One Molecule of Diphtheria Toxin Fragment A Introduced Into a Cell Can Kill the Cell

Cell. 1978 Sep;15(1):245-50. doi: 10.1016/0092-8674(78)90099-5.

Abstract

Erythrocyte ghosts containing a known number of molecules of purified fragment A of diphtheria toxin with a constant amount of FITC-BSA as a fluorescence marker were prepared by dialyzing a mixture of erythrocytes and these substances against hypotonic solution. These substances were then introduced into diphtheria toxin-resistant mouse L cells by virus-mediated cell fusion of the cells with the ghosts, and mononuclear recipients that has fused with only one erythrocyte ghost were separated in a flourescence-activated cell sorter (FACS) on the basis of their cell size and fluorescence intensity. After separation, the viability of cells containing known numbers of fragment A was examined by measuring colony-forming ability. The results demonstrated that a single molecule of fragment A was sufficient to kill a cell. This fact was confirmed by introduction into cells of fragment A from an immunologically related mutant toxin, CRM 176 (fragment A176); this has a completely functional fragment B region, but in cell extracts, the enzymic activity of its fragment A is about 10 fold less than that of wild toxin. The cytotoxicity of CRM 176 is about two hundredths of that of the wild-type (Uchida, Pappenheimer and Greany, 1973). As expected, about 100-200 fold excess of fragment A-176 was needed to kill the cells.

MeSH terms

  • Cell Survival / drug effects*
  • Diphtheria Toxin / administration & dosage
  • Diphtheria Toxin / toxicity*
  • Dose-Response Relationship, Drug
  • Erythrocyte Membrane
  • L Cells
  • Peptide Elongation Factors
  • Peptide Fragments / toxicity

Substances

  • Diphtheria Toxin
  • Peptide Elongation Factors
  • Peptide Fragments