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Miladi B, Bouallagui H, Dridi C, et al. Reprint of: A new tagged-TEV protease: Construction, optimisation of production, purification and test activity. Protein Expr Purif. 2011;doi: 10.1016/j.pep.2011.08.018.
Miladi, B., Bouallagui, H., Dridi, C., El Marjou, A., Boeuf, G., Di Martino, P., Dufour, F., & Elm'selmi, A. (2011). Reprint of: A new tagged-TEV protease: Construction, optimisation of production, purification and test activity. Protein expression and purification, . https://doi.org/10.1016/j.pep.2011.08.018
Miladi, Baligh, et al. "Reprint of: A new tagged-TEV protease: Construction, optimisation of production, purification and test activity." Protein expression and purification vol. (2011). doi: https://doi.org/10.1016/j.pep.2011.08.018
Miladi B, Bouallagui H, Dridi C, El Marjou A, Boeuf G, Di Martino P, Dufour F, Elm'selmi A. Reprint of: A new tagged-TEV protease: Construction, optimisation of production, purification and test activity. Protein Expr Purif. 2011 Sep 02; doi: 10.1016/j.pep.2011.08.018. Epub 2011 Sep 02. PMID: 21889987.
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Protein Expr Purif. 2011 Sep 02; doi: 10.1016/j.pep.2011.08.018. Epub 2011 Sep 02.

Reprint of: A new tagged-TEV protease: Construction, optimisation of production, purification and test activity.

Protein expression and purification

Baligh Miladi, Hassib Bouallagui, Cyrine Dridi, Ahmed El Marjou, Guilhem Boeuf, Patrick Di Martino, Florence Dufour, Abdellatif Elm'selmi

Affiliations

  1. Laboratory of Molecular Biology, Ecole de Biologie Industrielle, 32 Bd du Port, Cergy 95094, France.

PMID: 21889987 DOI: 10.1016/j.pep.2011.08.018

Abstract

The Tobacco Etch Virus (TEV) protease is frequently used in the cleavage of recombinant fusion proteins because of its efficiency and high specificity. In this work, we present a new recombinant form of TEV termed Streptag II-TEV for high-level production and purification of TEV protease from Escherichia coli and compare it to the hexahistidine (6xHis) tagged version of TEV. The effects of varying the host strain, the bacterial induction temperature (25, 30 and 37°C) and the IPTG inducer concentration on production and solubility of the two recombinant TEV proteases have been examined. Optimal Streptag II-TEV protein expression were obtained in the E. coli KRX strain under an induction temperature of 25°C in the presence of IPTG at 0.5mM. In these conditions, soluble Streptag II-TEV and 6xHis-TEV proteases accounted for about 25% and 18% of total soluble proteins, respectively. About 70% of Streptag II-TEV and 60% of 6xHis-TEV were detected in the supernatant. Streptag II-TEV protease purifies to near homogeneity (approximately 99%) via a simple, single step Strep-Tactin chromatography purification protocol based on the presence of Streptag II. The higher production of Streptag II-TEV coupled to its purification and cleavage efficiencies make it an attractive alternate to 6xHis-TEV.

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