1. In order to elucidate the physiological role of ADP activation for the threonine deaminase [EC 4. 2.1.16] of Clostridium tetanomorphum, metabolism of threonine and its regulation by the nucleotide were investigated. α-Ketobutyrate, a deamination product of L-threonine, was cleaved by a phosphoroclastic enzyme system to yield propionyl phosphate, CO2 and H2 in the presence of ferredoxin, CoA, and inorganic phosphate. Phosphate group in propionyl phosphate thus formed was transferred to ADP by a specific propionokinase to yield ATP and propionic acid. 2. Experiments with cell-free extracts fortified with added CoA, potassium phosphate and magnesium ions indicated that the rate of the formation of propionic acid from threonine was regulated by ADP concentration with a half saturation value of 0.14 mM. In addition, the activation of threonine deaminase by ADP was competitively abolished by ATP with a K1 value of 0.69 mM, which was almost 9 times higher than the Km for ADP under the experimental conditions employed. 3. A possible regulatory mechanism is proposed that the catabolism of threonine is linked to the generation of ATP and ADP accelerates the primary reaction, which appears to be the rate-limiting step in overall metabolism when the level of ATP decreases. ATP level is thus maintained by catabolic utilization of the substrate.