Enzymatic characterization of beta-D-galactoside alpha 2,3-trans-sialidase from Trypanosoma cruzi.
The substrate specificity, physico-chemical, and kinetic properties of the trans-sialidase from Trypanosoma cruzi have been investigated. The enzyme demonstrates activity towards a wide range of saccharide, glycolipid, and glycoprotein acceptors which terminate with a beta-linked galactose residue, and synthesizes exclusively an alpha 2-3 sialosidic linkage. Oligosaccharides which terminate in Gal beta 1-4(Fuc alpha 1-3)GlcNAc, Gal beta 1-3(Fuc alpha 1-4)GlcNAc, or Gal alpha 1- are not acceptor-substrates. The enzyme utilizes alpha 2,3-linked sialic acid when the donor species is an oligosaccharide and can also transfer, at a low rate, sialic acid from synthetic alpha-sialosides such as p-nitrophenyl-alpha-N-acetylneuraminic acid, but NeuAc alpha 2-3Gal beta 1-4(Fuc alpha 1-3)Glc is not a donor-substrate. The trans-sialidase has an apparent pH optimum of 7.9 and a temperature optimum of 13 degrees C. The kinetic properties of the enzyme suggest that the trans-sialylation reaction may occur via a rapid equilibrium random or steady-state ordered mechanism. A method for immobilizing the enzyme is described together with examples of its use for the synthesis of oligosaccharide and glycoprotein precursors of sialyl-Lewis and sialyl-Lewis.
Scudder
P
P
Division of Protein Biochemistry, Monsanto Co., St. Louis, Missouri 63167.
Doom
J P
JP
Chuenkova
M
M
Manger
I D
ID
Pereira
M E
ME
D000595:Amino Acid Sequence; D000818:Animals; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins; D007202:Indicators and Reagents; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009439:Neuraminidase / Q000302:isolation & purification / Q000378:metabolism*; D009842:Oligopeptides / Q000138:chemical synthesis; D009844:Oligosaccharides / Q000378:metabolism; D013379:Substrate Specificity; D013816:Thermodynamics; D014349:Trypanosoma cruzi / Q000201:enzymology*
D006023:Glycoproteins; D007202:Indicators and Reagents; D009842:Oligopeptides; D009844:Oligosaccharides; C069715:trans-sialidase; D009439:Neuraminidase