474 R. E. HANDSCHUMACHER AND A. I). WELCH 



inhibition caused by azaserine not only was partially nullified by the pres- 

 ence of excess glutamine, but also led to the accumulation of sufficient gly- 

 cinamide ribonucleotide and its iV^formyl derivative for rigorous charac- 

 terization studies. 133 Confirmation of these findings was rapid 134 and further 

 work with E. coli indicated that preformed purines or aminoimidazolecar- 

 boxamide, as well as the amino acids mentioned above, could prevent the 

 inhibitory action of azaserine. 135 The fact that these antagonists did not 

 inhibit the incorporation of formate into the thymine of DNA, and of pre- 

 formed adenine into liver, 136 intestine, and sarcoma- 180 in mice (indeed, 

 stimulation of the utilization of preformed purines has been observed) is 

 indicative of the specificity of its action. 137 In fact, profound blockade of 

 the synthesis of purines de novo by E. coli was obtained with doses which 

 were only one-tenth those needed to disturb amino acid metabolism. 138 It 

 has been reported that azaserine caused a marked increase in the size of 

 the glutamine pool and a decrease in other free amino acids in Scenedesmus 

 and Chlorella cells. 139 Presumably, this represents a blockade in many of 

 the transamination reactions upon which amino acid formation depends; 

 however, the concentrations of azaserine required were far greater than 

 those which inhibited the synthesis de novo of purines. Accumulation of 

 glutamine in the acid-soluble fraction of E. coli inhibited by azaserine also 

 has been reported. 140 



More detailed studies 141 of the nature of the interaction between azaserine 

 and the enzyme involved in the transfer of amino groups from glutamine 

 to formylglycinamide ribonucleotide have revealed competitive kinetics 

 over short periods when azaserine was added simultaneously. Under these 

 same conditions DON had an affinity for this enzyme which was approxi- 

 mately forty times greater than that of azaserine. If, however, the enzyme 

 was preincubated with azaserine or DON, an irreversible inactivation oc- 

 curred. Similarly, in the presence of excess formylglycinamide ribonucleo- 

 tide, the reaction kinetics would change from competitive to noncompeti- 



133 S. C. Hartman, B. Levenberg, and J. M. Buchanan, J. Biol. Chem. 221, 1057 (1956). 



134 A. J. Tomisek, H. J. Kelly, and H. E. Skipper, Arch. Biochem. Biophys. 64, 437 

 (1956). 



135 L. L. Bennett, Jr., F. M. Schabel, Jr., and H. E. Skipper, Arch. Biochem. Biophys. 

 64, 423 (1956). 



136 R. K. Barclay, E. Garfinkel, and M. Phillips, Proc. Am. Assoc. Cancer Research 

 2, 93 (1956). 



137 J. F. Fernandes, G. A. LePage, and A. Lindner, Cancer Research 16, 154 (1956). 



138 A. J. Tomisek, H. J. Kelly, and H. E. Skipper, Abstr. 128th Meeting Am. Chem. 

 Soc, Minneapolis, Minn. p. 5C (1955). 



139 S. A. Barker, J. A. Bassham, M. Calvin, and U. C. Quarck, J. Am. Chem. Soc. 78, 

 4632 (1956). 



140 A. J. Tomisek, M. R. Reid, and H. E. Skipper, Cancer Research 19, 489 (1959). 



141 B. Levenberg, I. Melnick, and J. M. Buchanan, J. Biol. Chem. 225, 163 (1957). 



