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Production of a non-toxic site-directed mutant of Clostridium perfringens -toxin which induces protective immunity in mice

Defence Evaluation and Research Agency, CBD Porton Down, Salisbury, Wiltshire SP4 0JQ, UK

ABSTRACT

A panel of ten site-directed mutants of Clostridium perfringens -toxin was generated. All of the mutated proteins expressed in Escherichia coli were recognized in immunoblots by a neutralizing mAb raised against wild-type native -toxin. The cytotoxicity of the site-directed mutated toxins was assayed in vitro against MDCK cells. One mutation resulting in loss of activity in the assay was identified. This non-toxic protein was derived by substituting a proline for the histidine at residue 106 of the toxin. Immunization of mice with the non-toxic mutated -toxin resulted in the induction of a specific antibody response and immunized mice were protected against 1000 LD₅₀ doses of wild-type recombinant -toxin.

Author for correspondence: Petra C. F. Oyston. Tel: +44 1980 613320. Fax: +44 1980 613284. e-mail: 100432.3200@compuserve.com

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