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:2.4.99.7 (sialyltransferase)
1,534 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The in vitro activity of sialyltransferase IV (SAT-IV), which catalyzes the transfer of sialic acid to the terminal galactose of different gangliotetraosylceramides (GA1, GM1a and GD1b), was examined in membrane-enriched preparations from mouse embryos at embryonic day 12 (E-12). Gangliosides GD1a and GT1b were the only reaction products using GM1a and GD1b as substrates, respectively. The Km values for GM1a and GD1b were 53 microM and 42 microM, respectively. Competitive inhibition experiments showed that the same enzyme (SAT-IV) catalyzed sialic acid transfer to the terminal galactose residues of both GM1a and GD1b. Two labeled ganglioside products were obtained, however, using GA1 as a substrate. One product was identified as ganglioside GM1b and the enzymatic reaction for its formation was maximal at pH 6.0, similar to that for GD1a and GT1b formation. The second product, synthesized by a different sialyltransferase, was identified as GD1 alpha based on results from TLC immunostaining, neuraminidase digestion, and periodate oxidation-borohydride reduction. The pH dependence curve for GD1 alpha formation had a different shape than that for GM1b formation with a maximum at pH 6.3. GD1 alpha is apparently synthesized from GM1b by an endosialyltransferase that catalyzes the transfer of a second sialic acid to the internal N-acetylgalactosamine of GM1b. The formation of both GM1b and GD1 alpha was linear over protein concentration. The ratio of GD1 alpha/GM1b formation varied from 0.25 to 1.20 depending on the GA1 substrate concentration.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Ganglioside biosynthesis in mouse embryos: sialyltransferase IV and the asialo pathway. 807 55

A new class of gangliosides, GT1a alpha and GQ1b alpha, were initially identified as cholinergic neuron-specific antigens in bovine brain. These gangliosides have in common alpha 2-6 NeuAc linked to the GalNAc residue in the gangliotetraose core structure. In this study, we have determined the biosynthetic pathways of GT1a alpha and GQ1b alpha using rat liver Golgi fraction. The results showed that GT1a alpha and GQ1b alpha were synthesized from GD1a and GT1b, respectively, by the action of a GalNAc alpha 2-6 sialyltransferase. It was also demonstrated that these two gangliosides were found to exist as extremely minor components in rat liver.
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PMID:Biosynthetic pathway for a new series of gangliosides, GT1a alpha and GQ1b alpha. 808 83

The developmental expression of the alpha 2,6- and alpha 2,8-linked sialic acid (Sia) residues in trout egg polysialoglycoproteins (PSGPs) was studied by correlating the temporal expression of these sugar residues, and the prerequisite sialyltransferases responsible for their synthesis, during oogenesis. The following new findings are reported. 1) Disialylated glycoproteins were identified in ovaries 4-6 months prior to ovulation. Three months prior to ovulation, a second more highly sialylated glycoprotein appeared. Structural studies confirmed that the two glycoproteins were discrete molecular species, designated PSGP(low Sia) and PSGP(high Sia), which differed only in their Sia content. PSGP(low Sia) contained mostly disialyl (Sia alpha 2,8-Sia alpha 2,6-) side chains, whereas PSGP(high Sia) contained alpha 2,8-linked oligo/polySia side chains ranging in length from 2 to over 20 Sia residues. The average degree of polymerization ([DP]av) was 6. 2) Biosynthetic studies using CMP-[14C]Neu5Ac indicated that three sialyltransferase activities were responsible for synthesis of the polysialyl residues of PSGPs: (i) alpha-N-acetylgalactosaminide alpha 2,6-sialyltransferase (alpha 2,6-ST), which catalyzed formation of the Sia residues alpha 2,6-linked to the proximal GalNAc residues in asialo-PSGP; (ii) alpha 2,6-sialoside alpha 2,8-sialyltransferase (alpha 2,8-ST or "initiase"), which catalyzed transfer of the first alpha 2,8-Sia residue to the alpha 2,6-linked Sia residue; and (iii) an alpha 2,8-polysialyltransferase (alpha 2,8-polyST or "polymerase"), responsible for synthesis of the alpha 2,8-linked poly/oligo Sia chains in PSGP(high Sia). Expression of these enzyme activities increased in accordance with the developmental appearance of each PSGP. 3) Structural characterization of the [14C]Sia-labeled side chains of each PSGP at different stages of development confirmed that synthesis of the disialyl unit containing a single alpha 2,8-Sia residue occurred before alpha 2,8-polysialylation. 4) In ovaries, 96% of the sialyltransferase activities were found in the Golgi-derived immature cortical vesicles or as soluble enzymes released from the fragile vesicles. Less than 4% of the activities were localized in the membrane (Golgi) fraction. In mature eggs, the sialyltransferases were also detected as soluble enzymes, and within the cortical vesicles.
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PMID:Developmental expression of trout egg polysialoglycoproteins and the prerequisite alpha 2,6-, and alpha 2,8-sialyl and alpha 2,8-polysialyltransferase activities required for their synthesis during oogenesis. 814 14

cDNA clones encoding GalNAc alpha 2,6-sialyltransferase (EC 2.4.99.3) have been isolated from chick embryo cDNA libraries using sequence information obtained from the conserved amino acid sequence of the previously cloned enzymes. The cDNA sequence included an open reading frame coding for 566 amino acids, and the deduced amino acid sequence showed 12% identity with that of Gal beta 1,4GlcNAc alpha 2,6-sialyltransferase from chick embryo. The primary structure of this enzyme suggested a putative domain structure, like that in other glycosyltransferases, consisting of a short NH2-terminal cytoplasmic domain, a signal-membrane anchor domain, a proteolytically sensitive stem region, and a large COOH-terminal active domain. The identity of this enzyme was confirmed by the construction of a recombinant sialyltransferase in which the NH2-terminal part (232 amino acid residues) was replaced with the immunoglobulin signal sequence. The expression of this recombinant in COS-7 cells resulted in secretion of a catalytically active and soluble form of the enzyme into the medium. The expressed enzyme exhibited activity toward only asialomucin and (asialo)fetuin, no significant activity being detected toward the other glycoprotein and glycolipid substrates tested. 14C-Sialylated glycols obtained from asialomucin re-sialylated with this enzyme were identical to NeuAc alpha 2,6-GalNAc-ol and GlcNAc beta 1,3(NeuAc alpha 2,6) GalNAc-ol. Synthetic GalNAc-SerNAc also served as an acceptor for alpha 2,6-sialylation. These results clearly showed that the expressed enzyme is GalNAc alpha 2,6-sialyltransferase.
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PMID:Molecular cloning and expression of GalNAc alpha 2,6-sialyltransferase. 828 7

To elucidate control mechanisms of O-glycan biosynthesis in leukemia and to develop biosynthetic inhibitors we have characterized core 2 UDP-GlcNAc:Gal beta 1-3GalNAc-R(GlcNAc to GalNAc) beta 6-N-acetylglucosaminyltransferase (EC 2.4.1.102; core 2 beta 6-GlcNAc-T) and CMP-sialic acid: Gal beta 1-3GalNAc-R alpha 3-sialyltransferase (EC 2.4.99.4; alpha 3-SA-T), two enzymes that are significantly increased in patients with chronic myelogenous leukemia (CML) and acute myeloid leukemia (AML). We observed distinct tissue-specific kinetic differences for the core 2 beta 6-GlcNAc-T activity; core 2 beta 6-GlcNAc-T from mucin secreting tissue (named core 2 beta 6-GlcNAc-T M) is accompanied by activities that synthesize core 4 [GlcNAc beta 1-6(GlcNAc beta 1-3)GalNAc-R] and blood group I [GlcNAc beta 1-6(GlcNAc beta 1-3)Gal beta-R] branches; core 2 beta 6-GlcNAc-T in leukemic cells (named core 2 beta-GlcNAc-T L) is not accompanied by these two activities and has a more restricted specificity. Core 2 beta 6-GlcNAc-T M and L both have an absolute requirement for the 4- and 6-hydroxyls of N-acetylgalactosamine and the 6-hydroxyl of galactose of the Gal beta 1-3GalNAc alpha-benzyl substrate but the recognition of other substituents of the sugar rings varies, depending on the tissue. alpha 3-sialyltransferase from human placenta and from AML cells also showed distinct specificity differences, although the enzymes from both tissues have an absolute requirement for the 3-hydroxyl of the galactose residue of Gal beta 1-3GalNAc alpha-Bn. Gal beta 1-3(6-deoxy)GalNAc alpha-Bn and 3-deoxy-Gal beta 1-3GalNAc alpha-Bn competitively inhibited core 2 beta 6-GlcNAc-T and alpha 3-sialyltransferase activities, respectively.
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PMID:Processing O-glycan core 1, Gal beta 1-3GalNAc alpha-R. Specificities of core 2, UDP-GlcNAc: Gal beta 1-3 GalNAc-R(GlcNAc to GalNAc) beta 6-N-acetylglucosaminyltransferase and CMP-sialic acid: Gal beta 1-3GalNAc-R alpha 3-sialyltransferase. 829 5

Incubation of synthetic Man beta 1-4GlcNAc beta-OMe, GalNAc beta 1-4GlcNAc beta-OMe, Glc beta 1-4GlcNAc beta-OMe, and GlcNAc beta 1-4GlcNAc beta-OMe with CMP-Neu5Ac and rat liver Gal beta 1-4GlcNAc alpha(2-6)-sialyltransferase resulted in the formation of Neu5Ac alpha 2-6Man beta 1-4GlcNAc beta-OMe, Neu5Ac alpha 2-6GalNAc beta 1-4GlcNAc beta-OMe, Neu5Ac alpha 2-6Glc beta 1-4GlcNAc beta-OMe and Neu5Ac alpha 2-6GlcNAc beta 1-4GlcNAc beta-OMe, respectively. Under conditions which led to quantitative conversion of Gal beta 1-4GlcNAc beta-OEt into Neu5Ac alpha 2-6Gal beta 1-4GlcNAc beta-OEt, the aforementioned products were obtained in yields of 4%, 48%, 16% and 8%, respectively. HPLC on Partisil 10 SAX was used to isolate the various sialyltrisaccharides, and identification was carried out using 1- and 2-dimensional 500-MHz 1H-NMR spectroscopy.
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PMID:Action of rat liver Gal beta 1-4GlcNAc alpha(2-6)-sialyltransferase on Man beta 1-4GlcNAc beta-OMe, GalNAc beta 1-4GlcNAc beta-OMe, Glc beta 1-4GlcNAc beta-OMe and GlcNAc beta 1-4GlcNAc beta-OMe as synthetic substrates. 835 30

Cell surface carbohydrates have been shown to be altered during cellular differentiation. Alveolar type II (ATII) cells in culture gradually lose their differentiated phenotype. Therefore, the aim of this study was: (1) to characterize changes in terminal carbohydrates of cell surface glycoproteins of rat ATII cells cultured for 1 to 5 days on plastic, and (2) to assess the concomitant changes in sialidase and sialyltransferase activity of ATII cell homogenates. Cells were surface-labeled with potassium-[3H]-borohydride after oxidation by sodium periodate at millimolar concentrations, galactose oxidase or neuraminidase plus galactose oxidase, allowing for the specific labeling of terminal sialic acids, terminal galactose/N-acetylgalactosamine (Gal/GalNAc), or terminal an penultimate Gal/GalNAc residues, respectively. Glycoproteins were separated by SDS-PAGE. On day 1, cells were heavily coated with sialic acids, since no labeling could be introduced with galactose oxidase alone. From day 1 to day 5, we observed a selective and progressive desialylation of two glycoproteins (200 and 165 kD). At the same time, the ATII cells' sialidase activity (pH 4.2) exhibited an 8-fold increase (60.3 +/- 4.0 pmol/min/mg protein on day 1 versus 406.9 +/- 3.7 pmol/min/mg protein on day 5), whereas the sialyltransferase activity increased 2-fold (212 +/- 8 fmol/min/mg protein on day 1 versus 395 +/- 82 fmol/min/mg protein on day 5) and the supernatant sialidase activity was unchanged (2.8 +/- 0.7 pmol/min/ml on day 5). Thus, the phenotypic changes of ATII cells in primary culture are accompanied by a partial cell surface desialylation and an increase in intracellular sialidase activity.
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PMID:Cell surface carbohydrates of rat alveolar type II cells in primary culture. 842 6

Sialyltransferase activities, SAT-3 (CMP-NeuAc:nLcOse4Cer alpha 2-3sialyltransferase) and SAT-4 (CMP-NeuAc:GgOse4Cer alpha 2-3sialyltransferase), in Colo 205 cells catalyze the transfer of sialic acid to the terminal galactose of GlcNc-- and GalNAc-containing glycolipid substrates, respectively. Competition kinetic studies with nLcOse4Cer and GM1 as substrates in a sialyltransferase assay show that these two activities are catalyzed by two different catalytic entities. The two enzymes were co-solubilized with taurochlorate and resolved by DEAE--Cibacron Blue--Sepharose column chromatography into two elution peaks. The column eluent with SAT-3 activity failed to transfer sialic acid to asialo alpha(1)-acid glycoprotein, indicating that this enzyme is different from the sialyltransferase (ST3N) that synthesizes NeuAc alpha 2-3Gal linkage in asparagine-linked oligosaccharides of glycoprotein. However, SAT-3 activity can be immunoprecipitated with a polyclonal antibody produced against a protein expressed in Escherichia coli as GST-fusion protein from an ECB cDNA homolog of an alpha 2-3 sialyltransferase SAT-3 or STZ) the has been cloned from human melanoma cell and human placenta. Thus a concentration-dependent decrease in the residual SAT-3 activity relative to SAT-4 activity was observed in the supernatant after precipitation of the immune complex. Expression of SAT-3 (STZ) cDNA was also detected in Colo 205 cell by RT-PCR, followed by sequence analysis of the RT-PCR product. Characterization of the catalytic reaction products of SAT-3 and SAT-4 with thin-layer chromatography, sialidase treatment, and binding to specific antibodies indicates that both SAT-3 and SAT-4 catalyze the formation of alpha 2-3 linkage between sialic acid and terminal galactose of glycolipid substrates.
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PMID:Characterization of two glycolipid: alpha 2-3sialyltransferases, SAT-3 (CMP-NeuAc:nLcOse4Cer alpha 2-3sialyltransferase) and SAT-4 (CMP-NeuAc:GgOse4Cer alpha 2-3sialyltransferase), from human colon carcinoma (Colo 205) cell line. 861

A cDNA encoding a novel sialyltransferase has been isolated employing the polymerase chain reaction using degenerate primers to conserved regions of the sialylmotif that is present in all eukaryotic members of the sialyltransferase gene family examined to date. The cDNA sequence revealed an open reading frame coding for 305 amino acids, making it the shortest sialyltransferase cloned to date. This open reading frame predicts all the characteristic structural features of other sialyltransferases including a type II membrane protein topology and both sialylmotifs, one centrally located and the second in the carboxyl-terminal portion of the cDNA. When compared with all other sialyltransferase cDNAs, the predicted amino acid sequence displays the lowest homology in the sialyltransferase gene family. Northern analysis shows this sialyltransferase to be developmentally regulated in brain with expression persisting through adulthood in spleen, kidney, and lung. Stable transfection of the full-length cDNA in the human kidney carcinoma cell line 293 produced an active sialyltransferase with marked specificity for the sialoside, Neu5Ac-alpha2,3Gal-beta1,3GalNAc and glycoconjugates carrying the same sequence such as G(M1b) and fetuin. The disialylated tetrasaccharide formed by reacting the sialyltransferase with the aforementioned sialoside was analyzed by one- and two-dimensional 1H and 13C NMR spectroscopy and was shown to be the Neu5Ac-alpha2,3Gal-beta1,3(Neu5Ac-alpha2,6)GalNAc sialoside. This indicates that the enzyme is a GalNAc alpha-2,6-sialyltransferase. Since two other ST6GalNAc sialyltransferase cDNAs have been isolated, this sialyltransferase has been designated ST6GalNAc III. Of these three, ST6GalNAc III displays the most restricted acceptor specificity and is the only sialyltransferase cloned to date capable of forming the developmentally regulated ganglioside G(D1alpha) from G(M1b).
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PMID:Molecular cloning of a developmentally regulated N-acetylgalactosamine alpha2,6-sialyltransferase specific for sialylated glycoconjugates. 863 73

cDNA and genomic clones encoding mouse Galbeta1, 3GalNAc-specific GalNAc alpha2,6-sialyltransferase (ST6GalNAc II) were isolated, and the structure organization of the gene was determined. The predicted amino acid sequence is 57.4% identical to the chick ST6GalNAc II sequence but 33.8% identical to the chick ST6GalNA I (GalNAc alpha2, 6-sialyltransferase) sequence. The ST6GalNAc II gene is constitutively expressed in various mouse tissues but highly expressed in lactating mammary gland and adult testis. The gene contains nine exons spanning about 25 kilobases of genomic DNA and encodes a messenger RNA of 1995 nucleotides. Primer extension and S1 nuclease protection analysis of submaxillary gland mRNA showed that the transcription of the ST6GalNAc II gene starts from 68 nucleotides upstream from the translation start site. Characterization of 5'-flanking genomic regions indicated that the Galbeta1,3GalNAc-specific GalNAc alpha2,6-sialyltransferase promoter is embedded in a G+C-rich domain and contains no TATA or CAAT box but has putative binding sites for transcription factors Sp1 and AP-2. Transient transfection experiments involving luciferase reporter genes demonstrated promoter activity in NIH3T3 cells.
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PMID:Molecular cloning and genomic analysis of mouse Galbeta1, 3GalNAc-specific GalNAc alpha2,6-sialyltransferase. 866 27


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