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Infect Immun. 1983 May; 40(2): 701-707

Purification and characterization of heat-stable enterotoxin from bovine enterotoxigenic Escherichia coli.

ABSTRACT

Heat-stable enterotoxin (ST) from Escherichia coli pathogenic for cattle was mass produced in a chemically defined medium. The toxin was concentrated and purified by sequentially applying batch adsorption chromatography on Amberlite XAD-2 resin, acetone fractionation, and preparative isoelectric focusing in a flatbed granulated gel. Reverse-phase high-performance liquid chromatography was used to purify the toxin further and to eliminate contaminating ampholytes. The toxin was purified more than 2,000-fold and had a minimal effective dose of less than 0.5 ng. It was biologically active after heating to 100 degrees C for 30 min and was not hydrolyzed by trypsin, pronase, and subtilisin, but it was inactivated by treatment with 0.1 M 2-mercaptoethanol or 4 X 10(-5) M dithiothreitol, suggesting that disulfide bonds are essential for retaining its biological activity. The amino acid analysis revealed 18 amino acid residues per molecule, which is in agreement with the composition of ST from a human strain of enterotoxigenic E. coli. The amino acid composition of our ST matched the published coding sequence of the last 18 codons of Tn1618, a transposon isolated from the bovine enterotoxigenic E. coli strain B41 and shown to be present also in some strains of porcine enterotoxigenic E. coli. These findings further support the existence of a form of ST common to bovine, porcine, and human strains of enterotoxigenic E. coli.