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Related Experiment Video

Updated: Jun 7, 2026

One-step Purification of Twin-Strep-tagged Proteins and Their Complexes on Strep-Tactin Resin Cross-linked With Bis(sulfosuccinimidyl) Suberate (BS3)

One-step Purification of Twin-Strep-tagged Proteins and Their Complexes on Strep-Tactin Resin Cross-linked With Bis(sulfosuccinimidyl) Suberate (BS3)

Published on: April 20, 2014

Optimisation of a multivalent Strep tag for protein detection.

Michael Busby¹, Lukas Kurt Josef Stadler, Paul Ko Ferrigno

¹Department of Chemistry, University of Oxford, Mansfield Road, Oxford OX1 3QR, United Kingdom.

Biophysical Chemistry

|October 26, 2010

Summary

Adding more Strep tags to proteins enhances their binding affinity to streptavidin. This study quantizes this effect using various assays and develops a novel FRET assay for protein-tag interactions.

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Last Updated: Jun 7, 2026

One-step Purification of Twin-Strep-tagged Proteins and Their Complexes on Strep-Tactin Resin Cross-linked With Bis(sulfosuccinimidyl) Suberate (BS3)

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Published on: April 20, 2014

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Tandem Affinity Purification of Protein Complexes from Eukaryotic Cells

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Published on: January 26, 2017

Area of Science:

Biochemistry
Protein Engineering
Biotechnology

Background:

The Strep tag is a peptide sequence that mimics biotin's binding to streptavidin.
Understanding Strep tag affinity is crucial for protein engineering and purification.

Purpose of the Study:

To investigate how the number of Strep tags affects binding affinity to streptavidin.
To develop and validate a novel FRET-based assay for Strep tag interactions.

Main Methods:

Appending Strep tag sequences (0-5) to a model protein (Stefin A Quadruple Mutant - SQM).
Assessing binding affinities using Western blots, microarray assays, and surface plasmon resonance spectroscopy.
Developing an interfacial Förster Resonance Energy Transfer (FRET) based assay.

Main Results:

Binding affinity increased with the number of Strep tags, reaching nanomolar levels with 5 inserts.
The increased affinity is attributed to a higher probability of binding interactions.
A novel, cost-effective FRET assay was developed for Strep tag interactions.

Conclusions:

The number of Strep tags directly correlates with enhanced streptavidin binding affinity.
The developed FRET assay offers a valuable tool for studying protein-tag interactions without expensive equipment.
This research has implications for protein purification and bioconjugation strategies.