Preparation and properties of chimeric toxins prepared from the constituent polypeptides of diphtheria toxin and ricin. Evidence for entry of ricin A-chain via the diphtheria toxin pathway.
A highly toxic conjugate of ricin A-chain and diphtheria toxin fragment B was prepared by disulfide exchange reaction. A similar conjugate between diphtheria toxin fragment A and ricin B-chain was nontoxic. Like native diphtheria toxin, the conjugate ricin A/diphtheria toxin B was much more toxic to Vero than to HeLa cells. Ricin was equally toxic to these cell lines. Lactose, which inhibits ricin binding, did not protect against the conjugate. Cells resistant to ricin, partly due to a reduced number of ricin-binding sites, were fully sensitive to the conjugate, indicating that the conjugate binds to diphtheria toxin receptors. The conjugate was fully toxic to two Vero cell mutants, resistant to diphtheria toxin because the elongation factor 2 could not be ADP-ribosylated by the diphtheria toxin A-fragment. Therefore, the inhibition of protein synthesis by the conjugate must be caused by the ricin A-chain. Ammonium chloride which prevents entry of diphtheria toxin, but not of ricin, also protected against the conjugate. Like diphtheria toxin, the conjugate was most toxic at low pH, whereas ricin is most active at pH above neutrality and inactive at low pH. The results indicate that the conjugate ricin A/diphtheria toxin B binds to diphtheria toxin receptors and inhibits cellular protein synthesis due to the action of ricin A-chain which appears to enter the cell by the diphtheria toxin pathway.
D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002522:Chlorocebus aethiops; D004167:Diphtheria Toxin / Q000494:pharmacology*; D004220:Disulfides; D004351:Drug Resistance; D006367:HeLa Cells / Q000187:drug effects; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007668:Kidney; D055503:Protein Multimerization; D012276:Ricin / Q000494:pharmacology*; D013329:Structure-Activity Relationship
D004167:Diphtheria Toxin; D004220:Disulfides; D000643:Ammonium Chloride; D012276:Ricin