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[Isolation of the theta-hemolysin of Cl. perfringens]
Abstract:
Teta-hemolysine was purified from Cl. perfringens strain BP6K 28 as follows: reprecipitation in the isoelectric point of the enzyme with 2 N H2SO4 containing 15% NaCl, DEAE cellulose chromatography, gel filtration on Sephadex G-100-G75 or Bio Gel P-60--P-100, rechromatography on DEAE-Sephadex A-50 and affinity chromatography. The biological activity of homogenous teta-hemolysine, estimated by complete hemolysis of human erythrocytes, exceeded 100, 000 theta E over mg protein but the enzyme was highly labile. Molecular weight of the enzyme was 53,000 daltons.
Insights
Clostridium perfringens theta-toxin was purified using multiple chromatography techniques. The resulting homogenous theta-hemolysin exhibited high biological activity but was unstable, with a molecular weight of 53,000 daltons.
Area of Science:
- Microbiology
- Biochemistry
Context:
- Clostridium perfringens is an opportunistic pathogen known to produce various toxins.
- Theta-toxin (also known as perfringolysin O) is a major virulence factor contributing to tissue damage.
Purpose:
- To describe the purification process of theta-hemolysin from Clostridium perfringens strain BP6K 28.
- To characterize the biological activity and molecular weight of the purified enzyme.
Summary:
- Theta-hemolysin was purified through a multi-step process involving isoelectric precipitation, DEAE cellulose chromatography, gel filtration, DEAE-Sephadex A-50 rechromatography, and affinity chromatography.
- The homogenous theta-hemolysin demonstrated significant biological activity, causing complete hemolysis of human erythrocytes at levels exceeding 100,000 theta E per mg protein.
- The purified enzyme was found to be highly labile, with a determined molecular weight of 53,000 daltons.
Impact:
- Provides a detailed method for obtaining homogenous theta-hemolysin, crucial for further structural and functional studies.
- Highlights the high specific activity of theta-hemolysin, underscoring its potent hemolytic and cytotoxic potential.
- Characterizes the lability and molecular weight of theta-hemolysin, offering essential data for understanding its stability and interactions.