UBC Theses and Dissertations
Changes in toxicity of Clostridium Botulinum type E toxin by chemical modification and enzymatic cleavage Ko, Arthur S.C.
The single residue of cysteine in Cl. botulinum type E strain Iwanai toxin has been linked with toxicity, by chemical modification using p-chloromercuribenzoate. A peptide containing the oysteine residue has been isolated by exhaustive tryptic digestion of the toxin molecule tagged with N-(4-dimethylamino-3,5-dinitrophenyl) maleimide, and subsequent gel filtration with Sephadex G-25 and descending paper chromatography. The toxic peptide of trypsin-activated toxin was isolated by fractionation through a composite Sephadex G-75 and G-50 column. By chymotryptic and tryptic digestion of the toxin at pH 5.8, a toxic fragment has been isolated by gel filtration with Sephadex G-25. On the basis of quantitative amino acid analyses, the molecular weights of the intact toxin, the trypsin-activated toxin and the chymotrypsin-trypsin fragmented toxin have been estimated to be 14,000-16,000, 10,000-12,000 and 4,000-6,000 respectively. Although the mechanism of tryptic activation was found to involve chiefly the removal of at least 18 amino acid residues from the N-terminus of the toxin molecule, the manner of reduction by cleavage has not been determined for the chymotrypsin-trypsin fragmented toxin.
Item Citations and Data