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Related Experiment Video

Updated: Jul 4, 2026

Intracellular Refolding Assay
07:18

Intracellular Refolding Assay

Published on: January 24, 2012

A medium or high throughput protein refolding assay.

Nathan P Cowieson1, Beth Wensley, Gautier Robin

  • 1Institute for Molecular Bioscience, University of Queensland, Brisbane, Australia.

Methods in Molecular Biology (Clifton, N.J.)
|June 11, 2008
PubMed
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This study introduces a simple assay to screen insoluble proteins for refolding success. The method identifies proteins amenable to refolding from inclusion bodies, aiding structural biology research.

Area of Science:

  • Structural Biology
  • Protein Biochemistry

Background:

  • Insoluble protein expression in Escherichia coli (E. coli) presents a significant challenge for high-throughput structural biology.
  • Refolding proteins from inclusion bodies into native conformations is crucial for increasing the number of targets available for structural studies.

Purpose of the Study:

  • To present a straightforward assay for screening insoluble protein targets.
  • To identify protein targets most amenable to refolding from inclusion bodies.

Main Methods:

  • The assay utilizes metal affinity resin to immobilize proteins during refolding.
  • It exploits the differential elution of folded versus misfolded proteins by imidazole.
  • Two assay implementations are described: a manual format for small-scale screening and an automated format for large-scale screening.

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Coupled Assays for Monitoring Protein Refolding in Saccharomyces cerevisiae

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Method for Efficient Refolding and Purification of Chemoreceptor Ligand Binding Domain
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Method for Efficient Refolding and Purification of Chemoreceptor Ligand Binding Domain

Published on: December 12, 2017

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Last Updated: Jul 4, 2026

Intracellular Refolding Assay
07:18

Intracellular Refolding Assay

Published on: January 24, 2012

Coupled Assays for Monitoring Protein Refolding in Saccharomyces cerevisiae
13:52

Coupled Assays for Monitoring Protein Refolding in Saccharomyces cerevisiae

Published on: July 9, 2013

Method for Efficient Refolding and Purification of Chemoreceptor Ligand Binding Domain
14:25

Method for Efficient Refolding and Purification of Chemoreceptor Ligand Binding Domain

Published on: December 12, 2017

Main Results:

  • The assay effectively distinguishes between folded and misfolded proteins based on imidazole elution profiles.
  • It integrates seamlessly into existing high-throughput structural biology pipelines.
  • The assay uses inclusion body preparations, a byproduct of expression and purification trials, requiring no additional facilities.

Conclusions:

  • This refolding assay provides a valuable tool for overcoming bottlenecks in structural biology.
  • It enhances the efficiency of identifying suitable protein targets for structural studies.
  • The assay's adaptability for both manual and automated screening supports diverse research needs.