STUDIES ON THE MODE OF ACTION OF DIPHTHERIA TOXIN

Abstract

When diphtheria toxin and NAD are added to soluble fractions containing aminoacyl transfer enzymes isolated from rabbit reticulocytes or from HeLa cells, free nicotinamide is released and, simultaneously, an inactive ADP ribose derivative of transferase II is formed. The reaction is reversible, and in the presence of excess nicotinamide, toxin catalyzes the restoration of aminoacyl transfer activity in intoxicated preparations. In living cultures of HeLa cells, the internal NAD concentration is sufficiently high to account for the rapid conversion, catalyzed by a few toxin molecules located in the cell membrane, of the entire cell content of free transferase II to its inactive ADP ribose derivative. Completely inactive ammonium sulfate fractions containing soluble proteins isolated from cells that have been exposed for several hours to excess toxin, can be reactivated to full aminoacyl transfer activity by addition of nicotinamide together with diphtheria toxin. Transferase II appears to be a highly specific substrate for the toxin-stimulated splitting of NAD and thus far no other protein acceptor for the ADP ribose moiety has been found.