36. BIOSYNTHESIS OF PYRIMIDINE NUCLEOTIDES 347 



interest to note that back-mutation to thymine-independence of the thy- 

 mine-less mutant, E. coli 15T-, does not involve a loss of ability to utilize 

 the free pyrimidine or the deoxy nucleoside. 168 



The pathway of utilization of thymidine has been elucidated by the ob- 

 servation of kinase activities in cell-free preparations of mamamlian 95 - 169 ' 17 ° 

 and bacterial 94 origin which effect the following transphosphorylations: 



Thymidine ATP > TMP ATP > TDP ATP > TTP 



The occurrence of TDP and TTP in extracts of thymus tissue has been 

 reported 93 and the involvement of TTP in DNA synthesis in soluble in 

 vitro enzyme systems has also been described. 94, 104, 169, 17 ° 



The coupling of thymidine phosphorylase with thymidine kinase offers 

 a pathway of thymine utilization. 



V. Alternate Pathways of Pyrimidine Nucleotide Synthesis 



Evidence pertaining to the operation of alternative pathways of pyrimi- 

 dine nucleotide synthesis has been accumulating in recent years. Consider- 

 ation has been given to the possible reversal of the reaction sequence 52 " 56 

 which effects the degradation of uracil to ^-alanine via dihydrouracil and 

 carbamyl-/3-alanine (jS-ureidopropionic acid). The enzymes involved in this 

 pathway have been extensively investigated. 52 ' m Fritzson 55 has indicated, 

 however, that the only effectively reversible step in rat liver slices is the 

 intercon version of dihydrouracil and carbamyl-0-alanine. As uracil is uti- 

 lized only to a negligible extent in rat liver for polynucleotide synthesis it 

 would be of interest to study the anabolic potentialities of the degradation 

 pathway in a tissue capable of more extensive uracil utilization. It should 

 be noted in this connection that dihydrouracil and dihydrocytosine (and 

 the corresponding ribonucleosides and deoxyribonucleosides) do not sup- 

 port the growth of a variety of pyrimidine-less mutants of E. coli} bf> Lagerk- 

 vist et al. m have also shown that N 15 -labeled dihydrouracil, carbamyl-/3- 

 alanine, and 0-alanine, unlike uracil, are not utilized in a significant manner 

 for polynucleotide pyrimidine synthesis in the Ehrlich ascites cell. Heidel- 

 berger et al.f 1 on the other hand, have reported that dihydrouracil-2-C 14 is 

 incorporated to a small extent into acid-soluble nucleotides by a high-speed 

 supernatant fraction of homogenates of rat liver or of the Flexner-Jobling 

 carcinoma. The extent of utilization of the dihydropyrimidine was slightly 

 greater than that of uracil. The significance of these observations is not 

 clear. The possibility that the observed incorporation of dihydrouracil-2- 



168 L. V. Crawford, Biochim. et Biophys. Acta 30, 428 (1958). 



169 F. J. Bollum, Federation Proc. 17, 193 (1958). 



170 F. J. Bollum and V. R. Potter, J. Biol. Chem. 233, 478 (1958). 



171 L. L. Campbell, Jr., J. Biol. Chem. 227, 693 (1957). 



172 See ref. 36a, page 328. 



