Coat and enterotoxin-related proteins in Clostridium perfringens spores.
Coat proteins from mature spores of two enterotoxin-positive (Ent+) and two enterotoxin-negative (Ent-) strains of Clostridium perfringens were solubilized using 50 mM-dithiothreitol and 1% sodium dodecyl sulphate at pH 9.7, and alkylated using 110 mM-iodoacetamide to prevent aggregation. The coat proteins and C. perfringens type A enterotoxin (CPE) were separated by SDS-PAGE and analysed by Western blotting using anti-CPE antibody. As previously reported, CPE aggregated in the presence of SDS, but no aggregation occurred at concentrations below 15 micrograms CPE ml-1. Two CPE-related proteins (34 and 48 kDa) were found in the solubilized spore coat protein of Ent+ strains while only the 48 kDa CPE-related protein was found in the spore coat fraction of Ent- strains. CPE-related proteins comprised 2.7% and 0.8% of the total solubilized coat protein of Ent+ and Ent- strains respectively. CPE-related proteins could be extracted from the spores with 1% SDS alone. They could also be released by disruption of whole spores, indicating that the CPE-related proteins may be in the spore core or trapped between the core and coat layers. The results suggest that CPE is not a major structural component of the coat fraction of C. perfringens spores.
Ryu
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Department of Food Science, University of Massachusetts, Amherst 01003.
Labbe
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D001426:Bacterial Proteins / Q000276:immunology / Q000302:isolation & purification*; D003016:Clostridium perfringens / Q000032:analysis*; D004229:Dithiothreitol; D004591:Electrophoresis, Polyacrylamide Gel; D004768:Enterotoxins / Q000276:immunology / Q000302:isolation & purification; D012967:Sodium Dodecyl Sulfate; D013171:Spores, Bacterial / Q000032:analysis
D001426:Bacterial Proteins; D004768:Enterotoxins; C035598:enterotoxin, Clostridium; D012967:Sodium Dodecyl Sulfate; D004229:Dithiothreitol