Studies on the Mode of Action of Diphtheria Toxin. 3. Site of Toxin Action in Cell-free Extracts

Extracts from HeLa cells treated with excess diphtheria toxin for several hours, until all protein synthesis has been arrested, are still able to stimulate the poly U-directed incorporation of phenylalanine into polypeptides at a moderate rate. Activity may be restored to normal levels or above by addition of a soluble enzyme fraction containing transferase II. Our results are in agreement with those of Collier who has recently shown that toxin inactivates transferase II in extracts from rabbit reticulocytes. We have further demonstrated that amino acid incorporation in extracts from intoxicated HeLa cells is limited by their transferase II content whereas, in extracts from normal cells, it is the ribosomes and to a lesser extent sRNA that are limiting. We have found that only soluble transferase II is inactivated by toxin; the ribosome-bound enzyme is resistant.