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Characterization of the peptide-N 4-(N-acetylglucosaminyl) asparagine amidase (PNGase Se) fromSilene alba cells

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Abstract

The peptide-N 4-(N-acetylglucosaminyl) asparagine amidase (PNGase Se) earlier described [Lhernould S., Karamanos Y., Bourgerie S., Strecker G., Julien R., Morvan H. (1992)Glycoconjugate J 9:191–97] was partially purified from culturedSilene alba cells using affinity chromatography. The enzyme is active between pH 3.0 and 6.5, and is stable in the presence of moderate concentrations of several other protein unfolding chemicals, but is readily inactivated by SDS. Although the enzyme cleaves the carbohydrate from a variety of animal and plant glycopeptides, it does not hydrolyse the carbohydrate from most of the corresponding unfolded glycoproteins in otherwise comparable conditions. The substrate specificity of this plant PNGase supports the hypothesis that this enzyme could be at the origin of the production of ‘unconjugated N-glycans’ in a suspension medium of culturedSilene alba cells.

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Abbreviations

GlcNAc:

N-acetylglucosamine

PNGase:

peptide-N 4-(N-acetylglucosaminyl) asparagine amidase

BSA:

bovine serum albumin

SDS-PAGE:

sodium dodecyl sulfate-polyacrylamide gel electrophoresis

TLC:

thin layer chromatography

HPAEC-PAD:

High pH anion exchange chromatography-pulsed amperometric detection

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Authors and Affiliations

  1. Laboratoire de Biologie Cellulaire Végétale et Valorisation des Espéces Ligneuses, Université de Limoges, 123, Avenue Albert Thomas, 87060, Limoges Cédex, France

    Sabine Lhernould & Henri Morvan

  2. Institut de Biotechnologie, Université de Limoges, 123, Avenue Albert Thomas, 87060, Limoges Cédex, France

    Yannis Karamanos

  3. Laboratoire des transports Intracellulaires, URA 203, UFR des Sciences, Université de Rouen, 76821, Mont St Aignan Cédex, France

    Patrice Lerouge

Authors
  1. Sabine Lhernould
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  2. Yannis Karamanos
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  3. Patrice Lerouge
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  4. Henri Morvan
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Lhernould, S., Karamanos, Y., Lerouge, P. et al. Characterization of the peptide-N 4-(N-acetylglucosaminyl) asparagine amidase (PNGase Se) fromSilene alba cells. Glycoconjugate J 12, 94–98 (1995). https://doi.org/10.1007/BF00731874

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  • DOI: https://doi.org/10.1007/BF00731874

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