CHEMICAL BONDS IN NUCLEIC ACIDS 441 



They were able to isolate from acid hydrolysates thymidine-3', 5 '-diphos- 

 phate (XXXI; R = thymine residue) and deoxycytidine-3', 5 '-diphosphate 

 (XXXI ; R = cytosine residue) and to establish their structure by synthesis; 

 they also obtained some evidence for the presence of small amounts of the 

 corresponding 3 ',5 '-diphosphate of 5-methyldeoxycytidine in their hydroly- 

 sates. The isolation of these substances is a clear indication that both the 

 3'- and 5 '-positions in the deoxyribonucleosides are involved in the inter- 

 nucleotidic linkage and emphasizes the difference between the hydrolytic 

 mechanism which operates in ribonucleic and deoxyribonucleic acids. 



O- 



I OPO3H2 



R-CH-CHj 



I OH 



CHj-OPOsH. R-CH-CH2 — \- 



CHz-OPOjHa 



H H H H 



XXXI XXXII 



Chemical hydrolysis of deoxyribonucleic acids is not a satisfactory 

 method for obtaining simple mononucleotides mainly on account of the 

 lability of the glycosidic linkage in the purine deoxyribonucleotides to acid, 

 and the resistance of the normal phosphodiester group to alkaline hy- 

 drolysis. Klein and Thannhauser/"** however, using an arsenate-inhibited 

 intestinal phosphatase, succeeded by enzymic hydrolysis in isolating four 

 nucleotides (deoxyadenylic, deoxyguanylic, deoxycy tidy lie, and thymidylic 

 acid) ; Volkin, Khym, and Cohn'"*^ also described their separation from simi- 

 lar hydrolysates. Similarity to the ribonucleoside-5 '-phosphates in their 

 ion-exchange characteristics and their dephosphorylation^'*^ by a specific 

 5 '-nucleotidase""* indicated that, contrary to earlier assumptions, they were 

 the 5'-phosphates of the respective nucleosides (XXXII; R = purine or 

 pyrimidine residue). Conclusive proof for this formulation has been pro- 

 vided at least in the case of the pyrimidine nucleotides by identification of 

 the natural substances through direct comparison with thy midine-5 '-phos- 

 phate (XXXII; R = thymine residue) and deoxycy tidine-5 '-phosphate 

 (XXXII; R = cytosine residue) prepared by unambiguous methods by 

 Michelson and Todd."' The almost quantitative recovery (ca. 92%) of the 

 deoxyribonucleoside-o '-phosphates from snake venom diesterase hydroly- 

 sates of deoxyribonuclease-treated deoxyribonucleic acids^"''^ shows that 



'••^ See also S. J. Thannhauser and B. Ottenstein, Z. physiol. ('hem. 114, 39 (1921); 



S. J. Thannhauser and G. Blanco, ibid. 161, 116 (1926). 

 '" W. Klein and S. J. Thannhauser, Z. -phisiol. Chem. 218, 173 (1933); 224, 252 (1934); 



231,96 (1935). 

 '" E. Volkin, J. X. Khym, and W. E. Cohn, J. Am. Chem. Soc. 73, 1533 (1951). 

 1" C. E. Carter, J. Ain. Chem. Soc. 73, 1537 (1951). 

 '« L. A. Heppel and R. J. Hilmoe, J. Biol. Chem. 188, 665 (1951). 

 '<' A. M. Michelson and A. R. Todd, J. Chem.. Soc. 1953, 951; 1954, 34. 



