Purification, characterization, and primary structure of Clostridium perfringens lambda-toxin, a thermolysin-like metalloprotease

F. U. Jin, Osamu Matsushita, Sej Ichi Katayama, Shengyong Jin, Chieko Matsushita, Junzaburo Minami, Akinobu Okabe

Research output: Contribution to journalArticle

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Abstract

The lambda-toxin of Clostridium perfringens type B NCIB10691 was purified by ammonium sulfate precipitation, followed by size exclusion, anion- exchange, and hydrophobic interaction chromatography. The purified toxin had an apparent molecular mass of 36 kDa, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The toxin possessed casein- hydrolyzing activity, which was inhibited specifically by metal chelators, indicating that the toxin is a metalloprotease. The gene encoding the lambda- toxin (lam), which was shown by Southern analysis to be located on a 70-kb plasmid, was cloned into Escherichia coli cells. Nucleotide and N-terminal amino acid sequencing revealed that the lam gene encodes a 553-amino-acid protein, which is processed into a mature form, the molecular mass of which was calculated to be 35,722 Da. The deduced amino acid sequence of the mature enzyme contains an HEXXH motif characteristic of zinc metalloproteases and is homologous to other known enzymes belonging to the thermolysin family. The purified toxin degraded various biologically important substances, such as collagen, fibronectin, fibrinogen, immunoglobulin A, and the complement C3 component. It caused an increase in vascular permeability and hemorrhagic edema on injection into the dorsal skin of mice. These results suggest that the toxin contributes to the pathogenesis of histolytic infection by lambda- toxin-producing C. perfringens.

Original languageEnglish
Pages (from-to)230-237
Number of pages8
JournalInfection and Immunity
Volume64
Issue number1
Publication statusPublished - 1996
Externally publishedYes

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Thermolysin
Metalloproteases
Complement C3
Protein Sequence Analysis
Ammonium Sulfate
Capillary Permeability
Enzymes
Chelating Agents
Caseins
Hydrophobic and Hydrophilic Interactions
Fibronectins
Sodium Dodecyl Sulfate
Fibrinogen
Immunoglobulin A
Genes
Anions
Chromatography
Zinc
Polyacrylamide Gel Electrophoresis
Amino Acid Sequence

ASJC Scopus subject areas

  • Immunology

Cite this

Purification, characterization, and primary structure of Clostridium perfringens lambda-toxin, a thermolysin-like metalloprotease. / Jin, F. U.; Matsushita, Osamu; Katayama, Sej Ichi; Jin, Shengyong; Matsushita, Chieko; Minami, Junzaburo; Okabe, Akinobu.

In: Infection and Immunity, Vol. 64, No. 1, 1996, p. 230-237.

Research output: Contribution to journalArticle

Jin, F. U. ; Matsushita, Osamu ; Katayama, Sej Ichi ; Jin, Shengyong ; Matsushita, Chieko ; Minami, Junzaburo ; Okabe, Akinobu. / Purification, characterization, and primary structure of Clostridium perfringens lambda-toxin, a thermolysin-like metalloprotease. In: Infection and Immunity. 1996 ; Vol. 64, No. 1. pp. 230-237.
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