Identification of residues critical for toxicity in Clostridium perfringens phospholipase C, the key toxin in gas gangrene: Residues critical for toxicity in phospholipase C
- Citation data:
European Journal of Biochemistry, ISSN: 0014-2956, Vol: 267, Issue: 16, Page: 5191-5197
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- Repository URL:
- http://hdl.handle.net/10669/29468; http://onlinelibrary.wiley.com/doi/10.1046/j.1432-1327.2000.01588.x/abstract;jsessionid=C9CB49492B0F301FFEFD4283817CDE46.f03t01
- Biochemistry, Genetics and Molecular Biology; Bacterial Toxins; Muscular Diseases; Molecular Models; Skeletal Muscle; Cell Survival
Clostridium perfringens phospholipase C (PLC), also called α-toxin, is the major virulence factor in the pathogenesis of gas gangrene. The toxic activities of genetically engineered α-toxin variants harboring single amino-acid substitutions in three loops of its C-terminal domain were studied. The substitutions were made in aspartic acid residues which bind calcium, and tyrosine residues of the putative membrane-interacting region. The variants D269N and D336N had less than 20% of the hemolytic activity and displayed a cytotoxic potency 10-fold lower than that of the wild-type toxin. The variants in which Tyr275, Tyr307, and Tyr331 were substituted by Asn, Phe, or Leu had 11-73% of the hemolytic activity and exhibited a cytotoxic potency 10- to 10-fold lower than that of the wild-type toxin. The results demonstrated that the sphingomyelinase activity and the C-terminal domain are required for myotoxicity in vivo and that the variants D269N, D336N, Y275N, Y307F, and Y331L had less than 12% of the myotoxic activity displayed by the wild-type toxin. This work therefore identifies residues critical for the toxic activities of C. perfringens PLC and provides new insights toward understanding the mechanism of action of this toxin at a molecular level.