Abstract
Glycosylation of nuclear and cytoplasmic proteins by O-linked N-acetylglucosamine (O-GlcNAc) monosaccharides is an abundant, ubiquitous, and transient post-translational modification. To characterize enzymes involved in removal of these sugars, a neutral and cytoplasmic N-acetyl-β-D-glucosaminidase (O-GlcNA-case) with strong selectivity for O-GlcNAc-synthetic glycopeptides has been purified over 22,000-fold from rat spleen homogenate. The purified O-GlcNAcase has two major polypeptides of apparent Mr = 54,000 (α subunit) and Mr = 51,000 (β subunit). Enzyme activity sediments at Mr = 106,000 on sucrose gradients, indicating that the native O-GlcNAcase is an αβ heterodimer. The O-GlcNA-case also shows substantially stronger relative activity against O-GlcNAc-synthetic glycopeptides than other hexosaminidases. Unlike acidic lysosomal hexosaminidases, O-GlcNAcase is not inhibited by GalNAc or its analogs, has no other detectable glycosidase activities, and does not cross-react with antibodies against acidic hexosaminidases. Subcellular fractionation and latency studies demonstrate the cytoplasmic and nucleoplasmic localization of the enzyme and its ubiquitous presence in tissues. These studies suggest that O-GlcNAcase is involved in the regulated removal of O-GlcNAc from O-GlcNAc-bearing glycoproteins in the nucleoplasmic and cytoplasmic compartments of cells.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 19321-19330 |
| Number of pages | 10 |
| Journal | Journal of Biological Chemistry |
| Volume | 269 |
| Issue number | 30 |
| State | Published - Jul 29 1994 |
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ASJC Scopus subject areas
- Biochemistry
Cite this
Purification and characterization of an O-GlcNAc selective N-acetyl-β-D-glucosaminidase from rat spleen cytosol. / Dong, Dennis L Y; Hart, Gerald Warren.
In: Journal of Biological Chemistry, Vol. 269, No. 30, 29.07.1994, p. 19321-19330.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Purification and characterization of an O-GlcNAc selective N-acetyl-β-D-glucosaminidase from rat spleen cytosol
AU - Dong, Dennis L Y
AU - Hart, Gerald Warren
PY - 1994/7/29
Y1 - 1994/7/29
N2 - Glycosylation of nuclear and cytoplasmic proteins by O-linked N-acetylglucosamine (O-GlcNAc) monosaccharides is an abundant, ubiquitous, and transient post-translational modification. To characterize enzymes involved in removal of these sugars, a neutral and cytoplasmic N-acetyl-β-D-glucosaminidase (O-GlcNA-case) with strong selectivity for O-GlcNAc-synthetic glycopeptides has been purified over 22,000-fold from rat spleen homogenate. The purified O-GlcNAcase has two major polypeptides of apparent Mr = 54,000 (α subunit) and Mr = 51,000 (β subunit). Enzyme activity sediments at Mr = 106,000 on sucrose gradients, indicating that the native O-GlcNAcase is an αβ heterodimer. The O-GlcNA-case also shows substantially stronger relative activity against O-GlcNAc-synthetic glycopeptides than other hexosaminidases. Unlike acidic lysosomal hexosaminidases, O-GlcNAcase is not inhibited by GalNAc or its analogs, has no other detectable glycosidase activities, and does not cross-react with antibodies against acidic hexosaminidases. Subcellular fractionation and latency studies demonstrate the cytoplasmic and nucleoplasmic localization of the enzyme and its ubiquitous presence in tissues. These studies suggest that O-GlcNAcase is involved in the regulated removal of O-GlcNAc from O-GlcNAc-bearing glycoproteins in the nucleoplasmic and cytoplasmic compartments of cells.
AB - Glycosylation of nuclear and cytoplasmic proteins by O-linked N-acetylglucosamine (O-GlcNAc) monosaccharides is an abundant, ubiquitous, and transient post-translational modification. To characterize enzymes involved in removal of these sugars, a neutral and cytoplasmic N-acetyl-β-D-glucosaminidase (O-GlcNA-case) with strong selectivity for O-GlcNAc-synthetic glycopeptides has been purified over 22,000-fold from rat spleen homogenate. The purified O-GlcNAcase has two major polypeptides of apparent Mr = 54,000 (α subunit) and Mr = 51,000 (β subunit). Enzyme activity sediments at Mr = 106,000 on sucrose gradients, indicating that the native O-GlcNAcase is an αβ heterodimer. The O-GlcNA-case also shows substantially stronger relative activity against O-GlcNAc-synthetic glycopeptides than other hexosaminidases. Unlike acidic lysosomal hexosaminidases, O-GlcNAcase is not inhibited by GalNAc or its analogs, has no other detectable glycosidase activities, and does not cross-react with antibodies against acidic hexosaminidases. Subcellular fractionation and latency studies demonstrate the cytoplasmic and nucleoplasmic localization of the enzyme and its ubiquitous presence in tissues. These studies suggest that O-GlcNAcase is involved in the regulated removal of O-GlcNAc from O-GlcNAc-bearing glycoproteins in the nucleoplasmic and cytoplasmic compartments of cells.
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UR - http://www.scopus.com/inward/citedby.url?scp=0028085881&partnerID=8YFLogxK
M3 - Article
C2 - 8034696
AN - SCOPUS:0028085881
VL - 269
SP - 19321
EP - 19330
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 30
ER -