Abstract The effects of the guanine analog, 6-thioguanine, have been evaluated in exponentially growing cultures of Bacillus cereus . The drug produced growth inhibition instantly, the duration but not the degree of which was related to the inhibitor concentration. Guanine prevented completely thioguanine-induced growth inhibition, in a competitive manner, whereas adenine and hypoxanthine were incapable of abolishing the inhibitory actions. Although thioguanine was anabolized into the ribomononucleotide, the quantity of drug incorporated into nucleic acids was extremely small. It is unlikely that this incorporation was responsible for growth inhibition. The drug was readily desulfurated and the sulfur portion incorporated into proteins. Thioguanine usually depressed the formation of protein and DNA in accordance with the decreased formation of cell mass, but RNA synthesis was decreased by a greater extent. Uracil incorporation into RNA pyrimidines was more specifically depressed than was RNA synthesis, whereas the conversion of orotic acid into RNA pyrimidines was enhanced during growth with thioguanine. The incorporation of amino acids into proteins was not specifically affected by drug treatment, but considerable fluctuations in replicate experiments were observed. The induction of penicillinase was unaltered by the analog. Bacterial flagella formation, on the other hand, was largely abolished, and the conversion of diaminopimelic acid into cell wall was decreased. In comparison experiments with the structurally closely related drug 6-mercaptopurine, that analog produced very similar biochemical actions as did thioguanine on nucleic acid and protein biosynthesis. The two drugs differed markedly, however, in the ability of purines to antagonize their growth-inhibitory properties. Other dissimilarities between the drugs included a lack of inhibitory effect by mercaptopurine (but not thioguanine) on guanine incorporation and a lesser enhancement of orotic acid incorporation into RNA pyrimidines in the presence of mercaptopurine. It is postulated that thioguanine and mercaptopurine produce a selectively damaging effect on the nucleic acids of B. cereus , perhaps on DNA, which leads to reduction of RNA synthesis and impairment of the formation of specific proteins.