ANTI-O-LINKED N-ACETYLGLUCOSAMINE CLONE

General description

Posttranslational modification of proteins by β-linked N-acetylglucosamine (β-GlcNAc) via the hydroxyl moieties on serine or threonine residues is termed O-linked β-GlcNAc or simply O-GlcNAc. O-GlcNAc is one of the most abundant posttranslational modifications within the nucleocytoplasmic compartments of all animals and plants. Unlike other types of protein glycosylations, O-GlcNAc occurs exclusively within the nuclear and cytoplasmic compartments and is generally not further modified to form more elongated structures. In addition, O-GlcNAcylation is a highly dynamic and reversible process. The O-GlcNAc transferase (OGT) attaches O-GlcNAc to proteins at specific serine or threonine residues, while O-GlcNAcase catalyzes the removal/hydrolysis of O-GlcNAc from proteins. In fact, a dynamic interplay between O-GlcNAcylation and serine/threonine phosphorylation plays an important role in regulating cellular signaling. Tau and RNA polymerase II (Pol II) are two well known proteins that undergo modification by O-GlcNAcylation. In Alzheimer’s diseased human brains, tau becomes extensively phosphorylated and less O-GlcNAcylated. Similarly, O-GlcNAc is removed and replaced with O-phosphate on the Poly II CTD when the elongation phase of transcription is initiated.

Specificity

Specifically recoginzes O-linked N-Acetylglucosamine (O-GlcNAc) moieties on O-GlcNAcylated proteins and peptides. Exhibits little reactivity toward free GIcNAc and no reactivity toward GalNac. Galactosyltransferase treatment of O-GlcNAcylated proteins results in galactosylation of O-GlcNAc via β1-4 linkage and a complete loss of binding by clone RL2 (Holt, G.D., et al. (1987). J. Cell Biol. 104(5):1157-1164).

Target structure is not species-specific.

Immunogen

Pore complex-lamina fraction purified from rat liver nuclear envelopes corresponding to Rat O-Linked N-Acetylglucosamine.

Application

Anti-O-Linked N-Acetylglucosamine Antibody, clone RL2 is an antibody against O-Linked N-Acetylglucosamine for use in Western Blotting, Immunocytochemistry, Affinity Binding Assay, Electron Microscopy, Immunoprecipitation.

Immunocytochemistry Analysis: 4.0 µg/mL from a representative lot detected O-Linked N-Acetylglucosamine in HeLa cells.
Immunocytochemistry Analysis: A representative lot immunostained nuclear envelopes, but not the nuclear interior, of digitonin-permeabilized HeLa cells. Clone RL2 stained the nuclear interior only among Triton X-100-permeabilized HeLa cells without intact nuclear envelopes (Adam, S.A., et al. (1990). J. Cell Biol. 111(3):807-816).
Affinity Binding Assay: A representative lot was radiolabeled with 125I and studied for its binding characteristics toward isolated rat liver nuclear envelopes (Snow, C.M., et al. (1987). J. Cell Biol. 104(5):1143-1156).
Electron Microscopy: A representative lot localized the O-GlcNAc immunoreactivity in isolated rat liver nuclear envelopes (Snow, C.M., et al. (1987). J. Cell Biol. 104(5):1143-1156).
Western Blotting Analysis: A representative lot detected O-GlcNAcylated proteins in rat liver nuclear envelopes preparations (Snow, C.M., et al. (1987). J. Cell Biol. 104(5):1143-1156; Holt, G.D., et al. (1987). J. Cell Biol. 104(5):1157-1164).
Immunoprecipitation Analysis: A representative lot immunoprecipitated O-GlcNAcylated proteins from solubilized rat liver nuclear envelopes preparations. Pretreatment of nuclear envelopes preparations with galactosyltrarnsferase prevented the immunoprecipitation of glycoproteins by clone RL2 (Snow, C.M., et al. (1987). J. Cell Biol. 104(5):1143-1156; Holt, G.D., et al. (1987). J. Cell Biol. 104(5):1157-1164).

Quality

Evaluated by Western Blotting in HeLa cell lysate.

Western Blotting Analysis: 1.0 µg/mL of this antibody detected O-Linked N-Acetylglucosamine in 10 µg of HeLa cell lysate.

Target description

Variable, depending on the size(s) of the O-GlcNAcylated protein(s).

Physical form

Format: Purified

Other Notes

Concentration: Please refer to lot specific datasheet.

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