Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.4.99.6 (sialyltransferase)
1,546 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The substrate requirements, linkage specificity, and kinetic mechanism of a pure sialyltransferase from porcine submaxillary glands have been examined. The enzyme transfers sialic acid from the donor nucleotide, CMP-NeuAc, into the sequence NeuAcalpha2 leads to 3Galbeta1 leads to 3GalNAc, which is found in both glycoproteins and gangliosides. It forms only the alpha2 leads to 3 linkage with the disaccharide Gal/beta1 leads to 3GalNAc or antifreeze glycoprotein, which, along with asialoglycoproteins containing the sequence Gal/beta1 leads to 3GalNAcalpha1 leads to O-Thr/Ser, are the best acceptor substrates. Low molecular weight galactosides linked beta1 leads to 3 to glycose residues other than N-acetylgalactosamine are poor acceptors with relatively high Km values, while those in beta1 leads to 4 or beta1 leads to 6 linkages have both high Km and low Vmax. With glycoprotein and ganglioside acceptors this substrate specificity appears to be even more strict, with the sequence Gal/beta1 leads to 3GalNAc serving as the exclusive acceptor. Thus the present enzyme is not responsible either for the sequence, NeuAcalpha2 leads to 3Galbeta1 leads to 4GlcNAc, found in the asparagine-linked chains of certain glycoproteins, or for the synthesis of hematoside, NeuAcalpha2 leads to 3Galbeta1 leads to 4Glcbeta1 leads to 1Cer. Initial rate kinetic studies, with and without inhibitors, suggest that the transferase has an equilibrium random order mechanism.
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PMID:Enzymatic characterization of beta D-galactoside alpha2 leads to 3 sialyltransferase from porcine submaxillary gland. 43 98

By means of affinity chromatography on CDP-hexanolamine-agarose, a CMP-N-acetylneuraminate: alpha-N-acetylgalactosaminide alpha 2 leads to 6 sialyltransferase (EC 2.4.99.1) has been purified 117,000-fold to homogeneity from Triton X-100 extracts of porcine submaxillary glands. The enzyme consists of several electrophoretic forms that can be partially resolved by chromatography on Sephadex G-200, the largest of which has a molecular weight of approximately 160,000 as estimated by sodium dodecyl sulfate-gel electrophoresis. Periodate oxidation studies show that the linkage formed by this enzyme with ovine submaxillary asialo-mucin as the acceptor substrate is NeuAc alpha 2 leads to 6GalNAc alpha 1 leads to O-Thr/Ser. On the basis of initial rate studies and the patterns of inhibition observed with alternate acceptor substrates, the transferase is proposed to have either a random equilibrium kinetic mechanism or an ordered steady state mechanism with the acceptor substrate binding first. Among a wide variety of oligosaccharides, glycoproteins, and simple glycosides (including p-nitrophenyl-alpha-N-acetylgalactosaminide), the only acceptor substrates for this enzyme are those glycoproteins containing the structure, R leads to 3GalNAc alpha 1 leads to O-Thr/Ser, where R may be H or a beta-galactoside.
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PMID:Purification to homogeneity and enzymatic characterization of an alpha-N-acetylgalactosaminide alpha 2 leads to 6 sialyltransferase from porcine submaxillary glands. 44 88

Sialyltransferase activity and cell-cell adhesion rates of aging WI-38 cells were studied to determine the possible basis for a previously described decrease in membrane bound sialic acid and loss of proliferation of senescent cells. Ectosialyltransferase was demonstrated on the surface of both young and old WI-38 cells. The sialyltransferase assays consist of an enzyme source which is either the surface of intact cells (ectoenzyme) or a Triton X-100 cell homogenate, the nucleotide sialic acid donor (cytidine monophosphate-N-acetylneuraminic acid), and an asialo-acceptor which may be endogenous to the enzyme preparation or may be added exogenously. When sialyltransferase activity is measured in the absence of exogenous acceptors, there is a greater amount of sialic acid transferred by odl cells. However, when exogenous acceptors are provided, the amount of transfer is stimulated to a greater extent in young cells equalizing the amount of sialic acid incorporated into young and old cells. This suggests that there are fewer asialoglycoproteins and that acceptor concentration is a limiting factor in assays of young cell sialyltransferase. The end result of this may be the previously described decreased amount of membrane-bound sialic acid of old cells. A change in the adhesiveness of old cells is described which may be related to the altered cell surface.
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PMID:Sialyltransferase activities of aging diploid fibroblasts. 45 90

Serum levels of sialyltransferase and sialic acid were measured in patients with malignant melanomas (n = 49), healthy control persons (n = 20), and patients with non-malignant skin disorders (n = 30). Both parameters were found to be higher in malignant melanoma patients than in healthy control persons, but they were not significantly higher in melanoma patients than in patients with benign skin disorders, unless widespread dissemination of metastases had occurred. The highest values were measured in patients with liver and lung metastases. No general correlation was found between sialyltransferase activities and sialic acid concentrations. Sialic acid concentrations seem to be a better index for tumor spreading than sialyltransferase activities. In early stages of the disease, shedding from tumor cells is not the major source of elevated serum levels of sialyltransferase and sialic acid, respectively.
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PMID:Sialyltransferase levels and sialic acid concentrations in sera of patients with malignant melanomas. 47 55

An inhibitory effect due to broken cells is observed when sialyltransferase (CMP-N-acetylneuraminate:D-galactosyl-glycoprotein N-acetylneuraminyltransferase, EC 2.4.99.1) is measured with mixture of intact and homogenized lymphocytes. This intracellular inhibitory factor ib purified and characterized as CMP-N-acetylneuraminic acid (CMP-NeuNAc) by its behavior in various chromatographic and electrophoretic systems and by its susceptibility to CMP-NeuNAc hydrolase. This endogenous CMP-NeuNAc leads to an isotopic dilution of the exogenous labelled CMP-NeuNAc explaining the apparently lower activity of homogenate when compared to whole cells. Consequently, the radioactivity bound to acceptors may not be related to a known number of sialyl residues transferred, calling into question the validity of comparing the incorporation of [14C]NeuNAc by homogenate and whole cells in order to assign sialyltransferase activity to ectoenzyme. A new approach is developed to detect ectoglycosyltransferases with whole cells, taking into account that both intracellular enzymes and endogenous precursor may be introduced by the small percentage of broken cells.
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PMID:Detection of ectosiallyltransferase activity using whole cells. Correction of misleading results due to the release of intracellular CMP-N-acetylneuraminic acid. 48 88

The sialyltransferase and galactosyltransferase activities of the Golgi-rich fraction from rat liver were enhanced by the binding of wheat germ agglutinin (WGA). The sialytransferase was more sensitive than the galactosyltransferase to the WGA. Maximal stimulation of the galactosyltransferase activity resulted from the binding of 60--80 micrograms WGA to the Golgi membrane, while only 40 micrograms of WGA produced a maximal enhancement in the sialyltransferase activity. Within 5 min of WGA binding, the Golgi sialytransferase activity was doubled. After the initial binding of WGA to the Golgi fraction, the galactosyltransferase activity was decreased by 30%. However, in 15 min the activity was doubled by the binding of WGA. The activities of both enzymes were further enhanced by incubation for up to 90 min. The stimulation of both sialyltransferase and galactosyltransferase activities by WGA was reversed by N-acetyl-D-glucosamine (GlcNAc), the specific inhibitor of agglutination by WGA. Complete reversal of the enhanced activity was observed after 20--30 min in the presence of 1 micromol GlcNAc. The association constant for the binding of WGA to the Golgi membranes was calculated to be 4.16 X 10(-6) M from a Steck-Wallach plot. The 'n' value or mean binding sites was calculated as 5.26 X 10(-5) M/mg of Golgi membrane protein.
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PMID:The effect of wheat germ agglutinin on sialyl and galactosyltransferases of rat liver Golgi membranes. 48 41

A simple assay for the determination of sialyltransferase activity is described. The method involves the isolation of the radioactive reaction product on cellulose paper discs washed with trichloroacetic acid, thereby greatly facilitating the handling of large numbers of assays. This procedure is both more accurate and more sensitive than that involving precipitation with protein denaturants and separation by centrifugation, and is applicable to both soluble and particulate enzyme preparations. The application of the method to the determination of sialytransferase in human plasma is detailed. Enzyme activity is elevated two-fold in both cancer patients and patients with non-malignant disease. This suggests that the diagnostic use of determinations of sialyltransferase activity may be limited by its non-specificity.
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PMID:The application of a simple, sensitive disc assay for sialyltransferase activity to particulate and soluble enzyme preparations and its use in measuring enzymatic activity in human plasma samples. 48 83

Six purified glycosyltransferase (a beta-galactoside alpha 2 leads to 6 sialyltransferase, a beta-galactoside alpha 2 leads to 3 sialyltransferase, an alpha-N-acetylgalactosaminide alpha 2 leads to 6 sialyltransferase, a beta-galactoside alpha 1 leads to 2 fucosyltransferase, a beta-N-acetylglucosaminide alpha 1 leads to 3 fucosyltransferase, and a (fucosyl alpha 1 leads to 2) galactoside alpha 1 leads to 3 N-acetyl-galactosaminyltransferase) have been used to study the biosynthetic pathways for formation of the nonreducing terminal oligosaccharide sequences in mammalian glycoproteins. The two glycoproteins used as model acceptor substrates in this study were human asialotransferrin, which contains the nonreducing terminal oligosaccharide sequence Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man, and antifreeze glycoprotein, which contains oligosaccarides with the structure, Gal beta 1 leads to 3GalNAc alph 1 leads O-Thr. Sequential action of the six glycosyltransferases on these model substrates led to the formation of previously described oligosaccharide structures. The studies reported here indicate that the substrate specificities of the individual enzymes dictate the structures that can be synthesized and the pathways by which they may be formed. The actions of a number of the transferasesare mutually exclusive, thereby prohibiting the formation of theoretically possible oligosaccharide structures. Oligosaccharides with the terminal sequence NeuAc alpha 2 leads to 3(Fuc alpha 1 leads to 2)Gal beta 1 leads to 3GalNAc and NeuAc alpha 2 leads to 6Gal beta 1 leads to 4(Fuc alpha 1 leads to 3)GlcNAc cannot be formed because the prior incorporation of sialic acid by the sialyltransferases yields products that are not acceptor substrates for the fucosyltransferases, and vice versa. Synthesis of other products requires that the enzymes act sequentially in a specific order. The structures NeuAc alpha 2 leads to 6(Fuc alpha 1 leads to 2)Gal beta 1 leads to 4GlcNAc, Fuc alpha 1 leads to 2Gal beta 1 leads to 4(Fuc alpha 1 leads to 3)GlcNAc, GalNAc alpha 1 leads to 3(Fuc alpha 1 leads to 2)Gal beta 1 leads to 4GlcNAc, and GalNAc alpha 1 leads to 3(Fuc alpha 1 leads to 2)Gal beta 1 leads to 3GalNAc can only be synthesized if the fucosyl alpha 1 leads to 2 galactose linkage is formed first. Synthesis of the pentasaccharide sequences GalNAc alpha 1 leads to 3(Fuc alpha 1 leads to 2)Gal beta 1 leads to 3(NeuAc alpha 2 leads to 6)GalNAc and GalNAc alpha 1 leads to 3(Fuc alpha 1 leads to 2)Gal beta 1 leads to 4(Fuc alpha 1 leads to 3)GlcNAc requires that the N-acetylgalactosaminyltransferase act last on the former structure and that the alpha 1 leads to 3 fucosyltransferase act last on the latter. In those instances where a product can be formed by one of two possible pathways, the comparisons of reaction rates indicate that one pathway is usually preferred...
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PMID:Biosynthesis of mammalian glycoproteins. Glycosylation pathways in the synthesis of the nonreducing terminal sequences. 50 Jul 30

Extracellular and surface bound Ca is essential to excitation-contraction (E-C) coupling in mammalian cardiac muscle. In intact hearts from cardiomyopathic hamster with congestive heart failure, a concomitant decrease in the Ca content of a superficial pool was associated with the reduced contractility. Ca binding to cardiac sarcolemmal ghosts prepared from these hearts revealed two binding sites by Scatchard plot. In normal hamsters, the low affinity site had a capacity of 114 nmol Ca.mg-1 protein, a KD of 1.5 mmol . litre-1 and was sensitive to neuraminidase treatment but not to 100 mmol . litre-1 Na, K, or Li, Ca binding in vitro approached a 1:1 relationship with the sialic acid content of the ghosts, 159 nmol . mg-1 protein. The activity of the enzyme responsible for glycosidically linking sialic acid to interstitial and sarcolemmal glycoproteins, sialyltransferase, was reduced from 1.80 to 0.41 pmol . mg-1 protein in the myopathic hearts. We suggest the functional defect in the hamster cardiomyopathy is a reduction in sialyltransferase activity leading to the deficiency in surface sialic acid residues. As a consequence, contractility is reduced, but Ca influx is increased. Reflex sympathetic activity increases Ca influx resulting in "Ca overload" and eventual cellular necrosis.
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PMID:Excitation-contraction coupling in normal and myopathic hamster hearts III: functional deficiencies in interstitial glycoproteins. 51 59

The effect of butyrate on glycolipid metabolism and morphological differentiation in the cell culture system of rat ascites hepatomas, AH 7974 of island-forming type and AH 7974F of free type, was studied. Both cell lines adhered to the substratum in the presence of 1 mM butyrate. In the case of AH 7974, the addition of butyrate induced a distinct morphological change but the other cell line showed no such conspicuous change. Butyrate-treated AH 7974 cells showed a 2 to 3-fold elevation of CMP-N-acetylneuraminic acid: lactosylceramide sialyltransferase activity to form N-acetylneuraminylgalactosylglucosylceramide (GM3). On the other hand, no enzyme activity could be detected in AH 7974F cells. Four glycosyltransferase activities involved in glycolipid synthesis, including sialyltransferase in AH 7974F cells, were reduced by butyrate. From these observations we concluded that sialyltransferase to form GM3, or TM3 itself, is prerequisite for the morphological alteration induced by butyrate.
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PMID:Effect of butyrate on glycolipid metabolism of two cell types of rat ascites hepatomas with different ganglioside biosynthesis. 52 38


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