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Journal ArticleDOI

A detailed study of the periodate oxidation of sialic acids in glycoproteins.

01 Jan 1989-Glycoconjugate Journal (Wallin and Dalholm)-Vol. 6, Iss: 1, pp 35-44
TL;DR: Periodate oxidation of terminalN-acetyl- andN-glycoloylneuraminic acid residues in the mucins from edible bird nest substance and pig submandibular gland can be carried out under conditions which exclusively give rise to the formation of the C-7 analogues of these sialic acids.
Abstract: Periodate oxidation of terminalN-acetyl- andN-glycoloylneuraminic acid residues in the mucins from edible bird nest substance and pig submandibular gland, respectively, can be carried out under conditions which exclusively give rise to the formation of the C-7 analogues of these sialic acids. In contrast, the C-8 compounds can be obtained in a maximum yield of about 40%. Under identical conditions,N-glycoloylneuraminic acid is oxidized about 1.5 times faster than theN-acetylated derivative. After release of the sialic acids by acid hydrolysis, the characterization of the oxidation products was carried out by TLC, by GLC and GLC-MS of the corresponding pertrimethylsilyl derivatives, and by 500-MHz1H-NMR spectroscopy. In addition, molar response factors for GLC analysis and extinction coefficients in the orcinol/Fe3+/HCl assay were determined.

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Citations
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PatentDOI
TL;DR: The coupling strategy which selectively modified a portion of the sialic acid residues of types II polysaccharide before coupling the polysacchide to TT preserved the epitope essential to protective immunity and enhanced the immunogenicity of the poly Saccharide.
Abstract: This invention relates to antigenic conjugate molecules comprising the capsular polysaccharide of Group B streptococcus type II which are covalently linked to protein. This invention also relates to vaccines and methods of immunizing mammals, including humans against infection by Group B streptococcus type II (GBS II). Multivalent vaccines comprising the conjugate molecules of this invention and antigens to other pathogenic bacteria are also claimed.

83 citations

Journal ArticleDOI
TL;DR: It is found that Plasmodium gallinaceum ookinetes readily adhered to midguts of Aedes aegypti, mimicking the natural recognition of the epithelium by the parasite, and this is the first receptor-ligand interaction demonstrated for the ookinete stage of a malaria parasite.
Abstract: During the course of its development in the mosquito and transmission to a new vertebrate host, the malaria parasite must interact with the mosquito midgut and invade the gut epithelium. To investigate how the parasite recognizes the midgut before invasion, we have developed an in vitro adhesion assay based on combining fluorescently labelled ookinetes with isolated midgut epithelia from blood-fed mosquitoes. Using this assay, we found that Plasmodium gallinaceum ookinetes readily adhered to midguts of Aedes aegypti, mimicking the natural recognition of the epithelium by the parasite. This interaction is specific: the ookinetes preferentially adhered to the lumen (microvillar) side of the gut epithelium and did not bind to other mosquito tissues. Conversely, the binding was not due to a non-specific adhesive property of the midguts, because a variety of other cell types, including untransformed P. gallinaceum zygotes or macrogametes, did not show similar binding to the midguts. High concentrations of glycosylated (fetuin, orosomucoid, ovalbumin) or non-glycosylated (bovine serum albumin) proteins, added as non-specific competitors, failed to compete with the ookinetes in binding assays. We also found that the adhesion of ookinetes to the midgut surface is necessary for sporogonic development of the parasite in the mosquito. Antibodies and other reagents that blocked adhesion in vitro also reduced oocyst formation when these reagents were combined with mature ookinetes and fed to mosquitoes. Chemical modification of the midguts with sodium periodate at pH 5.5 destroyed adhesion, indicating that the ookinete binds to a carbohydrate ligand on the surface of the midgut. The ligand is sensitive to periodate concentrations of less than 1 mmol l-1, suggesting that it may contain sialic-acid-like sugars. Furthermore, free N-acetylneuraminic acid competed with the ookinetes in binding aasays, while other monosaccharides had no effect. However, in agreement with the current belief that adult insects do not contain sialic acids, we were unable to detect any sialic acids in mosquito midguts using the most sensitive HPLC-based fluorometric assay currently available. We postulate that a specific carbohydrate group is used by the ookinete to recognize the midgut epithelium and to attach to its surface. This is the first receptor-ligand interaction demonstrated for the ookinete stage of a malaria parasite. Further characterization of the midgut ligand and its parasite counterpart may lead to novel strategies of blocking oocyst development in the mosquito.

79 citations

Journal ArticleDOI
TL;DR: Covalent cross-linking of the oxidized ligand to L-selectin can be demonstrated, suggesting Schiff base formation between lysine residues of the selectin and the newly formed aldehydes, and could be used to design drugs for disrupting leukocyte-endothelial interactions leading to pathological inflammation.
Abstract: The selectins are adhesion receptors that play key roles in leukocyte trafficking. Each has an N-terminal C-type lectin domain that binds to specific carbohydrates in a calcium-dependent manner. L-selectin recognizes sulfated, sialylated ligands on lymph node high endothelial venules. This recognition is abolished by strong periodate oxidation under conditions that destroy oligosaccharides. In contrast, mild periodate oxidation, which selectively oxidizes the side chain of sialic acid residues without affecting the underlying oligosaccharide, markedly enhances this interaction. The enhancement is calcium dependent, indicating that lectin recognition is maintained. Reduction of the sialic acid aldehydes generated by mild periodate to alcohol groups abolishes this effect. Covalent cross-linking of the oxidized ligand to L-selectin can be demonstrated, suggesting Schiff base formation between lysine residues of the selectin and the newly formed aldehydes. Such selectively oxidized sialylated ligands could be used to probe the lectin domains of the selectins and to identify lysine residues near the binding site. Also, this approach could be used to design drugs for disrupting leukocyte-endothelial interactions leading to pathological inflammation.

68 citations

Journal ArticleDOI
TL;DR: The reaction between ABAO and aldehydes is kinetically similar to oxime formations performed under stoichiometric aniline catalysis and the relationship between apparent rate constant and pH suggests that a protonated benzamidoxime acts as an internal general acid in Schiff-base formation.
Abstract: We describe the rapid reaction of 2-amino benzamidoxime (ABAO) derivatives with aldehydes in water. The ABAO combines an aniline moiety for iminium-based activation of the aldehyde and a nucleophilic group (Nu:) ortho to the amine for intramolecular ring closure. The reaction between ABAO and aldehydes is kinetically similar to oxime formations performed under stoichiometric aniline catalysis. We characterized the reaction by both NMR and UV spectroscopy and determined that the rate-determining step of the process is formation of a Schiff base, which is followed by rapid intramolecular ring closure. The relationship between apparent rate constant and pH suggests that a protonated benzamidoxime acts as an internal general acid in Schiff-base formation. The reaction is accelerated by substituents in the aromatic ring that increase the basicity of the aromatic amine. The rate of up to 40 M–1 s–1 between an electron-rich aldehyde and 5-methoxy-ABAO (PMA), which was observed at pH 4.5, places this reaction amo...

61 citations

Journal ArticleDOI
TL;DR: In this article, the reproducibility of the two-step RT-PCR protocol was investigated and the median coefficient of variation (standard deviation divided by the arithmetic mean) for the paired relative quantities obtained in the two parallel series for the four genes and amount to 7.7%.

58 citations

References
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Journal ArticleDOI
TL;DR: This chapter discusses the different aspects of thiobarbituric acid assay of sialic acid, which is suitable for measuring the release of bound sialoic acid by sialidase and hydrolysis of sIALic acid-containing material must be carried out for the measurement of total sialsic acids.

6,264 citations

Book ChapterDOI
TL;DR: This chapter presents literature data on the high-resolution, 1H-NMR spectroscopy of carbohydrates derived from glycoconjugates and discusses the results for carbohydrates related to the glycoproteins of N-glycosylic type.
Abstract: Publisher Summary This chapter discusses the application of high-resolution, 1H-nuclear magnetic resonance (NMR) spectroscopy to the structural analysis of carbohydrates related to glycoproteins. Glycoproteins are biopolymers consisting of a polypeptide backbone bearing one or more covalently linked carbohydrate chains. The carbohydrate chains of glycoproteins may be classified according to the type of linkage to the polypeptide backbone. N-Glycosylic chains are attached to the amide group of asparagine (Asn), whereas the O-glycosylic chains are linked to the hydroxyl group of amino acid residues such as serine (Ser), threonine (Thr), and hydroxylysine (Hyl). The high-resolution, 1H-NMR spectroscopy technique, in conjunction with methylation analysis, is extremely suitable for the structural studies of N-, as well as on O-, glycosylic glycans. For the interpretation of the 1H-NMR spectrum of a carbohydrate chain in terms of primary structural assignments, the concept of “structural reporter groups” was developed. This means that the chemical shifts of protons resonating at clearly distinguishable positions in the spectrum, together with their coupling constants and the line widths of their signals, bear the information essential to permit the assigning of the primary structure. This chapter presents literature data on the high-resolution, 1H-NMR spectroscopy of carbohydrates derived from glycoconjugates. It also discusses the results for carbohydrates related to the glycoproteins of N-glycosylic type.

797 citations

Journal ArticleDOI
TL;DR: This simple method of specifically introducing a radioactive label into cell surface sialic acids is useful in the study of cell surfaceSialic acid-containing glycoproteins.

482 citations

Book ChapterDOI

230 citations

Book ChapterDOI
01 Jan 1982
TL;DR: The discovery and the widespread occurrence of the sialic acids in mammalian tissues has been correlated with a range of different biological functions (see chapter J), and the known spectrum of derivatives covered in this chapter continues to expand.
Abstract: The discovery and the widespread occurrence of the sialic acids in mammalian tissues (Blix 1936, Klenk 1941) has been correlated with a range of different biological functions (see chapter J) which continues to expand. Although the literature on sialic acid occurrence is large (Gottschalk 1960, Blix and Jeanloz 1969, Tuppy and Gottschalk 1972, Schauer 1973, Ng and Dain 1976), it has been essentially limited to the vertebrates and some examples from the Echinodermata and Bacteria. Only one survey of sialic acid distribution in different phyla has been published (Warren 1963). Current data on sialic acid structure is compiled in Table 1, introducing the known spectrum of derivatives covered in this chapter.

168 citations