Abstract Rat liver NAD kinase (ATP:NAD 2'-phosphotransferase, EC 2.7.1.23) was purified about 70-fold. The Michaelis-Menten constants (Km) for NAD and ATP were 8 x 10-4 m and 2 x 10-3 m, respectively. NAD kinase activity was markedly inhibited by NADH and also NADPH. The Ki of NADH was approximately 1 x 10-4 m, and that of NADPH was approximately 5 x 10-5 m. Both inhibitions were competitive with NAD, and the inhibition of NADH and NADPH was partially additive. NADP had little or no inhibitory effect on the enzyme activity. p-Hydroxymercuribenzoate (2 x 10-5 m) inhibited NAD kinase activity. This inhibition was reversed by 1-cysteine or 2-mercaptoethanol. The possible role of the reduced pyridine nucleotides in the control of NADP synthesis has been considered.