title: Protein reagent modification of cholera toxin: characterization of effects on antigenic, receptor-binding and toxic properties.
abstract: The effects of protein modification procedures on the biologically most important properties of cholera toxin, i.e. the toxic activity, the GM1 receptor-binding capacity and the antigenic (antibody-fixing) properties, have been studied quantitatively using microgram amounts or less of toxin protein. Most of the 24 group-specific reagents used had either no inhibitory effect on the toxic or the combination of GM1-binding and antibody-fixing properties of cholera toxin, or they had a concomitant inhibitory effect on these activities. Separate testing of GM1- and antibody-binding revealed a close, but not absolute, structural association between these properties, Amino group reactive substances were particularly effective in decreasing the GM1-binding activity, while leucine aminopeptidase had no effect. This suggests that lysine residues may be involved in binding toxin to the acidic GM1 receptor. Sodium dodecylsulphate and mercaptoethanol, which caused dissociation of the subunits of cholera toxin as indicated by polyacrylamide gel electrophoresis, abolished toxicity without inhibiting the concomitant GM1- and antibody-binding properties of the toxin. Similar differential effects were also obtained with three reagents which did not seem to change the aggregation state of the toxin. These substances all had specificity for arginine, suggesting that arginyl residues of the toxin molecule may be involved in a 'toxic site' distinct from the receptor-binding site(s). A selective effect on the toxic site was also found by treating the toxin with carboxypeptidase or trypsin in the presence of urea; in the absence of urea no enzymic effect on any toxin property was noted.
mesh_terms: D000818:Animals; D000937:Antigen-Antibody Reactions / Q000187:drug effects; D000992:Antitoxins; D005732:Gangliosides / Q000378:metabolism; D006801:Humans; D007202:Indicators and Reagents / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D011817:Rabbits; D012882:Skin Tests; D014118:Toxins, Biological / Q000378:metabolism / Q000633:toxicity; D014734:Vibrio cholerae
chemical_list: D000992:Antitoxins; D005732:Gangliosides; D007202:Indicators and Reagents; D014118:Toxins, Biological