314 JOHN M. BUCHANAN 



a better inhibitor than azaserine by a factor of 40. When either inhibitor 

 is incubated with the enzyme in the absence of glutamine there is rapid 

 formation of an inhibitor-enzyme complex which does not dissociate even 

 upon addition of glutamine. The enzyme is thus irreversibly inactivated. 

 Recently radioactive azaserine has been prepared and has been found to 

 react with the purified enzyme in approximately stoichiometric amounts. 

 Because of the combination of azaserine with the enzyme even in the pres- 

 ence of glutamine the inhibition is partly competitive and partly noncom- 

 petitive in nature. 



Formylglycinamidine ribonucleotide is converted enzymically into the 

 first imidazole intermediate by reaction with ATP according to Eq. (7). 47 ' 50 

 The enzyme responsible for this reaction has been isolated from chicken 



Formvlglvcinamidine ribonucleotide + ATP — » 



(7) 

 5-aminoimidazole ribonucleotide + ADP + orthophosphate 



liver and purified with some difficulty due to its relative instability. The 

 activity of the enzyme during incubation shows a pronounced requirement 

 for potassium ions. Likewise, the enzyme is able to withstand dialysis and 

 prolonged storage only in solutions of potassium salts. The enzyme cata- 

 lyzes an irreversible reaction in which the energy for the closure of the 

 imidazole ring is obtained through the hydrolysis of an anhydride bond of 

 ATP with ADP and orthophosphate as products. 5-Aminoimidazole ri- 

 bonucleotide may be isolated by a procedure similar to that used for the 

 isolation of the other intermediates of these reactions. The ribonucleotide 

 is isolated by chromatography on an anion exchange column and is then 

 precipitated as the barium salt at neutral pH. 5-Aminoimidazole ribonu- 

 cleotide reacts with the Bratton and Marshall reagents to yield an orange- 

 colored product with an absorption maximum at 500 mp. The ribonucleotide 

 itself has only a nonspecific end absorption in the ultraviolet region. The 

 ribonucleotide is destroyed by heating at 100° in acid solution for a short 

 time. The relative instability of the intermediate under even moderate 

 conditions has made it difficult to isolate in pure form. 



5-Aminoimidazole reacts with C0 2 (or HC0 3 ~) in the next step 51 of this 

 reaction sequence to yield 5-amino-4-imidazolecarboxylic acid ribonucleo- 

 tide [Eq. (8)]. This compound exhibits a specific absorption maximum at 

 249 rn.fi, a property which has provided an excellent means for its determin- 

 ation. 



5-Aminoimidazole ribonucleotide + CO2 *=* 



5-amino-4-imidazolecarboxylic acid ribonucleotide 



50 B. Levenberg and J. M. Buchanan, /. Biol. Chem. 224, 1005 (1957). 

 61 L. N. Lukens and J. M. Buchanan, J. Am. Chem. Soc. 79, 1511 (1957). 



