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Related Concept Videos

Protein Networks02:26

Protein Networks

3.9K
An organism can have thousands of different proteins, and these proteins must cooperate to ensure the health of an organism. Proteins bind to other proteins and form complexes to carry out their functions. Many proteins interact with multiple other proteins creating a complex network of protein interactions.
These interactions can be represented through maps depicting protein-protein interaction networks, represented as nodes and edges. Nodes are circles that are representative of a protein,...
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Protein-protein Interfaces02:04

Protein-protein Interfaces

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Many proteins form complexes to carry out their functions, making protein-protein interactions (PPIs) essential for an organism's survival. Most PPIs are stabilized by numerous weak noncovalent chemical forces. The physical shape of the interfaces determines the way two proteins interact. Many globular proteins have closely-matching shapes on their surfaces, which form a large number of weak bonds. Additionally, many PPIs occur between two helices or between a surface cleft and a...
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Tagging and Fusion Proteins01:24

Tagging and Fusion Proteins

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Proteins are involved in several cellular processes and biochemical reactions. Analyzing a specific protein of interest requires it to be isolated from the other proteins in the cell. This is achieved by overexpressing the specific gene in a suitable host to produce large quantities of the target protein. A tag or label is recombined with the gene to produce a fusion protein containing the target protein and the tag. The tags on these fusion proteins can then be used for easy detection and...
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Related Experiment Video

Updated: Jun 8, 2025

Resolving Affinity Purified Protein Complexes by Blue Native PAGE and Protein Correlation Profiling
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Protocol for mapping differential protein-protein interaction networks using affinity purification-mass spectrometry.

Prashant Kaushal1, Manisha R Ummadi2, Gwendolyn M Jang2

  • 1Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA, USA; Institute for Quantitative and Computational Biosciences, University of California, Los Angeles, Los Angeles, CA, USA; Molecular Biology Institute, University of California Los Angeles, Los Angeles, CA, USA.

STAR Protocols
|November 3, 2024
PubMed
Summary

This study introduces a new proteomics protocol using affinity purification-mass spectrometry (AP-MS) to analyze protein complexes. The method helps identify changes in protein interactions linked to health and disease.

Keywords:
Mass SpectrometryMolecular BiologyProtein BiochemistryProtein expression and purificationProteomicsSystems biology

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Area of Science:

  • Molecular Biology
  • Proteomics
  • Biochemistry

Background:

  • Proteins function within complexes, crucial for cellular processes.
  • Alterations in protein complexes, due to mutations or signaling, impact health and disease.
  • Understanding protein-protein interactions is key to deciphering cellular mechanisms.

Purpose of the Study:

  • To present a robust affinity purification-mass spectrometry (AP-MS) protocol.
  • To enable the study of protein-protein interaction networks in mammalian cells.
  • To facilitate the identification of differential protein interactions under various conditions.

Main Methods:

  • Expression of affinity-tagged 'bait' proteins in mammalian cells.
  • Affinity purification of protein complexes.
  • Mass spectrometry for identification and quantification of interacting proteins.
  • Comparative analysis of protein interaction networks between conditions.

Main Results:

  • Successful identification and quantification of protein interactors.
  • Visualization of differential protein-protein interaction networks.
  • Demonstration of the protocol's applicability across cell types and biological contexts.

Conclusions:

  • The presented AP-MS protocol is a versatile tool for studying protein complex dynamics.
  • This method aids in understanding how protein interaction network changes contribute to phenotypic outcomes.
  • The protocol supports research in various biological areas, including health and disease.