Characterization of the peptide-N4-(N-acetylglucosaminyl) asparagine amidase (PNGase Se) from Silene alba cells.
The peptide-N4-(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 cultured Silene 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 cultured Silene alba cells.
Lhernould
S
S
Laboratoire de Biologie Cellulaire Végétale et Valorisation des Espéces Ligneuses, Université de Limoges, France.
Karamanos
Y
Y
Lerouge
P
P
Morvan
H
H
D000581:Amidohydrolases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D000645:Ammonium Sulfate / Q000737:chemistry; D000818:Animals; D001216:Asparagine / Q000737:chemistry / Q000378:metabolism; D002240:Carbohydrate Sequence; D002457:Cell Extracts / Q000737:chemistry; D002478:Cells, Cultured; D011232:Chemical Precipitation; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D002855:Chromatography, Thin Layer; D004591:Electrophoresis, Polyacrylamide Gel; D005453:Fluorescence; D006031:Glycosylation; D006801:Humans; D006863:Hydrogen-Ion Concentration; D019684:Magnoliopsida / Q000201:enzymology*; D008969:Molecular Sequence Data; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D010944:Plants / Q000737:chemistry; D013379:Substrate Specificity
D002457:Cell Extracts; D001216:Asparagine; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D000645:Ammonium Sulfate