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)

Membrane-associated sialyltransferase complexes in Escherichia coli K-235 can be dissociated by lipid deletion and reassembled by the addition of undecaprenyl phosphate, a unique membrane-bound lipid coenzyme. Following disruption of the cells by pressure disintegration and centrifugal fractionation, the sialyltransferase activity is assocatied with both a "particulate" and "soluble" complex. Kinetic studies as well as sugar nucleotide, metal ion, pH, ammonium sulfate, and thiol reagent requirements showed these two complexes contained functionally identical enzymatic activities. Isopycnic sucrose density gradient centrifugation studies carried out on unfractionated total membranes established that these sialytransferase activities were associated with membrane hybrids composed of different relative amounts of inner and outer membranes. Enzyme localization studies employing DPNH oxidase, a marker for the inner membrane, and relative phospholipid to protein composition determinations in the two complexes, provided added support for this conclusion. Sialyl polymer synthesis was not dependent on the incorporation of other monosaccharides and had no demonstrable metal ion requirement. Kinetic studies showed that the Km for cytidine 5-monophospho-N-acetylneuraminic acid in intact soluble and particulate enzyme preparations was 8.1 times 10-5M and 9.2 times 105M, respectively. Similarly, both enzyme complexes had nearly identical Vmax values. Following reassembly of delipidated enzyme preparations, however, there was a 10-fold increase in the Km value for the particulate enzyme and a 3-fold increase for the soluble enzyme. This increase was accompanied by an increase of approximately the same magnitude in the Vmax values. Since the lipid coenzyme was limiting in intact enzyme preparations, the increase in Vmax reflected an increase in the concentration of the active lipid in reconstituted complexes. Sialyl polymer synthesis in intact membrane preparations was stimulated by the exogenous addition of lipid. Insertion of the carrier lipid was dependent on temperature. At 37 degrees, a 120% increase in sialytransferase activity was observed while only a 35% increase was observed at 30 percent. At 20 degrees, no stimulation occurred. Fluidity of the lipid phase is apparently required for proper function of this membraneassociated enzyme complex. Thus, at 20 degrees, a temperature below the membrane lipid transition temperature, the lipids are relatively immobile.
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PMID:Properties of membrane-associated sialyltransferase of Escherichia coli. 109 67

Membranes from chick embryo epiphyseal cartilage were fractionated by equilibrium sucrose density gradient centrifugation and assayed for galactosyl xylose transferase, chondroitin polymerization and sulfation as well as the marker enzymes glucose-6-phosphatase, NADH cytochrome c reductase, galactosyl ovalbumin transferase, and sialyltransferase. The order of distribution of chondroitin sulfate synthesis from dense to light membranes correlated with the established sequence of events for its synthesis. The linkage region enzyme, viz. galactosyl xylose transferase, distributed with NADH cytochrome c reductase in an earlier and heavier cis compartment. Chondroitin polymerization and sulfation had a dual distribution similar to the galactosyl ovalbumin transferase and sialyltransferase in separate later and lighter medial and trans compartments, or in an extended medial or trans compartment. The galactosyl xylose transferase had a distribution distinctly different from that of the galactosyl ovalbumin transferase indicating that these distinct enzymes showed no cross-reactivity with their respective acceptor substrates. The dual distribution of chondroitin sulfate synthesis was consistent with our previous demonstration of the two nascent proteochondroitin populations produced by microsomal preparations from the same source. The results indicated separate subcellular locations for synthesis of the two forms.
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PMID:Subfractionation of chick embryo epiphyseal cartilage Golgi. Localization of enzymes involved in the synthesis of the polysaccharide portion of proteochondroitin sulfate. 185 50

The biosynthesis of N-glycoloylneuraminic acid in fractionated porcine submandibular glands was investigated. The following substrates: [3H]N-acetylmannosamine, free [14C]N-acetylneuraminic acid, CMP-[14C]N-acetylneuraminic acid, [14C]N-acetylneuraminic acid linked alpha(2----3) to galactose residues, or alpha(2----6) to Gal-beta(1----4)-GlcNAc residues of porcine submandibular mucin and [14C]N-acetylneuraminic acid linked alpha(2----6) to GalNAc residues of ovine submandibular gland mucin were incubated, in the presence of cofactors, with the soluble protein, heavy membrane and microsomal fractions of porcine submandibular glands. Radio thin-layer chromatographic analysis revealed that only one substrate, CMP-[14C]N-acetylneuraminic acid, was hydroxylated. The product was identified as CMP-[14C]N-glycoloylneuraminic acid by (i) co-chromatography with non-radioactive CMP-N-glycoloylneuraminic acid standard, (ii) acid hydrolysis to free [14C]N-glycoloylneuraminic acid, (iii) alkaline hydrolysis to yield N-glycoloylneuraminic acid and 2-deoxy-2,3-didehydro-N-glycoloylneuraminic acid and (iv) transfer of [14C]N-glycoloylneuraminic acid to asialo-fetuin by sialyltransferase. 85% of CMP-N-acetylneuraminic acid hydroxylase activity was present in the soluble protein fraction, with small amounts of activity in the two particulate fractions. The CMP-N-acetylneuraminic acid hydroxylase in the soluble protein fraction had an absolute requirement for Fe2+ ions and a reducing cofactor. NADPH and NADH were by far the most effective cofactors, smaller amounts of hydroxylation could, however, be supported by ascorbic acid and 6,7-dimethyl-5,6,7,8-tetrahydrobiopterin.
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PMID:The biosynthesis of N-glycoloylneuraminic acid occurs by hydroxylation of the CMP-glycoside of N-acetylneuraminic acid. 320 54