Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.4.99.7 (sialyltransferase)
 1,534 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Although it has previously been shown that bovine parvovirus (BPV) attaches to the sialated glycoprotein glycophorin A on erythrocytes, the nature of virus-binding moieties on mammalian nucleated cells is less clear. Buffalo lung fibroblasts (Bu), primary bovine embryonic kidney cells, Madin-Darby bovine kidney cells and bovine embryonic trachea (EBTr) cells were assessed for molecules capable of binding BPV. Competition studies were carried out on both erythrocyte and nucleated cell targets using a variety of sialated compounds and sialic acid-negative compounds. Glycophorin A was found to inhibit BPV binding, while mucin exhibited low-level inhibition. These two sialated compounds also blocked attachment of BPV-modified microsphere carriers to the Bu cell membrane. Influenza A virus was used as a sialic acid competitor and interfered with BPV attachment to erythrocytes and replication in Bu cells. Significantly, the enzyme sialidase removed BPV-binding sites from Bu and EBTr cells. The binding sites could be reconstituted on sialidase-treated cells by the enzymes alpha-2,3-O-sialyltransferase and alpha-2,3-N-sialyltransferase. These results indicated that BPV can attach to sialic acid on cell membranes and that the sialylglycoproteins available for virus attachment appear to contain both N- and O-linked carbohydrate moieties, but that not all members of the sialic acid family can bind BPV. Moreover, there may be other moieties that can bind BPV, which may act as either primary or secondary receptors.
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PMID:Attachment of bovine parvovirus to sialic acids on bovine cell membranes. 1526 59

Surface expressed negatively charged sialoglycans contribute to the regulation of adhesive cellular interactions and are thus involved in the growth and differentiation of hematopoietic progenitor cells. In particular, the cell surface sialylation state may govern the liberation of CD34+ hematopoietic precursors from bone marrow stroma cells and extracellular matrix. In order to assess the overall surface sialylation of live human CD34+ hematopoietic precursor cells, we applied a previously described flow cytometric enzyme assay. Cells with and without sialidase pretreatment were incubated in the presence of fluorescent CMP-sialic acid and exogenous ST6GalI. Thus sialylation of surface-expressed lactosamine residues was analysed. We demonstrated that surface lactosamines of CD34+ precursors derived from bone marrow and peripheral blood are over 95% sialylated, predominantly in alpha2-6 linkage. These results are in accordance with flow cytometric analysis of surface lectin staining. Sialic acid specific lectins MAA and SNA were strongly bound whereas SBA, VVA, and PNA became reactive only after sialidase pretreatment. CD34+ leukemia cell lines TF1 and KG1a also showed a high degree of surface sialylation, whereas cell line KG1 expressed to the largest extent free lactosamines. In these cell lines, alpha2-6 and alpha2-3 sialylated residues were present in equal amounts. In a variation of the flow cytometric enzyme assay, live cells were incubated without exogenous STGal I to measure the activity of endogenous ecto-sialyltransferase. Ecto sialyltransferase activity was observed in all CD34+ cells which was able to resialylate major surface glycoproteins such as HLA Class I, CD45, CD43, and CD34. The ecto-sialyltransferase may serve to maintain or increase surface sialylation rapidly without de novo synthesis.
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PMID:Cell surface sialylation and ecto-sialyltransferase activity of human CD34 progenitors from peripheral blood and bone marrow. 1575 Jul 86

A multifunctional sialyltransferase has been cloned from Pasteurella multocida strain P-1059 and expressed in E. coli as a truncated C-terminal His6-tagged recombinant protein (tPm0188Ph). Biochemical studies indicate that the obtained protein is (1) an alpha2,3-sialyltransferase (main function), (2) an alpha2,6-sialyltransferase, (3) an alpha2,3-sialidase, and (4) an alpha2,3-trans-sialidase. The recombinant tPm0188Ph is a powerful tool in the synthesis of structurally diverse sialoside libraries due to its relaxed substrate specificity, high solubility, high expression level, and multifunctionality.
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PMID:A multifunctional Pasteurella multocida sialyltransferase: a powerful tool for the synthesis of sialoside libraries. 1635 Oct 87

The negative effects of ammonium on recombinant protein productivity and glycosylation have been well documented, but the interaction of ammonium on glycosylation genes has not been completely elucidated. In this study, the effects of elevated ammonium on 12 glycosylation related genes in Chinese hamster ovary cells were evaluated by quantitative real time reverse transcriptase polymerase chain reaction. Numerous cytosol and endoplasmic reticulum (ER) localized genes associated with early glycosylation steps were insensitive to the ammonium condition. The initial expression of uridine diphosphate (UDP)-galactose transporter was higher for the ammonium-treated culture, while the initial expressions of cytosine monophosphate (CMP)-sialic acid transporter, beta(1,4)-galactosyltransferase, and UDP-glucose pyrophosphorylase were higher for the control culture. alpha(2,3)-sialyltransferase was observed to have lower expression level under the elevated ammonium condition compared to the control culture. This study indicates that galactosylation and sialylation inhibition is mainly due to decreased gene expression of galactosyltransferase, sialyltransferase, and CMP-sialic acid transporter and not due to sialidase. These unbalanced initial glycosylation and branching steps can explain the higher molecular heterogeneity under ammonium stress. Moreover, this study indicates that elevated ammonium has limited effects on the glycosylation genes associated with the ER and cytosol compared to the genes associated with the Golgi.
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PMID:Effects of elevated ammonium on glycosylation gene expression in CHO cells. 1638 Feb 82

Here, we describe an activity assay for sialyltransferases based on surface plasmon resonance (SPR). Different natural and synthetic oligosaccharides serving as acceptor substrates for the sialyltransferase ST3Gal-III (EC 2.4.99.6) were immobilized or synthesized on SPR chips. The chip was then exposed to different concentrations of a reaction mixture of ST3Gal-III and CMP-Neu5Ac either by injection or by external application of the reaction mixture to the chip surface. The binding of two lectins, one that specifically recognizes the unmodified acceptor, the other the sialylated oligosaccharide, was utilized to determine the extent of enzymatic turnover. In order to obtain enzymatic activities, the SPR data were correlated to data obtained from a classical radio assay. After regeneration, that is, cleavage of the sialic acid residues by using a sialidase, the chip is available for new experiments. The technique allows the rapid determination of sialyltransferase activity with only nanomolar quantities of acceptor substrates and should be of particular value in cases in which a large variety of samples, including cell lysates, have to be screened for their enzymatic activities.
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PMID:Assaying sialyltransferase activity with surface plasmon resonance. 1684 45

Ganglioside GM1 and its seven potential catabolic products: asialo-GM1, GM2, asialo-GM2, GM3, Lac-Cer, Glc-Cer and Cer, were labeled with tetramethylrhodamine (TMR) to permit ultra-sensitive analysis using laser-induced fluorescence (LIF) detection. The preparation involved acylation of the homogenous C(18)lyso-forms of GM1, Lac-Cer, Glc-Cer and Cer with the N-hydroxysuccinimide ester of a beta-alanine-tethered 6-TMR derivative, followed by conversion of these labeled products using galactosidase, sialidase, and sialyltransferase enzymes. The TMR-glycolipid analogs produced are detectable on TLC down to the 1 ng level by the naked eye. All eight compounds could be separated within 4 min in capillary electrophoresis where they could be detected at the zeptomole (ca. 1000 molecule) level using LIF.
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PMID:Synthesis of reference standards to enable single cell metabolomic studies of tetramethylrhodamine-labeled ganglioside GM1. 1706 78

CstII from bacterium Campylobacter jejuni strain OH4384 has been previously characterized as a bifunctional sialyltransferase having both alpha2,3-sialyltransferase (GM3 oligosaccharide synthase) and alpha2,8-sialyltransferase (GD3 oligosaccharide synthase) activities which catalyze the transfer of N-acetylneuraminic acid (Neu5Ac) from cytidine 5'-monophosphate (CMP)-Neu5Ac to C-3' of the galactose in lactose and to C-8 of the Neu5Ac in 3'-sialyllactose, respectively (Gilbert M, Karwaski MF, Bernatchez S, Young NM, Taboada E, Michniewicz J, Cunningham AM, Wakarchuk WW. 2002. The genetic bases for the variation in the lipo-oligosaccharide of the mucosal pathogen, Campylobacter jejuni. Biosynthesis of sialylated ganglioside mimics in the core oligosaccharide. J Biol Chem. 277:327-337). We report here the characterization of a truncated CstII mutant (CstIIDelta32(I53S)) cloned from a synthetic gene whose codons are optimized for an Escherichia coli expression system. In addition to the alpha2,3- and alpha2,8-sialyltransferase activities reported before for the synthesis of GM3- and GD3-type oligosaccharides, respectively, the CstIIDelta32(I53S) has alpha2,8-sialyltransferase (GT3 oligosaccharide synthase) activity for the synthesis of GT3 oligosaccharide. It also has alpha2,8-sialidase (GD3 oligosaccharide sialidase) activity that catalyzes the specific cleavage of the alpha2,8-sialyl linkage of GD3-type oligosaccharides and alpha2,8-trans-sialidase (GD3 oligosaccharide trans-sialidase) activity that catalyzes the transfer of a sialic acid from a GD3 oligosaccharide to a different GM3 oligosaccharide (3'-sialyllactoside). The donor substrate specificity study of the CstIIDelta32(I53S) GD3 oligosaccharide synthase activity indicates that the enzyme is flexible in using different CMP-activated sialic acids and their analogs for the synthesis of GD3 oligosaccharides containing natural and nonnatural modifications at the terminal sialic acid.
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PMID:Multifunctionality of Campylobacter jejuni sialyltransferase CstII: characterization of GD3/GT3 oligosaccharide synthase, GD3 oligosaccharide sialidase, and trans-sialidase activities. 1850 8

Endogenous polymorphonuclear leukocyte (PMN)-associated sialidase activity enhances PMN adhesion to and migration across the endothelium through the removal of sialylated cell-surface residues. We tested the hypothesis that PMNs also express sialyltransferase (ST) activity that restores sialyl residues to the PMN surface. We developed a highly sensitive fluorometric assay to demonstrate that intact human PMNs can mediate and accept sialyl residue transfer. This ST activity is inhibited by a ST inhibitor, CMP, which also inhibits the transendothelial migration of PMNs in response to IL-8 in vitro and in vivo. We conclude that intact PMNs express sialidase and ST activities that permit rapid modulation of their surface sialylation and their ability to adhere to and migrate across the endothelium.
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PMID:Expression of sialyltransferase activity on intact human neutrophils. 1866 29

Oseltamivir (Tamiflu) and zanamivir (Relenza), two extensively used clinically effective anti-influenza drugs, are viral sialidase (also known as neuraminidase) inhibitors that prevent the release of progeny virions and thereby limit the spread of infection. Recently mortalities and neuropsychiatric events have been reported with the use of oseltamivir, especially in pediatric cases in Japan, suggesting that these drugs might also inhibit endogenous enzymes involved in sialic acid metabolism, including sialidase, sialyltransferase, and CMP-synthase, in addition to their inhibitory effects on the viral sialidase. The possible inhibition could account for some of the rare side effects of oseltamivir. However, there has been little direct evidence in regard to the sensitivities of animal sialidases to these drugs. Here, we examined whether these inhibitors might indeed affect the activities of human sialidases, which differ in primary structures and enzyme properties but possess tertiary structures similar to those of the viral enzymes. Using recombinant enzymes corresponding to the four human sialidases identified so far, we found that oseltamivir carboxylate scarcely affected the activities of any of the sialidases, even at 1 mM, while zanamivir significantly inhibited the human sialidases NEU3 and NEU2 in the micromolar range (K(i), 3.7 +/- 0.48 and 12.9 +/- 0.07 microM, respectively), providing a contrast to the low nanomolar concentrations at which these drugs block the activity of the viral sialidases.
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PMID:Limited inhibitory effects of oseltamivir and zanamivir on human sialidases. 1869 48

Trans-sialidases catalyze the transfer of a sialic acid from one sialoside to an acceptor to form a new sialoside. alpha2,3-Trans-sialidase activity was initially discovered in the parasitic protozoan Trypanosoma cruzi, and more recently was found in a multifunctional Pasteurella multocida sialyltransferase PmST1. alpha2,8-Trans-sialidase activity was also described for a multifunctional Campylobacter jejuni sialyltransferase CstII. We report here the discovery of the alpha2,6-trans-sialidase activity of a previously reported recombinant truncated bacterial alpha2,6-sialyltransferase from Photobacterium damsela (Delta15Pd2,6ST). This is the first time that the alpha2,6-trans-sialidase activity has ever been identified. Kinetic studies indicate that Delta15Pd2,6ST-catalyzed trans-sialidase reaction follows a ping-pong bi-bi reaction mechanism. Cytidine 5'-monophosphate, the product of sialyltransferase reactions, is not required by the trans-sialidase activity of the enzyme but enhances the trans-sialidase activity modestly as a non-essential activator. Using chemically synthesized Neu5AcalphapNP and LacbetaMU, alpha2,6-linked sialoside Neu5Acalpha2,6LacbetaMU has been obtained in one-step in high yield using the trans-sialidase activity of Delta15Pd2,6ST. In addition to the alpha2,6-trans-sialidase activity, Delta15Pd2,6ST also has alpha2,6-sialidase activity. The multifunctionality is thus a common feature of many bacterial sialyltransferases.
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PMID:Trans-sialidase activity of Photobacterium damsela alpha2,6-sialyltransferase and its application in the synthesis of sialosides. 1988 Apr 25


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