Glycoconjugate Journal

, Volume 8, Issue 5, pp 434–441 | Cite as

CMP-N-acetylneuraminic acid hydroxylase activity determines the wheat germ agglutinin-binding phenotype in two mutants of the lymphoma cell line MDAY-D2

  • Lee Shaw
  • Shida Yousefi
  • James W. Dennis
  • Roland Schauer
Papers

Abstract

The dominant glycosylation mutants of MDAY-D2 mouse lymphoma cells, designated class 2 (D33W25 and D34W25) were selected for their resistance to the toxic effects of wheat germ agglutinin (WGA) and shown to express elevated levels of Neu5Gc. In accordance with this, the activity of CMP-Neu5Ac hydroxylase was found to be substantially higher in the mutant cells. The hydroxylase in the D33W25 mutant cells exhibited kinetic properties identical to those of the same enzyme from mouse liver. Growth rate experimentsin vivo andin vitro, where the mutant cells grew more slowly at low cell densities in serum-free medium and also formed slower growing tumours in syngeneic mice, indicate that CMP-Neu5Ac hydroxylase expression may be associated with altered growth of the mutant cells.

Keywords

sialic acid CMP-N-acetylneuraminic acid hydroxylase wheat germ agglutinin N-glycoloylneuraminic acid glycosylation mutants 

Abbreviations

WGA

wheat germ agglutinin

Neu5Ac

N-acetyl-β-d-neuraminic acid

Neu5Gc

N-glycology-β-d-neuraminic acid

CMP-Neu5Ac

cytidine-5′-monophospho-N-acetylneuraminic acid

CMP-Neu5Gc

cytidine-5′-monophospho-N-glycoloylneuraminic acid

FACS

fluorescence-activated cell sorting

buffer A

triethylamine hydrogen carbonate, pH 7.6 (concentration given at appropriate points in the text)

SFM

serum free medium

IMDM

Iscove's modified Dulbecco's medium

CMP-Neu5Ac hydroxylase

CMP-N-acetylneuraminate: NAD(P)H oxido-reductase (N-acetyl hydroxylating) (EC 1.14.99.18); CMP-sialate hydrolase (EC 3.1.4.40); sialic acid-pyruvate lyase (EC 4.1.3.3)

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Copyright information

© Chapman & Hall 1991

Authors and Affiliations

  • Lee Shaw
    • 1
  • Shida Yousefi
    • 2
  • James W. Dennis
    • 2
    • 3
  • Roland Schauer
    • 1
  1. 1.Biochemisches InstitutChristian-Albrechts UniversitätKiel 1Germany
  2. 2.Samuel Lunenfeld Research Institute600 University AvenueTorontoCanada
  3. 3.Department of Medical GeneticsUniversity of TorontoTorontoCanada

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