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Related Experiment Video

Updated: Jun 28, 2026

Generation of Native, Untagged Huntingtin Exon1 Monomer and Fibrils Using a SUMO Fusion Strategy

Generation of Native, Untagged Huntingtin Exon1 Monomer and Fibrils Using a SUMO Fusion Strategy

Published on: June 27, 2018

PHB-intein-mediated protein purification strategy.

Alison R Gillies¹, Reza Banki Mahmoud, David W Wood

¹Department of Chemical Engineering, Princeton University, Princeton, NJ, USA.

Methods in Molecular Biology (Clifton, N.J.)

|November 7, 2008

Summary

This study introduces a novel method for purifying recombinant proteins using polyhydroxybutyrate (PHB) particles, eliminating the need for complex chromatography. This tag-based purification strategy offers an efficient way to isolate target proteins from Escherichia coli.

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Last Updated: Jun 28, 2026

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Synthesis of an Intein-mediated Artificial Protein Hydrogel

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Published on: January 27, 2014

Area of Science:

Biotechnology
Protein Purification
Molecular Biology

Background:

Traditional recombinant protein purification often relies on complex and costly chromatographic techniques.
Affinity tags simplify purification but can require specific resins and elution conditions.
Escherichia coli is a common host for recombinant protein expression, but purification can be challenging.

Purpose of the Study:

To develop a simplified and efficient method for purifying recombinant proteins from Escherichia coli.
To replace conventional chromatographic methods with an in vivo affinity-based purification strategy.
To utilize polyhydroxybutyrate (PHB) particles as a purification matrix.

Main Methods:

Genetically fusing a PHB-binding protein (phasin) to the target protein via a self-cleaving intein linker.
Co-expressing the tagged protein and PHB particles in Escherichia coli.
Purifying the PHB-protein complex through cell lysis, washing, centrifugation, and resuspension.
Inducing self-cleavage of the intein linker via a pH shift to release the native target protein.

Main Results:

Successful purification of recombinant proteins using PHB particles as an affinity matrix.
Achieved yields of 35-40 microg of purified product per milliliter of cell culture.
Demonstrated efficient separation of the target protein from the PHB-phasin tag.

Conclusions:

The developed method offers a cost-effective and simplified alternative to traditional protein purification techniques.
This PHB-based purification strategy is applicable to a wide range of recombinant proteins and expression hosts.
The use of self-cleaving inteins allows for the recovery of native target proteins.