DOI: 10.1111/j.1432-1033.1995.0533k.x. The Preparation of Catalytically Active Human Cathepsin B from Its Precursor Expressed in <i>Escherichia coli</i> in the Form of Inclusion Bodies

The Preparation of Catalytically Active Human Cathepsin B from Its Precursor Expressed in Escherichia coli in the Form of Inclusion Bodies

10.1111/j.1432-1033.1995.0533k.x

Crossref journal-article

Wiley

European Journal of Biochemistry (311)

Abstract

A cDNA clone encoding human procathepsin B was expressed at a high level in Escherichia coli using a T7 polymerase expression system, resulting in the formation of insoluble cytoplasmic protein aggregates (inclusion bodies). The recombinant product was solubilized and renatured by refolding and reoxidation. The proenzyme was subsequently processed with pepsin to produce an enzymically active enzyme. By systematic variation of the parameters influencing the folding, formation of disulphide bonds, and processing of procathepsin B to the catalytically active mature form, a simple renaturation procedure was designed, allowing the production of about 3 mg purified active cathepsin B/1 E. coli culture broth. The enzyme obtained in this way consists of a single chain and, as a consequence of pepsin treatment, possesses a three‐amino‐acid extension at its N‐terminus. The enzyme has similar kinetic and immunological properties to native human cathepsin B.

Bibliography

Kuhelj, R., Dolinar, M., PungerÄar, J., & Turk, V. (1995). The Preparation of Catalytically Active Human Cathepsin B from Its Precursor Expressed in Escherichia coli in the Form of Inclusion Bodies. European Journal of Biochemistry, 229(2), 533â539. Portico.

Authors 4

Robert Kuhelj (first)
Marko Dolinar (additional)
Jože Pungerčar (additional)
Vito Turk (additional)

References 40 Referenced 98

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{'key': 'e_1_2_3_3_1', 'first-page': '256', 'article-title': 'Lysosomal cysteine proteinases', 'volume': '1', 'author': 'Barrett A. J.', 'year': '1988', 'journal-title': 'ISI Atlas Sci. Biochem.'} / ISI Atlas Sci. Biochem. / Lysosomal cysteine proteinases by Barrett A. J. (1988)
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{'key': 'e_1_2_3_25_1', 'first-page': '163', 'article-title': 'Expression of rat and mouse cathepsin B precursors in Escherichia coli', 'volume': '369', 'author': 'Mort J. S.', 'year': '1988', 'journal-title': 'Biol. Chem. Hoppe-Seyler'} / Biol. Chem. Hoppe-Seyler / Expression of rat and mouse cathepsin B precursors in Escherichia coli by Mort J. S. (1988)
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{'key': 'e_1_2_3_41_1', 'first-page': '509', 'article-title': 'Some characterizations of cathepsin B and α‐N‐benzoylarginine‐β‐naphtylamide hydrolase from bovine lymph nodes', 'volume': '53', 'author': 'Zvonar‐Popoviˇ T.', 'year': '1980', 'journal-title': 'Croat. Chem. Acta'} / Croat. Chem. Acta / Some characterizations of cathepsin B and α‐N‐benzoylarginine‐β‐naphtylamide hydrolase from bovine lymph nodes by Zvonar‐Popoviˇ T. (1980)

Dates

Type	When
Created	20 years, 5 months ago (March 4, 2005, 9:30 a.m.)
Deposited	1 year, 9 months ago (Nov. 22, 2023, 5:46 p.m.)
Indexed	1 day, 23 hours ago (Aug. 27, 2025, 12:27 p.m.)
Issued	30 years, 4 months ago (April 1, 1995)
Published	30 years, 4 months ago (April 1, 1995)
Published Online	20 years, 5 months ago (March 3, 2005)
Published Print	30 years, 4 months ago (April 1, 1995)

Funders 0

None

BibTeX

@article{Kuhelj_1995, title={The Preparation of Catalytically Active Human Cathepsin B from Its Precursor Expressed in Escherichia coli in the Form of Inclusion Bodies}, volume={229}, ISSN={1432-1033}, url={http://dx.doi.org/10.1111/j.1432-1033.1995.0533k.x}, DOI={10.1111/j.1432-1033.1995.0533k.x}, number={2}, journal={European Journal of Biochemistry}, publisher={Wiley}, author={Kuhelj, Robert and Dolinar, Marko and Pungerčar, Jože and Turk, Vito}, year={1995}, month=apr, pages={533–539} }

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