Clostridium botulinum type D toxin: purification, molecular structure, and some immunological properties

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Abstract
Clostridium botulinum type D progenitor toxin was purified. The addition of ribonucleic acid to the whole culture helped initial acid precipitation of the toxin. As with type B, both L (16S) and M toxins (12S) obtained from a hemagglutinin-positive strain, whereas M toxin only was produced by a hemagglutinin-negative strain. M toxin (molecular weight, 300,000) consisted of one molecule each of a toxic (molecular weight, 170,000) and a nontoxic component (molecular weight, 130,000); L toxin consisted of both components plus hemagglutinin. The specific toxicity of M toxin was 5 X 10(8) mean lethal doses per mg of N; that of L toxin was 2.4 X 10(8) mean lethal doses per mg of N. These toxins were fully or nearly fully active, but in un-nicked form. Trypsinization caused nicking in the toxic component, forming a molecule made up of two peptide chains with molecular weights of 110,000 and 60,000; there was little or no increase in toxicity. The toxic component of type D was not antigenically related to that of type C, whereas the nontoxic component was antigenically indistinguishable from that of type C. The toxicities of both L nad M toxins of the hemagglutinin-positive strain were increased twofold by trypsinization. Neither toxin contained the C2 toxic factor elaborated by C and D strain.