The present invention relates to the isolation of polypeptides derived from the Clostridium botulinum neurotoxin and the use thereof as immunogens for the production of vaccines and antitoxins, as well as research and drug development applications. Clostridium botulinum is responsible for food bone botulism, a severe and often deadly disease. Botulinum neurotoxin binds to neural cells and are translocated into the cytosol. The toxin then prevents neurotransmitter release by cleaving a SNARE protein. A double mutant E224A/E262 full length botulinum neurotoxin Type A holo form was successfully cloned and purified, which lacks the endopeptidase activity involved in the toxic action of the BoNT, and thus leading to its detoxification (DR BoNT/A). This new molecule can be used as an antidote against botulism, and also as a vaccine candidate for botulism. Due to the poor availability and extreme toxicity of native holo-toxin, a nontoxic form of the holo-toxin is highly desired for research and vaccine development. The full length DR BoNT/A protein has been expressed in E. coli as a soluble form.
