58 W. G. OVEREND AND M. STACEY 



c. Phosphates 



Manson and Lampen'"® reported that they obtained phosphorolysis and 

 arsenolysis of hypoxanthine deoxyriboside by enzyme preparations from 

 calf thymus gland and rat liver. An acid-stable phosphate was isolated as a 

 product of phosphorolysis. It was concluded that this ester was 2-deoxy-D- 

 ribose-5-phosphate and that it was formed from 2-deoxy-D-ribose-l -phos- 

 phate by mutase action. [Cf. Glock, Chapter 22, and Schlenk, Chapter 24.] 

 The same authors^ "^ obtained indications for the formation of 2-deoxy-D- 

 ribose-l -phosphate during the phosphorolysis of thymidine. Reverse reac- 

 tions have been demonstrated since it was shown that hypoxanthine deoxy- 

 D-riboside was formed enzymically from deoxy-D-ribose-1 -phosphate and 

 hypoxanthine.^^^ Similarly, guanine deoxyriboside could be formed by using 

 guanine in place of hypoxanthine.^"^ Manson and Lampen^^" also observed 

 nucleoside synthesis with the enzyme found in cell-free extracts of Es- 

 cherichia coli. Determination of the equilibrium constant indicates that the 

 synthesis rather than the splitting of hypoxanthine deoxy-D-riboside is 

 favored.^"* 



By enzymic phosphorolysis of guanine deoxy-D-riboside Friedkin^"^ was 

 able to isolate the labile phosphate in the form of a crystalline cyclohexyl- 

 amine salt. This was a salt of the sugar phosphate designated above as 2- 

 deoxy-D-ribose-1 -phosphate and used in the experiments on nucleoside 

 synthesis. Several reasons are forwarded to support the structure assigned 

 to this compound. The ester, which contains one mole of 2-deoxy-D-ribose 

 per mole of phosphorus, is extremely acid-labile — 50 % of the phosphorus 

 present is released as inorganic phosphate within 10-15 minutes upon 

 hydrolysis at pH 4 at 23°. This lability of the phosphate ester linkage points 

 to carbon atom 1 as the site of esterification, and indeed a free aldehyde 

 group is also released upon hydrolysis at pH 4. 



The phosphate resulting from the mutase action on 2-deoxy-D-ribose-l- 

 phosphate was isolated as the barium salt. Hydolysis with A^ hydrochloric 

 acid for 7 minutes liberated 45 % of the organic-bound phosphate. Since 

 the sugar phosphate was reducing and gave no formaldehyde on periodate 

 oxidation, it was, in view of its formation from hypoxanthine deoxy-D- 

 riboside, assumed to be 2-deoxy-D-ribose-5-phosphate. 



Arsenolysis of hypoxanthine deoxy-D-riboside resulted in the formation 

 of hypoxanthine and free 2-deoxy-D-ribose.^°^ Probably the primary product 



3°^ L. A. Manson and J. O. Lampen, Abstracts Papers 114th Meeting Am. Chem. Soc 



53C (1948); /. Biol. Chem. 191, 95 (1951). 

 307 L. A. Manson and J. O. Lampen, Federation Proc. 8, 224 (1949). 

 SOS M. Friedkin, /. Biol. Chem. 184, 449 (1950). 



309 M. Friedkin and H. M. Kalckar, /. Biol. Chem. 184, 437 (1950). 

 3'o L. A. Manson and J. 0. Lampen, Federation Proc. 9, 397 (1950). 



