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Purification and characterization of α(2-6)-sialyltransferase from human liver

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Abstract

A Galβ1-4GlcNAc α(2-6)-sialyltransferase from human liver was purified 34 340-fold with 18% yield by dye chromatography on Cibacron Blue F3GA and cation exchange FPLC. The enzyme preparation was free of other sialyltransferases. It did not contain CMP-NeuAc hydrolase, protease, or sialidase activity, and was stable at −20°C for at least eight months. The donor substrate specificity was examined with CMP-NeuAc analogues modified at C-5 or C-9 of theN-acetylneuraminic acid moiety. Affinity of the human enzyme for parent CMP-NeuAc and each CMP-NeuAc analogue was substantially higher than the corresponding Galβ1-4GlcNAc α(2-6)-sialyltransferase from rat liver.

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Abbreviations

FPLC:

fast protein liquid chromatography

NeuAc:

5-N-acetyl-d-neuraminic acid

9-amino-NeuAc:

5-acetamido-9-amino-3,5,9-trideoxy-d-glycero-2-nonulosonic acid

9-acetamido-NeuAc:

5,9-diacetamido-3,5,9-trideoxy-d-glycero-β-d-2-nonulosonic acid

9-benzamido-NeuAc:

5-acetamido-9-benzamido-3,5,9-trideoxy-d-glycero-β-d-galacto-2-nonulosonic acid

9-fluoresceinyl-NeuAc:

9-fluoresceinylthioureido-NeuAc

5-formyl-Neu:

5-formyl-β-d-neuraminic acid

5-aminoacetyl-Neu:

5-aminoacetyl-β-d-neuraminic acid

CMP-NeuAc:

cytidine-5′-monophospho-N-acetylneuraminic acid

GM1 :

Galβ1-3GalNAcβ1-4(NeuAcα2-3)Galβ1-4Glc-ceramide

ST:

sialyltransferase

DTE:

1,4-dithioerythritol

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Authors and Affiliations

  1. Institut für Biochemie II der Universität Heidelberg, Im Neuenheimer Feld 328, 6900, Heidelberg 1, FRG

    Udo Sticher, Hans J Gross & Reinhard Brossmer

Authors
  1. Udo Sticher
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  2. Hans J Gross
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  3. Reinhard Brossmer
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Additional information

Enzyme: Galβ1-4GlcNAc α(2-6)-sialyltransferase, EC 2.4.99.1.

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Sticher, U., Gross, H.J. & Brossmer, R. Purification and characterization of α(2-6)-sialyltransferase from human liver. Glycoconjugate J 8, 45–54 (1991). https://doi.org/10.1007/BF00731642

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  • DOI: https://doi.org/10.1007/BF00731642

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