36. BIOSYNTHESIS OF PYRIMIDINE NUCLEOTIDES 



HCO^ 



343 



5 10 10 ATP 



N.N-methenyl - FH 4 ^ » N -CHO- FH 4 * 



FH 



i*S 



FH 4 

 glycine serine 



HCHO ^_ N 5 n'°- methylene- FH 4 ■* 



OH 



-*FH 4 



a 



OH 



2 < 



R 



deoxyuridine -5- 

 phosphate 



XJ 



OH 



N > V CH 3 



R 

 thymidine -5- 

 phosphate 



Fig. 4. The thymidine-5'-phosphate synthetase reaction together with certain 

 pathways of "1-C" metabolism. 



able evidence indicated deoxyuridine-5'-phosphate to be the primary "1-C" 

 acceptor. 



Blakley 150 has similarly studied the methylation of deoxyuridine by 

 Dowex-1 -treated soluble enzymes of thymus. In this system, the formation 

 of methyl-labeled thymidine from deoxyuridine and serine-3-C 14 required 

 the addition of FH 4 , ATP, Mg++, and DPNH. Uridine, 4,5-dihydrouri- 

 dine, and 4,.~-dihydrodeoxyuridine were not involved in the methylation 

 reaction (cf. Greenberg and Humphreys 151 ). The statement that deoxyuri- 

 dine-5'-phosphate is not utilized as effectively as deoxyuridine is difficult 

 to reconcile with the observations of the other groups. 



The over-all thymidylate synthetase reaction is shown in Fig. 4 together 

 with the pathways of incorporation of methyl group precursors into the 

 thymine ring. The assignation of structure of the immediate "1-C" donor as 

 Y 5 , JV 10 -methylene-FH 4 may be tentatively made on the basis of the evidence 

 of Blakley 134 and Kisliuk. 152 Kisliuk 152 has shown that formaldehyde 

 binding by FH 4 is dependent on the presence of unsubstituted N 5 H- 

 and N 10 H-positions. Blakley 134 has indicated that the primary site of inter- 

 action of formaldehyde and FH 4 is at the N 5 H-position and that the struc- 

 ture of the adduct contains a A^iV^-methylene bridge. He has further 

 shown that the only difference between synthetic A^ 5 ,A^ 10 -methylene-FH 4 

 and the product of enzymic interaction of serine and FH 4 is in stereoisomeric 

 respects. The data do not, however, rule out the possibility of enzymic in- 

 volvement of an A^-hydroxymethyltetrahydrofolic acid in facile equilibrium 

 with the iV 5 ,iV 10 -methylene-FH 4 . The metabolic interrelationships of for- 



150 R. L. Blakley, Biochim. et Biophys. Acta 24, 224 (1957). 



151 D. M, Greenberg and (.1. K. Humphreys, Federation Proc. 17, 234 (1958). 



152 R. L. Kisliuk, J. Biol. ('hem. 227, 805 (1957). 



