Affiliation:
1. Department of Microbiology, University of Kansas, Lawrence, Kansas 66045
Abstract
Heat-labile enterotoxin (LT) produced by a human strain of enterotoxigenic
Escherichia coli
(286C
2
) was purified to homogeneity from pH extracts of fermentor-grown cells by ultrafiltration, (NH
4
)
2
SO
4
fractionation, hydrophobic chromatography on norleucine-Sepharose 4B, hydroxylapatite chromatography, and Bio-Gel P-150 filtration. Purified LT preparations exhibited biological activity comparable to that of cholera toxin in four bioassays specific for the two enterotoxins (Y-1 adrenal tumor cells, Chinese hamster ovary cells, pigeon erythrocyte lysates, and skin permeability test). The overall yield of LT protein was 20%, which represented a 500-fold purification over pH extracts. A native molecular weight of 73,000 was determined by gel electrophoresis. The toxin dissociated upon treatment with sodium dodecyl sulfate, pH 7.0, into two components with molecular weights of 44,000 and 30,000. Purified LT preparations were remarkably stable over a wide range of storage conditions, temperatures, and pH's. The biological activity was increased by incubation with trypsin and completely destroyed by pronase and proteinase K, whereas deoxyribonuclease I, ribonuclease, and phospholipase D had no effect. The amino acid composition of purified LT was quite different from that of cholera toxin. Neither carbohydrate nor lipopolysaccharide was present in purified preparations. The purification scheme appeared applicable to LT produced by other human and porcine enterotoxigenic strains, but reflected the amount of LT produced by each strain. These data show that LT and cholera toxin share many common chemical and physical properties, but must be purified by different techniques.
Publisher
American Society for Microbiology
Subject
Infectious Diseases,Immunology,Microbiology,Parasitology
Reference60 articles.
1. Purification and chemical characterization of the heat-stable enterotoxin produced by porcine strains of enterotoxigenic Escherichia coli;Alderete J. F.;Infect. Immun.,1978
2. Resolution of bacterial proteins by polyacrylamide gel electrophoresis on slabs;Ames G. F.;J. Biol. Chem.,1974
3. Radial passive immune homolysis assay for detection of heatlabile enterotoxin produced by individual colonies of Escherichia coli or Vibrio cholerae;Bramucci M. G.;J. Clin. Microbiol.,1978
4. Chervenka C. H. 1973. A manual of methods for the analytical ultracentrifuge p. 39-55. Spinco Division of Beckman Instruments Inc. Palto Alto Calif.
5. Craig J. P. 1971. Cholera toxin p. 189-254. In S. Kadis T. C. Minfic and S. J. Ajl (ed.) Academic Press Inc. New York.
Cited by
105 articles.
订阅此论文施引文献
订阅此论文施引文献,注册后可以免费订阅5篇论文的施引文献,订阅后可以查看论文全部施引文献