Phyllis F. Cohen and Roberta F. Colman* abstract: This paper explores the hypothesis that only one of the ionic forms of isocitrate participates in the metal-dependent reaction catalyzed by DPN-specific isocitrate dehydrogenase and suggests that activators can function indirectly by modifying the distribution of the various forms of isocitrate. Determinations of the Michaelis constants for DPN and total isocitrate at 1.25 dim MnS04 showed that the Km for DPN was relatively constant, but that the Km for total isocitrate increased nearly 100-fold as the pH was raised from 6.0 to 8.0. Higher concentrations of MnS04 increased the Knl for total isocitrate at each pH. An analysis of the distribution of the various forms of isocitrate, facilitated by a computer pro-gram, revealed that the Km for dibasic isocitrate (approximately 30.0 gM) was independent of both pH and manganese concentration. The observed variation in the Km for total iso-citrate can therefore be explained as a reflection of differential modifications of the equilibria between the ionic forms of iso-citrate caused by changes in the concentrations of hydrogen and manganous ions. Free dibasic isocitrate is the actual sub-