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Related Experiment Video

Updated: Jun 17, 2026

Engineering Cell-permeable Protein

Engineering Cell-permeable Protein

Published on: December 28, 2009

Engineering cell-permeable protein.

Bernhard Münst¹, Christoph Patsch, Frank Edenhofer

¹Stem Cell Engineering Group, Institute of Reconstructive Neurobiology, University of Bonn - Life & Brain Center and Hertie Foundation.

Journal of Visualized Experiments : Jove

|December 30, 2009

Summary

Researchers engineered a cell-permeable Cre recombinase protein for direct delivery into mammalian cells. This protein transduction technique overcomes delivery limitations, enabling precise genetic modifications using the Cre/loxP system.

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Published on: October 17, 2013

Area of Science:

Molecular Biology
Cell Biology
Biotechnology

Background:

Protein transduction allows direct delivery of active proteins into mammalian cells.
Cell-penetrating peptides (CPPs), like the HIV-1 Tat-derived peptide, facilitate cellular uptake.
The Cre/loxP system is crucial for conditional genome editing, but active Cre delivery is challenging.

Purpose of the Study:

To engineer a cell-permeable version of the DNA-modifying enzyme Cre recombinase.
To demonstrate the construction, production, and application of this engineered protein.
To validate the functionality of the cell-permeable Cre protein in mammalian cells.

Main Methods:

Utilized the pSESAME vector system for gene insertion and domain/tag cloning.
Engineered fusion proteins incorporating CPPs and nuclear localization signals (NLS).
Expressed and purified recombinant cell-permeant proteins from E. coli.
Validated intracellular recombinase activity in cell culture.

Main Results:

Successfully constructed and produced a cell-permeable Cre recombinase.
Demonstrated that rearranging tags in the pSESAME vector system can optimize protein properties like yield and solubility.
Validated the functional activity of the recombinant Cre protein within cells.

Conclusions:

The developed protein transduction technique enables efficient delivery of active Cre recombinase.
The pSESAME vector system provides a versatile platform for engineering cell-permeable proteins.
This approach facilitates advanced applications of the Cre/loxP system for conditional genome engineering.