A histidine auxotrophic (hisA) mutant of Kiebsiella pneumoniae is phenotypically Nif-when grown with 20,ug of histidine ml-' but Nif+ when supplied with histidine at 100 Fg ml-'. Reversion to Nif' at 20 ,ug of histidine ml' occurs phenotypically by the addition Qf 2-thiazolyl-DL-alanine or genetically by mutation in hisG; 2-thiazolyl-DL-alanine inhibits and hisG encodes phosphoribosyl phosphotransferase, the first enzyme of the histidine biosynthetic pathway which consumes ATP. Physiological studies of the hisA mutant JS85 showed that after removal of NH4+ from a culture of the mutant grown with 20 ,g of histidine ml-, synthesis of nitrogenase polypeptides occurred at a rate similar to that in the wild type for about 3 h and acetylene reduction activity reached about 10% of the fully derepressed wild-type level. Shortly thereafter the concentration of intracellular adenylates decreased; in particular, ATP fell to about 10% of normal levels. Also, nitrogenase proteins (nifHDK products) and the nifJ gene product stopped being synthesized. These effects were not due to impairment of growth or protein synthesis by histidine starvation. Inhibition of phosphoribosyl phosphotransferase with 2-thiazolyl-DL-alanine restored nitrogenase activity and synthesis, indicating that the effect of the hisA mutation on nif expression was probably a consequence of lowered energy resources that occurred during anaerobic N starvation. Nif+ when supplied with 100 ,ug of histidine ml-' (7). Other histidine auxotrophs with mutations in hisC, hisB, hisD, hisF, or hisG were Nif+ even in low-histidine medium, although nitrogenase activity was slightly reduced in some of them. These results indicated that histidine starvation per se did not cause the Nif-phenotype in the hisA mutants. In liquid cultures of one hisA mutant, JS85, nitrogenase activity was never more than 10% of wild-type levels when histidine was supplied at 20 jxg ml-l but was 100% when histidine was at 100 pLg ml-. ATP levels in JS85 decreased to 10% of that of His' strains during derepression and appeared to correlate with a cessation of nitrogenase synthesis.The reversal of JS85 to Nif+ by the addition of adenine or 2-thiazolyl-DL-alanine (2TA) to low-histidine, N-free agar medium also indicated that histidine starvation was not the cause of the Nif phenotype (7). 2TA is an analog of histidine that is not incorporated into proteins and has no effect on protein synthesis but inhibits the activity of the first enzyme of the histidine biosynthetic pathway. This is phosphoribosyl phosphotransferase, an ATP-consuming enzyme (Fig. 1). The hisA mutation prevents formation of the biosynthetic intermediate 5-'aminoimidazole-4-carboxamide-1-ribonucleotide, which normally restores the purine precursor pool. Thus, physiological evidence indicated that a loss of ATP or a consequent energy imbalance correlated with the Nif-phenotype of JS85.This hisA mutant, JS85, has now been further characterized by examining Nif* revertants, by measuring adenylate levels and rates of nitrogenase syn...