Characterization of the neurotoxin isolated from a Clostridium baratii strain implicated in infant botulism.
Botulism is widely known to result from ingestion of food containing botulinum neurotoxin produced in situ by certain strains of Clostridium botulinum. Infant botulism caused by C. botulinum, unlike the food-borne intoxication, is the toxicoinfectious form of botulism (S. S. Arnon, p. 331-345, in G. E. Lewis, ed., Biomedical Aspects of Botulism, 1981). The strain of Clostridium baratii implicated in infant botulism produced a neurotoxin that was neutralized with antiserum for botulinum neurotoxin serotype F (J. D. Hall, L. M. McCroskey, B. J. Pincomb, and C. L. Hatheway, J. Clin. Microbiol. 21:654-655, 1985). We developed a procedure to culture the toxigenic C. baratii (strain 6341) in dialysis bags and a simple purification scheme (precipitation of 900-ml culture supernatant with ammonium sulfate and two anion-exchange chromatographic steps at pH 5.5 and 8.0) that yielded up to 150 micrograms of purified neurotoxin. It is an approximately 140-kDa single-chain protein and has the following sequence of amino acid residues at the N terminus: Pro-Val-Asn-Ile-Asn-Asn-Phe-Asn-Tyr-Asn-Asp-Pro-Ile-Asn-Asn-Thr-Thr-Ile- Leu. Comparison of this amino acid sequence with those of the botulinum neurotoxin serotypes A, B, and E showed 40 to 50% identical residues in comparable positions. The specific toxicity of the neurotoxin, approximately 2 x 10(6) 50% lethal doses for mice per mg of protein injected, was not enhanced significantly by mild trypsinization, although the protease cleaved the neurotoxin within a disulfide loop that generated at least two primary fragments, approximately 47 and approximately 86 kDa, that remained linked by an interchain disulfide. These two fragments resembled the light and heavy chains of the well-characterized neurotoxin serotypes A, B, C, D, E, and F produced by C. botulinum.
Giménez
J A
JA
Department of Food Microbiology and Toxicology, University of Wisconsin, Madison 53706-1187.
Giménez
M A
MA
DasGupta
B R
BR
D000595:Amino Acid Sequence; D000818:Animals; D001906:Botulism / Q000209:etiology*; D003013:Clostridium / Q000378:metabolism* / Q000472:pathogenicity; D006801:Humans; D007928:Lethal Dose 50; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009498:Neurotoxins / Q000302:isolation & purification* / Q000633:toxicity; D011817:Rabbits
D009498:Neurotoxins