| 1. |
- Bourgeois, C, et al.
(författare)
-
Heparin-like structures on respiratory syncytial virus are involved in its infectivity in vitro.
- 1998
-
Ingår i: Journal of Virology. - 0022-538X .- 1098-5514. ; 72:9, s. 7221-7227
-
Tidskriftsartikel (refereegranskat)abstract
- Addition of heparin to the virus culture inhibited syncytial plaque formation due to respiratory syncytial virus (RSV). Moreover, pretreatment of the virus with heparinase or an inhibitor of heparin, protamine, greatly reduced virus infectivity. Two anti-heparan sulfate antibodies stained RSV-infected cells, but not noninfected cells, by immunofluorescence. One of the antibodies was capable of neutralizing RSV infection in vitro. These results prove that heparin-like structures identified on RSV play a major role in early stages of infection. The RSV G protein is the attachment protein. Both anti-heparan sulfate antibodies specifically bound to this protein. Enzymatic digestion of polysaccharides in the G protein reduced the binding, which indicates that heparin-like structures are on the G protein. Such oligosaccharides may therefore participate in the attachment of the virus.
|
|
| 2. |
|
|
| 3. |
- Gustafson, Stefan, et al.
(författare)
-
Accessible hyaluronan receptors identical to ICAM-1 in mouse mast-cell tumours
- 1995
-
Ingår i: Glycoconjugate Journal. - 0282-0080 .- 1573-4986. ; 12:3, s. 350-355
-
Tidskriftsartikel (refereegranskat)abstract
- Immunohistochemical studies of the hyaluronan (HA)-receptor (R), originally found on liver endothelial cells (LEC) and related to the intercellular adhesion molecule 1 (ICAM-1), showed that polyclonal antibodies against HARLEC (HA receptor on LEC) also stain structures in mouse mastocytomas, mainly vessels. To test if intravenously administered HA might target the tumour receptors in vivo, mice carrying an inoculated mastocytoma in one hind leg muscle were injected in the tail vein with 125I-tyrosine (T)-labelled HA and killed 75 min after injection when organs and tissues were checked for radioactivity. When doses exceeding the binding capacity of the liver were injected, a significant increase in radioactivity (up to five-fold) within the tumour tissue was found. The weight adjusted difference between control and tumour tissue was greater for smaller tumours, probably due to necrosis in the larger. HA-staining of tumours from animals receiving 125I-T-HA, showed HA in areas that also stained weakly for ICAM-1 using monoclonal antibodies. ICAM-1 staining was dramatically increased after hyaluronidase treatment of the sections, indicating that the HA is bound to these receptors and thereby blocks antibody recognition.
|
|
| 4. |
- Hodson, N, et al.
(författare)
-
Identification that KfiA, a protein essential for the biosynthesis of the Escherichia coli K5 capsular polysaccharide, is an alpha -UDP-GlcNAcglycosyltransferase : The formation of a membrane-associated K5 biosynthetic complex requires KfiA, KfiB and KfiC.
- 2000
-
Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 275:35, s. 27311-27315
-
Tidskriftsartikel (refereegranskat)abstract
- The Escherichia coli K5 capsular polysaccharide consists of the repeat structure -4)GlcA-beta(1,4)-GlcNAc-alpha(1-and requires the KfiA, KfiB, KfiC, and KfiD proteins for its synthesis, Previously, the KfiC protein was shown to be a beta-UDP-GlcA glycosyltransferase, and KfiD was shown to be a UDP-Glc dehydrogenase. Here, we demonstrate that KfiA is an alpha-UDP-GlcNAc glycosyltransferase and that biosynthesis of the K5 polysaccharide involves the concerted action of the KfiA and KfiC proteins. By site-directed mutagenesis, we determined that the acidic motif of DDD, which is conserved between the C family of glycosyltransferases, is essential for the enzymatic activity of KfiA III addition, by Western blot analysis, we determined that association of KfiA with the cytoplasmic membrane requires KfiC but not KfiB, whereas the interaction of KfiC with the cytoplasmic membrane was dependent on both KfiA and KfiB. Likewise, KfiB was only detectable in cytoplasmic membrane fractions when both KfiA and KfiC were present. These data suggest that the interaction between the KfiA, KfiB, and KfiC proteins is essential for the stable association of these proteins with the cytoplasmic membrane and the biosynthesis of the K5 polysaccharide.
|
|
| 5. |
|
|
| 6. |
|
|
| 7. |
- Kitagawa, H, et al.
(författare)
-
N-acetylgalactosamine (GalNAc) transfer to the common carbohydrate-protein linkage region of sulfated glycosaminoglycans : identification of UDP-GaINAc:chondro oligosaccharide aNacetylgalactosaminyltransferase in fetal bovine serum
- 1995
-
Ingår i: The Journal of Biological Chemsitry. - 0021-9258. ; 270:38, s. 22190-22195
-
Tidskriftsartikel (refereegranskat)abstract
- During the course of a study to elucidate the role ofmodification of the common polysaccharide-protein linkagestructure, GlcAb1–3Galb1–3Galb1–4Xylb1-O-Ser, inbiosynthetic sorting mechanisms of the different sulfatedglycosaminoglycan chains, a novel N-acetylgalactosamine(GalNAc) transferase was discovered in fetalbovine serum. The enzyme catalyzed the transfer of[3H]GalNAc from UDP-[3H]GalNAc to linkage tetrasaccharideand hexasaccharide serines synthesized chemicallyand to various regular oligosaccharides containingterminal D-glucuronic acid (GlcA), which were preparedfrom chondroitin and chondroitin sulfate using testicularhyaluronidase digestion. The labeled products obtainedwith the linkage tetra- and hexasaccharideserines and with the tetrasaccharide (GlcAb1–3GalNAc)2were resistant to digestion with chondroitinase AC-IIand b-N-acetylhexosaminidase but sensitive to a-Nacetylgalactosaminidasedigestion, indicating that theenzyme is an a-N-acetylgalactosaminyltransferase. Thisfinding is in contrast to that of Rohrmann et al. (Rohrmann,K., Niemann, R., and Buddecke, E. (1985) Eur. J.Biochem., 148, 463–469), who reported that a correspondingproduct was susceptible to digestion with b-Nacetylhexosaminidase.The presence of a sulfate groupat C4 of the penultimate GalNAc or Gal units markedlyinhibited the transfer of GalNAc to the terminal GlcA,while a sulfate group at C6 of the GalNAc had little effecton the transfer. Moreover, a slight but significant transferof [3H]GalNAc was observed to an oligosaccharidecontaining terminal 2-O-sulfated GlcA as acceptor,whereas no incorporation was detected into oligosaccharidescontaining terminal unsaturated or 3-O-sulfatedGlcA units. These results suggest that this novelserum enzyme is a UDP-GalNAc:chondro-oligosaccharidea1–3- or 1–4-N-acetylgalactosaminyltransferase.
|
|
| 8. |
- Lidholt, Kerstin, et al.
(författare)
-
Assessment of glycosaminoglycan-protein linkage tetrasaccharides as acceptors for GalNAc- and GlcNAc-transferases from mouse mastocytoma.
- 1997
-
Ingår i: Glycoconjugate Journal. - 0282-0080 .- 1573-4986. ; 14:6, s. 737-742
-
Tidskriftsartikel (refereegranskat)abstract
- Two glycosaminoglycan-protein linkage tetrasaccharide-serine compounds, GlcA beta 1-3Gal beta 1-3Gal beta 1-4Xyl beta 1-O-Ser and GlcA beta 1-3Gal(4-O-sulfate)beta 1-3Gal beta 1-4Xyl beta 1-O-Ser, were tested as hexosamine accepters, using UDP-[H-3]GlcNAc and UDP-[H-3]GalNAc as sugar donors, and solubilized mouse mastocytoma microsomes as enzyme source. The nonsulfated Ser-tetrasaccharide was found to function as an acceptor for a GalNAc residue, whereas the Ser-tetrasaccharide containing a sulfated galactose unit was inactive. Characterization of the radio-labelled product by digestion with alpha-N-acetylgalactosaminidase and beta-N-acetylhexosaminidase revealed that the [H-3]GalNAc unit was alpha-linked, as in the product previously synthesized using serum enzymes, and not beta-linked as found in the chondroitin sulfate polymer. Heparan sulfate/heparin biosynthesis could not be primed by either of the two linkage Ser-tetrasaccharides, since no transfer of [H-3]GlcNAc from UDP-[H-3]GlcNAc could be detected. By contrast, transfer of a [H-3]GlcNAc unit to a [GlcA beta 1-4GlcNAca1-4](2)-GlcA beta 1-4-aMan hexasaccharide acceptor used to assay the GlcNAc transferase involved in chain elongation, was readily detected. These results are in agreement with the recent proposal that two different N-acetylglucosaminyl transferases catalyse the biosynthesis of heparan sulfate. Although the mastocytoma system contains both the heparan sulfate/heparin and chondroitin sulfate biosynthetic enzymes the Ser-tetrasaccharides do not seem to fulfil the requirements to serve as accepters for the first HexNAc transfer reactions involved in the formation of these polysaccharides.
|
|
| 9. |
|
|
| 10. |
- Lidholt, Kerstin, et al.
(författare)
-
Biosynthesis of heparin : Modulation of polysaccharide chain length in a cell-free system
- 1988
-
Ingår i: Biochemical Journal. - 0264-6021 .- 1470-8728. ; 254:2, s. 571-578
-
Tidskriftsartikel (refereegranskat)abstract
- The formation of heparin-precursor polysaccharide (N-acetylheparosan) was studied with a mouse mastocytoma microsomal fraction. Incubation of this fraction with UDP-[3H]GlcA and UDP-GlcNAc yielded labelled macromolecules that could be depolymerized, apparently to single polysaccharide chains, by alkali treatment, and thus were assumed to be proteoglycans. Label from UDP-[3H]GlcA (approx. 3 microM) is transiently incorporated into microsomal polysaccharide even in the absence of added UDP-GlcNAc, probably owing to the presence of endogenous sugar nucleotide. When the concentration of exogenous UDP-GlcNAc was increased to 25 microM the rate of incorporation of 3H increased and proteoglycans carrying polysaccharide chains with an Mr of approx. 110,000 were produced. Increasing the UDP-GlcNAc concentration to 5 mM led to an approx. 4-fold decrease in the rate of 3H incorporation and a decrease in the Mr of the resulting polysaccharide chains to approx. 6000 (predominant component). When both UDP-GlcA and UDP-GlcNAc were present at high concentrations (5 mM) the rate of polymerization and the polysaccharide chain size were again increased. The results suggest that the inhibition of polymerization observed at grossly different concentrations of the two sugar nucleotides, UDP-GlcA and UDP-GlcNAc, may be due either to interference with the transport of one of these precursors across the Golgi membrane or to competitive inhibition of one of the glycosyltransferases. The maximal rate of chain elongation obtained, under the conditions employed, was about 40 disaccharide units/min. The final length of the polysaccharide chains was directly related to the rate of the polymerization reaction.
|
|
