Isolation and characterization of functional Shiga toxin subunits and renatured holotoxin

• Published in: Molecular microbiology Vol. 3; no. 9; p. 1231
• Main Authors: Donohue-Rolfe, A, Jacewicz, M, Keusch, G T
• Format: Journal Article
• Language: English
• Published: England 01.09.1989
• Subjects: Bacterial Toxins - isolation & purification; Bacterial Toxins - toxicity; Binding, Competitive; Cross-Linking Reagents; Cytotoxins - isolation & purification; HeLa Cells; Humans; Imidoesters; Peptide Fragments - isolation & purification; Peptide Fragments - toxicity; Protein Biosynthesis - drug effects; Protein Conformation; Shiga Toxins; Shigella dysenteriae - analysis
• ISSN: 0950-382X
• DOI: 10.1111/j.1365-2958.1989.tb00273.x

Shiga toxin is a protein toxin produced by Shigella dysenteriae type I strains. In this report we present a procedure for the separation of functionally intact toxin A and B chains and for their reconstitution to form biologically active molecules. In agreement with the findings of others, the isolated A chain was shown to be a potent in vitro inhibitor of eukaryotic protein synthesis. The isolated B chain bound to HeLa cells and competitively inhibited the binding and cytotoxic activity of holotoxin. These findings show that the functional role of the B chain is to recognize cell surface functional receptors. By labelling the B subunit alone, prior to renaturation of holotoxin, the polypeptide chains were shown to associate noncovalently with a stoichiometry of one A chain and five B chains.