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Related Concept Videos

Molecular Chaperones and Protein Folding03:00

Molecular Chaperones and Protein Folding

The native conformation of a protein is formed by interactions between the side chains of its constituent amino acids. When the amino acids cannot form these interactions, the protein cannot fold by itself and needs chaperones. Notably, chaperones do not relay any additional information required for the folding of polypeptides; the native conformation of a protein is determined solely by its amino acid sequence. Chaperones catalyze protein folding without being a part of the folded protein.
The...
Protein Folding01:25

Protein Folding

Proteins are chains of amino acids linked together by peptide bonds. Upon synthesis, a protein folds into a three-dimensional conformation, critical to its biological function. Interactions between its constituent amino acids guide protein folding, and hence the protein structure is primarily dependent on its amino acid sequence.
Protein Structure Is Critical to Its Biological Function
Proteins perform a wide range of biological functions such as catalyzing chemical reactions, providing...
Protein Folding01:22

Protein Folding

Overview
Protein Folding Quality Check in the RER01:29

Protein Folding Quality Check in the RER

ER is the primary site for the maturation and folding of soluble and transmembrane secretory proteins. The calnexin cycle is a specific chaperone system that folds and assesses the confirmation of N-glycosylated proteins before they can exit the ER lumen. The primary players of this quality check pipeline are the lectins, ER-resident chaperones, and a glucosyl transferase enzyme. In case the calnexin system in the lumen fails to salvage a misfolded protein, it is transported to the cytoplasm...
Export of Misfolded Proteins out of the ER01:32

Export of Misfolded Proteins out of the ER

After folding, the ER assesses the quality of secretory and membrane proteins. The correctly folded proteins are cleared by the calnexin cycle for transport to their final destination, while misfolded proteins are held back in the ER lumen. The ER chaperones attempt to unfold and refold the misfolded proteins but sometimes fail to achieve the correct native conformation. Such terminally misfolded proteins are then exported to the cytosol by ER-associated degradation or ERAD pathway for...
Bacterial Protein Maturation01:26

Bacterial Protein Maturation

Bacterial protein maturation is a tightly regulated process that ensures newly synthesized polypeptides achieve correct functional conformations. This maturation involves a series of modifications, folding events, and quality control steps, often assisted by specialized chaperone proteins.N-Terminal ModificationsThe maturation of bacterial polypeptides begins cotranslationally as the polypeptide exits the ribosome. The first amino acid, N-formylmethionine (fMet), is typically modified at the...

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Related Experiment Video

Updated: Jun 19, 2026

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

Refolding solubilized inclusion body proteins.

Richard R Burgess1

  • 1McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, Wisconsin, USA.

Methods in Enzymology
|November 7, 2009
PubMed
Summary

Recombinant protein purification often yields insoluble inclusion bodies (IBs). This work details strategies to solubilize IBs and refold proteins into active forms, crucial for biotechnology and research.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Protein Chemistry

Background:

  • Recombinant protein expression in hosts like Escherichia coli is standard practice.
  • Many expressed proteins aggregate into insoluble inclusion bodies (IBs).
  • Washed inclusion bodies offer a relatively pure source of target protein.

Purpose of the Study:

  • To outline techniques for solubilizing inclusion bodies.
  • To describe strategies for refolding proteins from inclusion bodies into their native, biologically active state.
  • To provide guidance on overcoming challenges in recombinant protein refolding.

Main Methods:

  • Isolation and washing of inclusion bodies.
  • Solubilization of inclusion bodies using denaturants.

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Purification and Refolding to Amyloid Fibrils of (His)6-tagged Recombinant Shadoo Protein Expressed as Inclusion Bodies in E. coli

Published on: December 19, 2015

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  • Refolding of solubilized proteins by altering buffer conditions (e.g., pH, redox potential, additives).
  • Assessment of protein refolding and biological activity.

    • Main Results:

    • Inclusion bodies provide a concentrated source of recombinant proteins.
    • Successful solubilization requires appropriate chemical agents.
    • Optimized refolding conditions are protein-specific and critical for regaining function.
    • A systematic approach to refolding can overcome aggregation issues.

    Conclusions:

    • Solubilization and refolding of inclusion bodies are key steps in obtaining active recombinant proteins.
    • The process requires careful optimization for each individual protein.
    • Effective strategies enable the recovery of functional proteins from insoluble aggregates.