Related Experiment Videos
The presence of N-glycosylated proteins in cell nuclei
A Ferraro1, P Grandi, M Eufemi
1Department of Biochemical Sciences A. Rossi Fanelli, University La Sapienza, Rome.
Abstract:
The protein-DNA crosslinking capability of cis-dichloro diammineplatinum has been exploited to check the intranuclear location of N-glycosylated proteins. When intact liver cells were treated with this reagent, a number of glycoproteins, recognized by Concanavalin A, have been shown to become crosslinked to DNA; many of them have been recognized as nuclear matrix components. The recognition by this lectin was abolished by treatment with N-glycosidase F, showing the presence of N-glycosidic bonds between the sugar moiety and the protein. Most of the glycoproteins appeared to have high mannose oligosaccharide chains, but sialic acid containing oligosaccharides were also identified.
Insights
This study used cis-diammineplatinum to identify nuclear glycoproteins in liver cells. These N-glycosylated proteins were found to crosslink with DNA, indicating their presence within the cell nucleus and association with the nuclear matrix.
Area of Science:
- Biochemistry
- Cell Biology
- Molecular Biology
Background:
- N-glycosylated proteins play crucial roles in cellular functions.
- Understanding the intranuclear localization of glycoproteins is essential for deciphering their functions.
- cis-diammineplatinum is a known protein-DNA crosslinking agent.
Purpose of the Study:
- To investigate the intranuclear location of N-glycosylated proteins using cis-diammineplatinum.
- To identify nuclear matrix components among these glycoproteins.
Main Methods:
- Treatment of intact liver cells with cis-diammineplatinum.
- Detection of glycoproteins using Concanavalin A.
- Enzymatic cleavage with N-glycosidase F to confirm N-glycosidic bonds.
- Analysis of oligosaccharide chain composition (high mannose and sialic acid-containing).
Main Results:
- cis-diammineplatinum treatment resulted in crosslinking of several glycoproteins to DNA.
- Many of these crosslinked glycoproteins were identified as nuclear matrix components.
- N-glycosidase F treatment abolished Concanavalin A recognition, confirming N-glycosidic linkages.
- The identified glycoproteins predominantly featured high mannose oligosaccharide chains, with some also containing sialic acid.
Conclusions:
- N-glycosylated proteins are present within the nucleus and associated with the nuclear matrix.
- cis-diammineplatinum is an effective tool for probing the nuclear localization of glycoproteins.
- The study provides insights into the composition and nuclear association of specific glycoprotein populations.