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Molecular cloning of a developmentally regulated n-acetylgalactosamine alpha 2,6-sialyltransferase specific for sialylated glycoconjugates

Academic Article
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Overview

authors

  • Sjoberg, E. R.
  • Kitagawa, H.
  • Glushka, J.
  • vanHalbeek, H.
  • Paulson, James

publication date

  • March 1996

journal

  • Journal of Biological Chemistry  Journal

abstract

  • 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).

subject areas

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Carbohydrate Sequence
  • Chickens
  • Cloning, Molecular
  • Consensus Sequence
  • Conserved Sequence
  • DNA Primers
  • DNA, Complementary
  • Gene Expression Regulation, Enzymologic
  • Glycoconjugates
  • Humans
  • Magnetic Resonance Spectroscopy
  • Membrane Proteins
  • Mice
  • Molecular Sequence Data
  • Multigene Family
  • Oligosaccharides
  • Open Reading Frames
  • Polymerase Chain Reaction
  • Rats
  • Recombinant Proteins
  • Sequence Homology, Amino Acid
  • Sialic Acids
  • Sialyltransferases
  • Substrate Specificity
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Identity

International Standard Serial Number (ISSN)

  • 0021-9258

PubMed ID

  • 8631773
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Additional Document Info

start page

  • 7450

end page

  • 7459

volume

  • 271

issue

  • 13

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