I. REPLICATION OF DNA IN CELL-FREE SYSTEMS 13 



and Potter, 1953) suggests that Sable's observation is probably related 

 to the reports of Klenow and Anderson (1957) and Klenow and Lichtler 

 (1957) that adenylate kinase (myokinase) can use adenine deoxyribo- 

 nuclcotides in place of ribonucleotides. This possibility is supported by 

 the observation of Lehman et al. (1958) that heated extracts of E. coli 

 |iartially retain their ability to phosphoiylate deoxyadenylate while 

 losing their capacity to phosphorylate other deoxyribonucleotides. Heat 

 stability is a characteristic property of muscle adenylate kinase (Colo- 

 wick and Kalckar, 1943). Hecht et al. (1954) reported the phosphoryla- 

 tion of dCMP to dCDP and dCTP in a cytoplasmic fraction of rat liver 

 and found that dTTP was formed from dTMP in extracts of regenerating 

 liver. Ochoa and Heppel (1957) have observed the phosphoiylation of 

 (lAAIP and dCMP in extracts of Azotobacter. Bollum (1958) and Canel- 

 lakis and Mantsavinos (1958) reported the enzymatic synthesis of all 

 four deoxyribonucleoside triphosphates in extracts of regenerating rat 

 liver, and Kielley (1961) has studied these reactions in mouse liver. 



It is interesting that normal rat liver, which can catalyze the phos- 

 l)horylation of dAIVIP, dCMP, and dGMP by ATP, has at most a very 

 low capacity to phosphoiylate dTMP. The rapid increase in thymidylate 

 kinase activity in regenerating rat liver has been demonstrated in several 

 laboratories (Bollum and Potter, 1959; Canellakis, 1959; Canellakis et 

 al., 1959; Smellie, 1960). This increase is inhibited by post-operative 

 X-irradiation of the whole animal (Bollum et al, 1960). Weissman et al. 

 (1960) and Smellie (1960) have studied the sequence of reactions: 



Deoxythymidine ^ dTMP ^ dTDP -^ dTTP 



They claim that there is a sequential induction or appearance of thymi- 

 dine kinase, thymidylate kinase, and thymidine diphosphate kinase in 

 regenerating rat liver and in an L-strain of fibroblasts grown in tissue 

 culture. 



Gray et al. (1960) and Smellie (1960) claim there is a factor in 

 normal rat liver which inhibits thymidylate kinase when added to 

 extracts of regenerating liver. It is heat-labile, nondialyzable, and has no 

 effect on dAMP, dOAIP, and dCMP kinases. It will be interesting to 

 learn more of this inhibitor which could play a role in regulating dTTP 

 formation. 



Bianchi et al. (1961) have reported that dTDP is not an intennediate 

 in dTTP synthesis in mouse lymph nodes and leukemic spleen. They 

 suggest that a pyrophosphate group is transferred from ATP according 

 to the following reaction : 



dTMP + ATP -> dTTP + AMP 



