BIOSYNTHESIS OF PURINES AND PYRIMIDINES 307 



and magnesium ions were also necessary for the reaction to proceed: 



i i 1 t-\r\ I -K-TT carbamylgutamate . , . , 



L-aspartate + COo + PsHs — at? Mg^-^ * L-ureidosuccinate 



Several enzymic steps are involved and it seems that the first step — the 

 activation of CO2 and NH3 — is analogous to the first step in citrulUne syn- 

 thesis.'^* The second step involves condensation of aspartate with the acti- 

 vated intermediate and catalyzation by an enzyme different from the "con- 

 densing enzyme" in citrulline synthesis. 



Lieberman et al}^^ have purified an enzyme (ureidosuccinase) from Z. 

 oroticum which forms L-aspartate, CO2 , and NH3 from L-ureidosuccinate. 

 The enzyme requires Mn++ or Fe"^ for full activity and does not seem to be 

 identical with the one described in rat liver. 



Evidence for an alternative pathway for pyrimidine biosynthesis in 

 pigeon liver was obtained by Schulman and Badger, "° who found that the 

 labeled ureido-carbon from L-citrulline was utilized for the synthesis of 

 carbon 2 of the pyrimidine ring both in vivo and in vitro. This finding might 

 indicate that arginosuccinate is an intermediate in orotic a6id formation 

 in pigeon liver. '^"^ 



The transformation of orotate-C'^ into uracil derivatives by soluble 

 enzymes was studied independently in Romberg's laboratory and by Hurl- 

 bert and Reichard.'^' Both groups found that uridine-5'-phosphate (UMP) 

 was the first uracil derivative formed from orotic acid, ribose-5'-phosphate, 

 and ATP, and that UMP could be further degraded to uracil and uridine, 

 or phosphorylated to uridine pyrophosphate derivatives.'^' •''*' 



The mechanism of UMP formation from orotic acid has been clarified 

 through the brilliant work of Romberg and co-workers.'^' '^--'^^ With a 

 purified enzyme from pigeon liver it was shown that 5'-phosphoribosylpyro- 

 phosphate (PRPP) is formed from ribose-5'-phosphate and ATP. Orotic 

 acid condenses with PRPP to form orotidine-5'-phosphate, which is then 

 decarboxylated to UMP. These latter two reactions were demonstrated 

 with purified enzymes from yeast. Schematically the whole reaction se- 



'3* S. Grisolia in "Phosphorus Metabolism," ( McElroy and Glass, eds.), Vol. 1, 



p. 619. Johns Hopkins Press, Baltimore, 1951. 

 ''' I. Lieberman, A. Kornberg, E. S. Simms, and S. R. Kornberg, Federation Proc. 



13, 252 (1954). 

 '" M. P. Schulman and S. J. Badger, Federation Proc. 13, 292 (1954). 

 140a jYn alternative explanation might be that by a reversal of the second step in 



citrulline synthesis"' labeled Compound X (the intermediate containing "active" 



NH3 and CO2) is formed from citrulline-C'^ Compound X would then condense 



with aspartate to form labeled ureidosuccinate. 

 '^» R. B. Hurlbert and P. Reichard, Acta Chem. Scand. 8, 701 (1954). 

 '^2 A. Kornberg, I. Lieberman, and E. S. Simms, /. Am. Chem. Soc. 76, 2027 (1954). 

 1" L Lieberman, A. Kornberg, and E. S. Simms, /. Am. Chem. Soc. 76, 2844 (1954). 



