolactotetraosylceramide, lactotetraosylceramide, the Le(a)-5 glycosphingolipid, as well as a weak binding to gangliotriaosylceramide and gangliotetraosylceramide, was found in analogy with binding patterns that previously have been described for other bacteria classified as lactosylceramide-binding.
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| 34. |
- Thomsson, Kristina A, 1969, et al.
(författare)
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Detailed O-glycomics of the Muc2 mucin from colon of wild-type, core 1- and core 3-transferase-deficient mice highlights differences compared with human MUC2.
- 2012
-
Ingår i: Glycobiology. - : Oxford University Press (OUP). - 1460-2423 .- 0959-6658. ; 22:8, s. 1128-39
-
Tidskriftsartikel (refereegranskat)abstract
- The heavily O-glycosylated mucin MUC2 constitutes the major protein in the mucosal layer that acts as a physical barrier protecting the epithelial layer in the colon. In this study, Muc2 was purified from mucosal scrapings from the colon of wild-type (WT) mice, core 3 transferase knockout (C3Gnt(-/-)) mice and intestinal epithelial cell-specific core 1 knockout (IEC C1Galt1(-/-)) mice. The Muc2 O-glycans were released by reductive β-elimination and analyzed with liquid chromatography-mass spectrometry in the negative-ion mode. Muc2 from the distal colon of WT and C3Gnt(-/-) knockout mice carried a mixture of core 1- or core 2-type glycans, whereas Muc2 from IEC C1Galt1(-/-) mice carried highly sialylated core 3- and core 4-type glycans. A large portion of NeuAc in all mouse models was positioned on disialylated N-acetyllactosamine units, an epitope not reported on human colonic MUC2. Mass spectra and proton NMR spectroscopy revealed an abundant NeuAc linked to internally positioned N-acetylglucosamine on colonic murine Muc2, which also differs markedly from human MUC2. Our results highlight that murine colonic Muc2 O-glycosylation is substantially different from human MUC2, which could be one explanation for the different commensal microbiota of these two species.
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| 35. |
- Ångström, Jonas, 1950, et al.
(författare)
-
Chemical characterization of penta-, hexa-, hepta-, octa-, and nonaglycosylceramides of rat small intestine having a globoside-like terminus.
- 1982
-
Ingår i: The Journal of biological chemistry. - 0021-9258. ; 257:2, s. 682-8
-
Tidskriftsartikel (refereegranskat)abstract
- A novel series of glycosphingolipids has been isolated from the nonepithelial part of rat small intestine. A mixed fraction containing 3 major components corresponding to glycolipids with 5, 6, and 7 sugars and 2 minor components with 8 and 9 sugars was characterized. The structure of the major components was deduced by mass spectrometry and proton NMR spectroscopy of nondegraded permethylated and permethylated-reduced (LiAlH4) derivatives and gas-liquid chromatography of degradation products of native, permethylated, and permethylated-reduced glycolipids. The structures of the penta-, hexa-, and hepta-glycosylceramides were found to be GalNAcp beta 1 leads to (3Galp alpha 1 leads to)2-44Galp beta 1 leads to 4Glcp beta 1 leads to 1Cer. By analogy reasoning, supported by mass spectrometry, the octa- and nonaglycosylceramides were concluded to have 1 and 2 additional internal leads to 3Galp alpha 1 leads to 3 structures, respectively. A pentaglycosylceramide fraction from another rat strain was also isolated. The NMR spectra were in agreement with 2 isomeric structures of which 1 had the internal alpha 1 leads to 4 linkage replaced by an alpha 1 leads to 3 linkage. The fatty acids of all components were nonhydroxy 16:0 to 24:0 acids with the 18:0 homologue as dominating species. The major base was sphingosine and possibly monohydroxy 18:1 base in the larger glycolipids. This is a novel series of structures with a terminal saccharide identical with isoglobotetraoxylceramide (cytolipin R). The glycosyltransferase for the terminal GalNAc beta 1 leads to 3 of cytolipin R may possibly be identical with the enzyme adding the terminal sugar of this novel series. This is supported by the presence in the same tissue of probable precursor glycolipids with 4 to 8 hexoses.
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| 36. |
- Ångström, Jonas, 1950, et al.
(författare)
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Default biosynthesis pathway for blood group-related glycolipids in human small intestine as defined by structural identification of linear and branched glycosylceramides in a group O Le(a-b-) nonsecretor.
- 2004
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Ingår i: Glycobiology. - : Oxford University Press (OUP). - 0959-6658 .- 1460-2423. ; 14:1, s. 1-12
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Tidskriftsartikel (refereegranskat)abstract
- Glycoconjugates of the GI tract are important for microbial interactions. The expression of histo-blood group glycosyltransferases governs both the expression of blood group determinants and in part the structure and size of the glycoconjugates. Using neutral glycolipids isolated from the small intestine of a rare blood group O Le(a-b-) ABH secretor-negative (nonsecretor) individual we were able to map the "default" pathway of the individual lacking ABO, Lewis, and secretor glycosyltransferases. Structures were deduced with combined analysis of mass spectrometry (MALDI-TOF and ESI-MS/MS), and 1H NMR (500 and 600 MHz). All structures present at a level >5% were structurally resolved and included two extended structures: Galbeta4(Fucalpha3)GlcNAcbeta3(Galbeta4[Fucalpha3]GlcNAcbeta6)Galbeta4GlcNAcbeta3Galbeta4Glcbeta1Cer and Galbeta3GlcNAcbeta3(Galbeta4[Fucalpha3]GlcNAcbeta6)Galbeta3GlcNAcbeta3Galbeta4Glcbeta1Cer. The first, a novel component, is based on a type 2 chain and bears the Lex glycotopes on both its branches. The second, a major component, is based on a type 1 chain, which bears a 3-linked type 1 precursor (Lec) glycotope and a 6-linked Lex glycotope on its branches. This latter structure is identical to that previously isolated from plasma and characterized by MS and GC-MS but not by NMR. Structural resolution of these structures was supported by reanalysis of the blood group H-active decaosylceramides previously isolated from rat small intestine. Other minor linear monofucosylated penta-, hepta-, and difucosylated octaosylceramides, some bearing blood group determinants, were also identified. The cumulative data were used to define a default biosynthesis pathway where it can be seen that carbohydrate chain extension, in the absence of blood group glycosyltransferases, is controlled and regulated by non-blood group fucosylation and branching with type 2 Galbeta4GlcNAc branches.
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| 37. |
- Ångström, Jonas, 1950, et al.
(författare)
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Separation and characterization of hematosides with different sialic acids and ceramides from rat small intestine. Different composition of epithelial cells versus non-epithelial tissue and of duodenum versus jejunum-ileum.
- 1981
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Ingår i: Journal of biochemistry. - 0021-924X. ; 90:4, s. 909-21
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Tidskriftsartikel (refereegranskat)abstract
- The hematosides (sialyl-lactosylceramides) of rat small intestine were separated as their acetylated derivatives. The isolated fractions were characterized by mass spectrometry and degradative methods, and the two major fractions also by NMR spectroscopy. From these results hematosides with different sialic acid and ceramide type could be assigned to thin-layer chromatographic bands. This allowed a structural interpretation of the chromatographic patterns observed for different parts of the small intestine. Thus, epithelial cells of ileum contained only hematoside with N-glycoloylneuraminic acid. Duodenum lacked this compound and instead the epithelial cells contained hematoside with N-acetylneuraminic acid. In non-epithelial tissue or both duodenum and jejunum-ileum the major hematoside had N-acetyl-neuraminic acid. The hematosides of epithelial cells had ceramide containing 18 : 0 trihydroxy base combined with 16, 20, 22, 24 : 0, and 24 : 1 hydroxy fatty acids (major part) or non-hydroxy fatty acids. In the non-epithelial hematosides the ceramide consisted of 18 : 1 dihydroxy base combined with 16, 18, 20, 22, 24 : 0, and 24 : 1 non-hydroxy fatty acids.
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| 38. |
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