Purification and sequence determination of heat‐stable enterotoxin elaborated by a cholera toxin‐producing strain of Vibrio cholerae O1

Abstract

Four molecular species of heat-stable enterotoxins elaborated by a cholera toxin-producing strain of Vibrio cholerae O1 were isolated from its culture supernatant. The amino acid sequence of one of the enterotoxins was determined to be Phe-Ile-Lys-Gln-Val-Asp-Glu-Asn-Gly-Asn-Leu-Ile-Asp-Cys-Cys-Glu-Ile-Cys- Cys-Asn-Pro-Ala-Cys-Phe-Gly-Cys-Leu-Asn with three intramolecular disulfide linkages. The other enterotoxins had shorter amino acid sequences in the N-terminal regions, but possessed the same sequence in their C-terminal regions including the three disulfide linkages. The enterotoxins with the shorter N-terminal sequences showed more potent toxicities, and the minimum effective dose of the longest one with 28 amino acid residues was 10-folds of that of the shortest one.

Amino acid sequence of a novel heat-stable enterotoxin produced by a yst gene-negative strain of Yersinia enterocolitica

“…This observation indicates that A-2 is the C-terminal fragment in Y-STb, but A-1 is not [13]. The amino acid sequence of A-1 was identical to that of a part (4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14) of the N-terminal amino acid sequence of the purified Y-STb, which is consistent with the data from FAB-MS and amino acid analysis ( Fig. 3 and Table 1).…”

“…An aliquot of each fraction was tested for toxicity in suckling mice and the active fractions were pooled. The fractions containing toxic materials were concentrated, dialyzed against distilled water, and then subjected to immunoaffinity chromatography using a monoclonal antibody (MAb), 2F, under conditions described previously [9]. The toxic fractions obtained by immunoaffinity chromatography were purified further by reversed-phase highperformance liquid chromatography (RP-HPLC) (see legend of Fig.…”

Characterization of a highly toxic, large molecular size heat‐stable enterotoxin produced by a clinical isolate of Yersinia enterocolitica

“…The MED of T-2 was nearly identical to that of T-l, indicating that the conversion of Nterminal Gln to pGlu does not affect the enterotoxic activity. In ST molecules produced by V. cholerae O1, the molecules with the shorter N-terminal sequences showed more potent toxicities, and the MED of the largest one with 28 amino acid residues was ten times more than that of the smallest one with 17 amino acid residues [8]. In contrast, Y-STc was the most toxic in the ST family despite having the longest sequence.…”

“…3). The length of the secreted mature polypeptides ranges from only 17 amino acid residues for NAG-ST and O1-ST produced by E cholerae [6,8] sequences of Y-STc and the pro-form of Y-STa revealed that there is a sequence similarity of 49% (26 residues/53 residues) (Fig. 3).…”

“…Heat-stable enterotoxins (STs) are peptide toxins produced by bacterial enteropathogens such as enterotoxigenic Escherichia coli [1,2], Yersinia enterocolitica [3][4][5] and Vibrio cholerae [6][7][8], and are responsible for acute diarrhea in humans. ST binds to a membrane-bound guanylate cyclase which serve as a specific cell surface receptor for ST, followed by elevation of the intracellular cGMP level [9][10][11].…”