Isolation and characterization of functional Shiga toxin subunits and renatured holotoxin.

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.