Purification and Chemical Characterization of the Heat-Labile Enterotoxin Produced by Enterotoxigenic
            <i>Escherichia coli</i>

Kunkel, Steven L.; Robertson, D C

doi:10.1128/iai.25.2.586-596.1979

Cited by 104 publications

(34 citation statements)

References 57 publications

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“…Although LT and CT share some common biological activities, immunological determinants, molecular structure, and enzymatic activities (4,10,17,24,28,35), their antigenic and amino acid compositions are clearly distinguishable (4,5,8,34). Moreover, immunological and molecular heterogeneity of LTs of human and porcine origin have recently been reported (14,21,37).…”

Section: Discussionmentioning

confidence: 99%

Purification and some properties of a non-o1 Vibrio cholerae enterotoxin that is identical to cholera enterotoxin

Yamamoto¹,

Takeda²,

Miwatani³

et al. 1983

Infect Immun

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Cholera-like enterotoxin was isolated and purified from the culture supernatant of a non-O1 strain of Vibrio cholerae, E8498, isolated from the environment. Enterotoxin was purified by aluminum hydroxide absorption and elution and successive gel filtrations on Sephadex G-100, Bio-Gel A-5m, and Sephadex G-75. Purified enterotoxin gave a single stained band on polyacrylamide gel disc electrophoresis, and the mobility was the same as that of cholera enterotoxin. The specific biological activity of the purified enterotoxin was almost the same as that of cholera enterotoxin in the Chinese hamster ovary cell assay, fluid accumulation in mouse ligated intestine, increase in vascular permeability in rabbit skin, and passive immune hemolysis. Sodium dodecyl sulfate-polyacrylamide slab gel electrophoresis showed that the purified enterotoxin consisted of subunits A and B, identical to those of cholera enterotoxin, and Ouchterlony double gel diffusion tests indicated that the two toxins were immunologically identical. Enterotoxins prepared from several non-O1 strains isolated from human patients were also immunologically identical to cholera enterotoxin.Sodium azide and sodium EDTA were added to culture supernatants at final concentrations of 0.02 and 0.05%, respectively.Purified CT. CT from V. cholerae 569B, serovar 01, purified by the method described by Ohtomo et al. 1128 on July 15, 2020 by guest http://iai.asm.org/ Downloaded from

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Section: Discussionmentioning

confidence: 99%

Purification and some properties of a non-o1 Vibrio cholerae enterotoxin that is identical to cholera enterotoxin

Yamamoto¹,

Takeda²,

Miwatani³

et al. 1983

Infect Immun

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“…LT, obtained from E. co/i strain 41 1-5, cultivated as previously described , was dissolved in PBS to 1125 BD-units/ml (BD-units according to Craig (1965)). The washed bacteria were sonicated at 22 kHz for 2 min., extracted with PBS and fractionated by fractionated precipitation with ammonium sulfate (Kunkel & Robertson 1979).…”

Section: Methodsmentioning

confidence: 99%

Reversal of Enterotoxic Diarrhoea by Anaesthetic and Membrane‐stabilizing Agents

Lönnroth

Jennische

1982

Acta Pharmacologica et Toxicologica

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The effect of membrane‐stabilizing and sedative drugs on enterotoxic diarrhoea was studied in mice. Fluid secretion was induced in ligated loops of the small intestine by challenge with cholera toxin (CT), heatlabile enterotoxin (LT) from Escherichia coli or dibuturyl‐cyclic AMP (dB‐cAMP). Chlorpromazine, melperone, diazepam, mebumal, ketamine and ethanol all inhibited CT‐induced hypersecretion, ED50 being 1.5, 4, 4, 35, 70 and 1500 mg/kg, respectively. The drugs also blocked CT‐stimulation of adenylate cyclase, which mediates the action of CT. Concomitant with the antisecretory effect a sedative effect was induced, as judged by the motility and righting reflex of the animals. Hypersecretion by E. coli and LT was also totally blocked by chlorpromazine, diazepam, ketamine and ethanol. In contrast, the secretion by dB‐cAMP which bypasses adenylate cyclase in its action, was not affected by diazepam and was only partly reversed by chlorpromazine, ketamine and ethanol. The reversal of dB‐cAMP‐secretion is probably due to enhanced absorption, rather than inhibition of adenylate‐cyclase. The use of membrane‐stabilizing drugs may represent a new principle for pharmacological treatment of diarrhoea.

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“…CT is closely related to the heat-labile enterotoxin (LT) of Escherichia coli (34). The mode of action of LT is similar to that of CT, and LT has been implicated in the pathogenesis of secretory diarrhea in humans and in animals (13).Both CT and LT have been purified to homogeneity (3,9,18,22), and the structural genes for both toxins have been cloned (30,35,37). Each toxin is composed of two different polypeptide subunits designated A and B.…”

mentioning

confidence: 99%

“…Both CT and LT have been purified to homogeneity (3,9,18,22), and the structural genes for both toxins have been cloned (30,35,37). Each toxin is composed of two different polypeptide subunits designated A and B.…”

mentioning

confidence: 99%

Characterization of monoclonal antibodies that react with unique and cross-reacting determinants of cholera enterotoxin and its subunits

Holmes¹,

Twiddy²

1983

Infect Immun

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Seventeen selected hybridoma cell lines that produced monoclonal antibodies against cholera enterotoxin (CT) were isolated and characterized. All of the monoclonal antibodies contained the kappa light chain; 14 were of the immunoglobulin Gl (IgGl) isotype and 3 were IgG2a. The 17 monoclonal antibodies were divided into a minimum of seven different specificity groups based on their abilities to bind to the following purified test antigens in solid-phase radioimmunoassays: CT, the A and B polypeptides of CT (CT-A and CT-B, respectively), and the heat-labile enterotoxins designated LTh and LTp from Escherichia coli. The binding of these antibodies to the following subunits and fragments of CT was also determined in Western blots: pentameric CT-B, monomeric CT-B, intact CT-A, and the Al fragment of CT-A. Each of the monoclonal antibodies was tested for neutralization of CT and for precipitation with CT in immunodiffusion tests. Antigenic determinants were identified on CT that were not present either on CT-A or CT-B. One class was unique for CT and another was shared with LTh and LTp. Antibodies directed against these holotoxin-specific determinants had no neutralizing activity. Most of the monoclonal antibodies that reacted strongly with CT-A or CT-B also reacted strongly with CT holotoxin; however, one class of antibody reacted strongly with CT-A but weakly with CT. Among the monoclonal antibodies against CT-A or CT-B, some were specific for CT and others crossreacted with LTh and LTp or with LTh only. The most potent neutralizing antibodies were against CT-B, and all of our monoclonal antibodies against CT-B had some neutralizing activity. In contrast, only some of the monoclonal antibodies against CT-A had neutralizing activity, and their specific activities were low. We found no direct correlations between the ability of monoclonal antibodies to neutralize CT and to cross-react with LTh or LTp. None of the epitopes recognized by our monoclonal anti-CT antibodies was present on CT-A and CT-B.Cholera enterotoxin (CT) is a heat-labile enterotoxin produced by Vibrio cholerae (9). It has an essential role in the pathogenesis of cholera and causes secretory diarrhea by activating adenylate cyclase in the mucosal epithelium of the small intestine (13). CT is closely related to the heat-labile enterotoxin (LT) of Escherichia coli (34). The mode of action of LT is similar to that of CT, and LT has been implicated in the pathogenesis of secretory diarrhea in humans and in animals (13).Both CT and LT have been purified to homogeneity (3,9,18,22), and the structural genes for both toxins have been cloned (30,35,37). Each toxin is composed of two different polypeptide subunits designated A and B. Each holotoxin contains one A polypeptide and five copies of the B polypeptide held together by noncovalent bonds (14,15). Comparisons of primary structures demonstrated that the A and B polypeptides from CT have extensive homology with those from LT (6,36). Hyperimmune antisera prepared against either of the purified toxins will ne...

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Purification and Chemical Characterization of the Heat-Labile Enterotoxin Produced by Enterotoxigenic Escherichia coli

Cited by 104 publications

References 57 publications

Purification and some properties of a non-o1 Vibrio cholerae enterotoxin that is identical to cholera enterotoxin

Purification and some properties of a non-o1 Vibrio cholerae enterotoxin that is identical to cholera enterotoxin

Reversal of Enterotoxic Diarrhoea by Anaesthetic and Membrane‐stabilizing Agents

Characterization of monoclonal antibodies that react with unique and cross-reacting determinants of cholera enterotoxin and its subunits

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