Abstract
The specificity of recombinant (2-->3)-alpha-sialyltransferase (ST3Gal-III), expressed in baculovirus-infected insect cells, has been determined with various oligosaccharide acceptors and sugar-nucleotide donors using a fluorescence based assay. Recombinant ST3Gal-III tagged with a polyhistidine tail was immobilized on Ni(2+)-NTA-Agarose as an active enzyme for use in the synthesis of three sialylated oligosaccharides: (i) the divalent molecule [alpha-Neu5Ac-(2-->3)-D-Galp-(1-->4)-beta-D-GlcpNAc-O-CH(2)](2)-C-(CH(2)OBn)(2) (12); (ii) the dansylated derivative, alpha-Neu5Ac-(2-->3)-D-Galp-(1-->3)-beta-D-GlcpNAc-O-(CH(2))(6)-NH-dansyl and; (iii) the tetrasacharide alpha-Neu5Ac-(2-->3)-beta-D-Galp-(1-->4)-beta-D-GlcpNAc-(1-->2)-alpha-D-Manp-O-CH(3). Compound 12 was itself prepared from the divalent N-acetyllactosamine molecule built on pentaerythritol by a chemo-enzymatic route.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Baculoviridae / genetics
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Carbohydrate Sequence
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Cell Line
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Cloning, Molecular
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Enzymes, Immobilized / chemistry
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Enzymes, Immobilized / metabolism
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Gene Expression
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Genetic Vectors / genetics
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Lewis Blood Group Antigens
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Lewis X Antigen
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Molecular Sequence Data
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Molecular Structure
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Nickel / chemistry*
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Oligosaccharides / chemical synthesis*
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Oligosaccharides / metabolism
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Rats
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Recombinant Proteins / chemistry
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Recombinant Proteins / metabolism*
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Sepharose / chemistry*
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Sialyltransferases / chemistry
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Sialyltransferases / genetics
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Sialyltransferases / metabolism*
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Spodoptera
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Substrate Specificity
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beta-Galactoside alpha-2,3-Sialyltransferase
Substances
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Enzymes, Immobilized
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Lewis Blood Group Antigens
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Lewis X Antigen
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Oligosaccharides
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Recombinant Proteins
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lewis(a)-lewis(x)
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Nickel
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Sepharose
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Sialyltransferases
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beta-Galactoside alpha-2,3-Sialyltransferase