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Related Concept Videos

Covalently Linked Protein Regulators02:04

Covalently Linked Protein Regulators

Proteins can undergo many types of post-translational modifications, often in response to changes in their environment. These modifications play an important role in the function and stability of these proteins. Covalently linked molecules include functional groups, such as methyl, acetyl, and phosphate groups, and also small proteins, such as ubiquitin. There are around 200 different types of covalent regulators that have been identified.
These groups modify specific amino acids in a protein.
Regulated Protein Degradation02:58

Regulated Protein Degradation

It is vital to regulate the activity of enzymatic as well as non-enzymatic proteins inside the cell. This can be achieved either through creating a balance between their rate of synthesis and degradation or regulating the intrinsic activity of the protein. Both these regulation mechanisms play an essential role in the normal functioning of cells.
Protein degradation plays two important roles in the cells. It helps to protect cells from misfolded or damaged proteins before they lead to a...

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Related Experiment Video

Updated: Jun 8, 2026

In Vitro Analysis of E3 Ubiquitin Ligase Function
06:06

In Vitro Analysis of E3 Ubiquitin Ligase Function

Published on: May 14, 2021

Novel approach for characterizing ubiquitin E3 ligase function.

Jeffrey G Marblestone1, K G Suresh Kumar, Michael J Eddins

  • 1Progenra, Inc., Malvern, Pennsylvania, USA. Marblestone@progenra.com

Journal of Biomolecular Screening
|September 25, 2010
PubMed
Summary
This summary is machine-generated.

A new assay platform enables rapid, cost-effective screening of E3 ubiquitin ligase activity, crucial for understanding disease and accelerating drug discovery. This method aids in identifying novel inhibitors for E3 ligases involved in various pathologies.

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Evaluation of Substrate Ubiquitylation by E3 Ubiquitin-ligase in Mammalian Cell Lysates
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Evaluation of Substrate Ubiquitylation by E3 Ubiquitin-ligase in Mammalian Cell Lysates

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Related Experiment Videos

Last Updated: Jun 8, 2026

In Vitro Analysis of E3 Ubiquitin Ligase Function
06:06

In Vitro Analysis of E3 Ubiquitin Ligase Function

Published on: May 14, 2021

Functional Characterization of RING-Type E3 Ubiquitin Ligases In Vitro and In Planta
10:27

Functional Characterization of RING-Type E3 Ubiquitin Ligases In Vitro and In Planta

Published on: December 5, 2019

Evaluation of Substrate Ubiquitylation by E3 Ubiquitin-ligase in Mammalian Cell Lysates
09:47

Evaluation of Substrate Ubiquitylation by E3 Ubiquitin-ligase in Mammalian Cell Lysates

Published on: May 10, 2022

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Drug Discovery

Background:

  • The ubiquitin-proteasome system regulates cellular events; its dysregulation is linked to disease.
  • E3 ubiquitin ligases are key regulators, coordinating substrate ubiquitylation and degradation.
  • Current limitations in high-throughput screening (HTS)-compliant E3 ligase assays hinder inhibitor discovery.

Purpose of the Study:

  • To develop a novel HTS-compliant assay platform for quantifying E3 ligase activity.
  • To demonstrate the broad utility of this platform across different E3 ligase families.
  • To validate the platform's effectiveness in identifying E3 ligase inhibitors.

Main Methods:

  • Utilized a ubiquitin binding domain's affinity for polyubiquitin chains.
  • Developed an assay to analyze E3 ligase-dependent polyubiquitin chain formation.
  • Applied the assay to RING and HECT family E3 ligases.

Main Results:

  • Successfully demonstrated the assay's broad applicability to diverse E3 ligases.
  • Identified inhibitors of the E3 ligase CARP2 using the developed platform.
  • Established a rapid and cost-effective method for monitoring E3 ligase activity.

Conclusions:

  • The novel assay platform is HTS-compliant and broadly applicable to E3 ligases.
  • This platform facilitates the discovery of E3 ligase inhibitors for therapeutic development.
  • Expeditious quantification of E3 ligase activity is essential for drug discovery in disease-associated pathways.