I06 INTERMEDIARY METABOLISM AND GROWTH I 



-^^P-thymidylate is also incorporated into the polynucleotide (Kornberg et al., 



1956a, b). 



ATP ATP ATP 



Thymidine > thymidylate > thymidine triphosphate > DNA 



Polymerization of thymidine triphosphate (TTP) requires ATP, a heat stable 

 DNA fragment, provisionally regarded as a primer, and two enzyme fractions 

 which have been purified more than 100 fold. TDP can replace TTP in the reac- 

 tion but a decision as to the more immediate precursor requires further purifica- 

 tion of the system (Kornberg ^irt/., 1956b). The polyphosphates of adenine, cytosine 

 and guanine deoxyribosides are also used for polynucleotide synthesis by the 

 crude enzyme preparations but the rates are appreciably slower than that for the 

 polymerization of TPP by the purified enzymes, suggesting the presence of 

 different enzymes for each of the deoxyribonucleoside triphosphates. Mixtures of 

 the triphosphates give additive rates, further suggesting diflferent enzymes. 



The enzymatic formation of the di- and tri-phosphates of deoxycytidine and 

 thymidine from deoxycytidylate and thymidylate takes place in the cytoplasmic 

 fraction of rat liver homogenates (Hecht et al., 1954) while the phosphorylation 

 of deoxyadenylic acid to the di- and tri-phosphates occurs in kidney and muscle 

 (Sable et al., 1954). 



F. Phospholipid Biosynthesis 

 I. Ethanolmnine and choline formation 



The inability of the rat to form labelled phospholipid-choline from glycine- i-^^^C, 

 whereas both glycine-2-^'*C and serine-3-''*C yield choline which is labelled in the 

 two carbon moiety (Arnstein, 1 95 1 ) supports the view that this latter compound 

 is formed by a decarboxylation of serine to ethanolamine (Du Vigneaud et al., 

 1946) and the subsequent methylation of the ethanolamine. A similar mechanism 

 apparently occurs in plant leaves (Bregoff and Delwiche, 1955). 



The role of ethanolamine as a choline precursor is supported by experiments 

 on the nutritional requirements of JVeurospora mutants (Horowitz, 1946; Fig. 47). 



I I 



Serine ?-»- Ethanolamine t-^-Methyl-ethonolamine ] — »-Dimethyi- _ 



/ ' I ethanolamine 



" 



Glycine Cholineless 1 Cholineless 2 Choline 



Fig. 47. Choline biosynthesis from serine and ethanolamine. 



Monomethylethanolamine accumulates in the culture medium of one cholineless 

 mutant (#2) which is active in supporting the growth of another cholineless 

 mutant (#1). Direct proof was obtained by demonstrating that in rats, ethanol- 

 amine-'^N, ethanolamine- i,2-^'*C, or deuterium labelled monomethylethanol- 

 amine, are readily incorporated into phospholipide choline {P'llgeram et al., 1953; 

 Du Vigneaud et al., 1946). The methyl groups of choline are derived from the 

 methyl group of methionine, from formate, or from other one-carbon donors 

 (Bregoff and Delwiche, 1955; Arnstein, 1951; Elwyn et al., 1955; Du Vigneaud, 

 1952; Section IV, B4, p. 56). 



