Modification of the glycolipid-binding specificity of vero cytotoxin by polymyxin B and other cyclic amphipathic peptides

Abstract

Polymyxin B, an amphipathic cycle decapeptide produced by Bacillus polymyxa, is routinely used in the extraction of the components from the periplasmic space of germ-negative bacteria. Vero cytotoxin 1 (VT1) is an Escherichia coli-elaborated subunit toxin which binds to the glycolipid globotriosylceramide (Gal-.alpha.1-4-Gal .beta.1-4-Glc-ceramide [Gb3]) and has been strongly implicated in the etiology of the hemolytic uremic syndrome and hemorrhagic colitis. We now shown by in vitro glycolipid-binding assays that in the presence of low concentrations of polymyxin B, globotetraosylceramide (GalNAc.beta.1-3Gal.alpha.1-4Gal.beta.1-4Glc-ceramide [Gb4]) is also recognized by both the VT1 B (binding) subunit and holotoxin. Melittin, a 26-amino-acid cyclic peptide of similar amphipathic nature, produced the same effect, whereas a hydrophobic blocking agent did not. Triton X-100 did not increased binding of VT1 to Gb4 but prevented glycolipid binding in toto at concentrations above 0.5%. Caution is therefore advised in the analysis of VT1 glycolipid binding in the presence of amphipathic peptides.