Related Experiment Video

Updated: Aug 8, 2026

Blue Native Polyacrylamide Gel Electrophoresis (BN-PAGE) for Analysis of Multiprotein Complexes from Cellular Lysates
12:03

Blue Native Polyacrylamide Gel Electrophoresis (BN-PAGE) for Analysis of Multiprotein Complexes from Cellular Lysates

Published on: February 25, 2011

Two-dimensional electrophoresis reveals a nuclear matrix-associated nucleolin complex of basic isoelectric point

J Gotzmann1, A Eger, M Meissner

  • 1Institute of Tumor Biology-Cancer Research, University of Vienna, Austria.

Electrophoresis
|April 4, 1998
PubMed

Insights

Researchers identified a novel form of nucleolin (a protein) associated with the nuclear matrix. This nuclear matrix-bound nucleolin interacts with ribonucleoproteins and a basic component, resisting standard dissociation methods.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Nuclear matrix and intermediate filaments serve crucial structural roles in eukaryotic cells.
  • Nucleolin is a well-characterized nucleolar protein involved in various cellular processes.
  • Understanding protein associations within the nuclear matrix is key to deciphering nuclear organization.

Purpose of the Study:

  • To characterize a novel protein recognized by a monoclonal antibody raised against nuclear matrix scaffolds.
  • To investigate the properties and localization of this antigen, particularly its relationship with nucleolin.
  • To elucidate the composition and interactions of nuclear matrix-bound proteins.

Main Methods:

  • Generation of a monoclonal antibody against nuclear matrix/intermediate filament scaffolds.
  • Immunoblotting and immunofluorescence microscopy to detect and localize the antigen.
  • Partial peptide sequencing for protein identification.
  • Two-dimensional gel electrophoresis to analyze protein properties and fractions.
  • Fractionation of cellular components and solubilization studies using RNAse and SDS.

Main Results:

  • A monoclonal antibody recognized a ~100 kDa protein in human cells and tissues, localized to nucleolar structures.
  • Peptide sequencing indicated the antigen is homologous to human nucleolin.
  • Two forms of the antigen were detected by 2D electrophoresis: one acidic (pI 5.5-6.1) and one basic (pI 8.5-9).
  • The acidic form was soluble, while the basic form was nuclear matrix-bound and RNAse-sensitive.
  • RNAse treatment released the basic component, which, upon SDS treatment and re-analysis, shifted to the acidic pI.

Conclusions:

  • The nuclear matrix harbors a nucleolin-like protein associated with ribonucleoproteins.
  • This nuclear matrix-bound nucleolin exists in a complex with a basic component, resistant to standard dissociation.
  • RNAse treatment suggests the involvement of RNA in maintaining the integrity or association of this nucleolin complex within the nuclear matrix.

More Related Videos

Visualization of miniSOG Tagged DNA Repair Proteins in Combination with Electron Spectroscopic Imaging (ESI)13:06

Visualization of miniSOG Tagged DNA Repair Proteins in Combination with Electron Spectroscopic Imaging (ESI)

Published on: September 24, 2015

Horizontal Gel Electrophoresis for Enhanced Detection of Protein-RNA Complexes06:36

Horizontal Gel Electrophoresis for Enhanced Detection of Protein-RNA Complexes

Published on: July 28, 2017

Related Concept Videos

The Nucleolus02:55

The Nucleolus

The nucleolus is the most prominent substructure of the nucleus. When it was first discovered, it was considered to be an isolated organelle that forms fibrils and granules. In 1931, the relationship between the nucleolus and chromosomes was first described by Heitz. He observed that the appearance and size of nucleolus varies depending on the stage of the cell cycle. He also noticed constricted regions on different chromosomes clustered together at definite cell cycle stages. These regions,...
Nuclear Protein Sorting01:34

Nuclear Protein Sorting

Nuclear protein sorting is the selective trafficking of histones, polymerases, gene regulatory proteins into the nucleus and exporting RNAs and ribosomes to the cytosol. It is a tightly controlled process that regulates gene expression within a cell.
Proteins targeted to the nucleus carry nuclear localization signals or NLS recognized by import receptors in the cytosol. Similarly, proteins with nuclear export signals are recognized by export receptors. Import and export receptors are...
Nuclear Localization Signals and Import01:46

Nuclear Localization Signals and Import

Proteins targeted to the nucleus carry short stretches of amino acid sequences called the nuclear localization signal or NLS. Classical nuclear localization signals are of two types: monopartite and bipartite NLS. Monopartite classical NLS (cNLS) consists of a single cluster of 4-8 amino acids. Bipartite cNLS consists of two clusters of  2-3 amino acids and a 9-12 residue long proline-rich linker bridging the two clusters. Signal clusters are rich in positively charged amino acids such as...
SDS-PAGE01:27

SDS-PAGE

Gel electrophoresis is a method that separates biological macromolecules like nucleic acids or proteins by forcing them to pass through a gel matrix under an electric field.
A variation of gel electrophoresis, termed  polyacrylamide gel electrophoresis (PAGE), is commonly used for separating proteins according to their molecular size by passing them through a polyacrylamide gel. Because of the varying charges associated with amino acid side chains, PAGE can be used to separate intact proteins...
Western Blotting01:15

Western Blotting

Western blotting is an analytical technique for protein identification. It has various applications in immunology and medicine, including detecting diseases like bovine spongiform encephalopathy, mad cow disease, and human and feline immunodeficiency virus from biological samples.
The technique begins with separating proteins from the sample using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), followed by protein transfer, immunoblotting, and finally, protein detection.
Two-dimensional Gel Electrophoresis01:22

Two-dimensional Gel Electrophoresis

Two-dimensional gel electrophoresis is a high-resolution protein separation method first introduced by O' Farrell and Klose in 1975. This method involves protein separation by two dimensions, mass and charge, making it more accurate than one-dimensional gel electrophoresis.
The first dimension separation uses the isoelectric focusing or IEF technique performed on immobilized pH gradient (IPG) strips that separate proteins according to their isoelectric points.
Biological samples, such as  cells...