Purification and refolding of recombinant human proMMP-7 (pro-matrilysin) expressed in Escherichia coli and its characterization.

Purification and refolding of recombinant human proMMP-7 (pro-matrilysin) expressed in Escherichia coli and its characterization. Research Abstract Details 

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Purification and refolding of recombinant human proMMP-7 (pro-matrilysin) expressed in Escherichia coli and its characterization. Abstract Text:

M Itoh,K Masuda,Y Ito,T Akizawa,M Yoshioka,K Imai,Y Okada,H Sato,M Seiki,

Human matrix metalloproteinase-7 (MMP-7 = matrilysin) was overproduced in Escherichia coli as a recombinant zymogen (31 kDa), the C-terminus of which bears artificial hexa-histidines. Most of the enzyme was isolated from the insoluble fraction of the cell lysate and purified by a single step using Ni-NTA resin after solubilization of the precipitates with 8 M urea solution. The resin-bound recombinant protein was refolded into a form that is activatable by p-amino-phenylmercuric acetate in an autocatalytic manner. The activated enzyme cleaved a synthetic peptide substrate at the reported site for MMP-7. Digestion of carboxymethylated transferrin (a natural substrate of MMP-7) by the recombinant proteinase generated fragments with the same peptide map as in the case of native purified MMP-7. The autocatalytic activation and enzyme reaction were entirely dependent on the presence of calcium and zinc ions. The enzyme activity to cleave carboxymethylated transferrin was inhibited by tissue inhibitors of metalloproteinases-1 and -2, MMP-specific inhibitors. The activity of the recombinant MMP-7 was also inhibited by a synthetic peptide derived from a part of the cysteine switch that maintains the zymogen in an inactive state. Thus, we report here a simple means of preparing a large quantity of recombinant proMMP-7 that can be used to study the activation mechanism and to screen synthetic inhibitors.

Purification and refolding of recombinant human proMMP-7 (pro-matrilysin) expressed in Escherichia coli and its characterization. Publishing Authors By Initials

M Itoh,K Masuda,Y Ito,T Akizawa,M Yoshioka,K Imai,Y Okada,H Sato,M Seiki,

For similar proteins: blood proteins: acute-phase proteins: transferrin research abstracts see: proteins: blood proteins: acute-phase proteins: transferrin research

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Purification and refolding of recombinant human proMMP-7 (pro-matrilysin) expressed in Escherichia coli and its characterization. Journal Published:

PUBLICATION TYPE: Research Support, Non-U.S. Gov

Journal: Journal of biochemistry

VOLUME: 119

Page Numbers: 667-73

Journal Abbreviation: J. Biochem.

ISSN: 0021-924X

DAY: 19

MONTH: Apr

YEAR: 1996

Purification and refolding of recombinant human proMMP-7 (pro-matrilysin) expressed in Escherichia coli and its characterization. Information

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LANGUAGE: eng

NlmUniqueID: 376600

Purification and refolding of recombinant human proMMP-7 (pro-matrilysin) expressed in Escherichia coli and its characterization. Keywords Mesh Terms:

KEYWORDS: Transferrin

MESH TERMS: metabolism

Chemical & Substance for Abstract: Purification and refolding of recombinant human proMMP-7 (pro-matrilysin) expressed in Escherichia coli and its characterization. Information

Substance Name: promatrilysin

Registry Number: EC 3.4.24.-

Grant and Affiliation Information for Purification and refolding of recombinant human proMMP-7 (pro-matrilysin) expressed in Escherichia coli and its characterization.

AFFILIATION: Department of Analytical Chemistry, Faculty of Pharmaceutical Sciences, Setsunan University, Osaka.

Country: JAPAN

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MEDLINETA: J Biochem

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