Glutathione is synthesized from gamma-glutamylcysteine and glycine via the action of glutathione synthetase. It is known that gamma-glutamylcyclotransferase is present in many cells and may convert gamma-glutamylcysteine to 5-oxoproline and cysteine, but until now there has not been a credible explanation for the apparent suppression of the gamma-glutamylcyclotransferase reaction during glutathione synthesis. Our data suggest that the gamma-glutamylcyclotransferase and glutathione synthetase pathways are regulated by a simple kinetic mechanism that favors the synthesis of glutathione.