VOL. 12 (ig53) BIOSYNTHESIS OF NUCLEOSIDES AND NUCLEOTIDES 255 



ribose-i-phosphate stems either from ingested inosine or from hexosephosphates 

 generated in ordinary carbohydrate metabohsm^®. 



Role of ribose-i ,^-diphosphate in nucleotide formation and in the biosynthesis of purines. 

 The possibihty of alternative pathways for ribose-i -phosphate as an inter mediary 

 metabohte, is a crucial point in a discussion of nucleotide synthesis. Two independent 

 investigators studying h3'poxanthine synthesis in pigeon liver have found evidence for 

 ribose-5-phosphate as the carrier of both incomplete purine precursors and complete 

 purines. Greenberg^^ found that ^*C labelled formic acid added to pigeon liver extract 

 appeared in the hypoxanthine of 5-inosinic acid (hypoxanthine-ribose-5-phosphate) in 

 markedly higher concentration than in that of inosine of free hypoxanthine [cf. Table II). 

 Inosine had a conspicuously low isotope concentration and Greenberg was therefore 

 led to believe that in the synthesis of purine nucleotides the nucleosides were bypassed. 



TABLE II 



INOSINE NOT PRECURSOR OF IMP-5 IN DE NOVO SYNTHESIS 



Specific activity, cts./per/^M 



Inosine- 5-phosohMe Inosine Hypoxanthine 



18,600 9,900 4.730 



Experiment H-27, vessel i. System i ml of 



1 : 2 liver homogenate i j.iM. of inosine, 1 2 /iM 



of ^*C-formate (68- 10^ cts. per ^mol), time 



21 minutes. 



From G. R. Greenberg, /. Biol. Chem., 190 

 (1951) 623. 



The nucleosides were considered to be breakdown products from nucleic acids and nucleo- 

 tides and not steps on the biosynthetic pathway to the latter compounds. Such a by- 

 passing of nucleosides was also indicated in some studies on nicotinamide nucleotide 

 synthesis in hemolysates^. A third observation of importance was the stimulation of 

 inosinic acid synthesis in pigeon liver extract by the combined action of adenosine tri- 

 phosphate and ribose-5-phosphate (Buchanan^^). 



It will be noticed that the studies performed on unfractionated extracts of tissues 

 have given, in general, some extra information about nucleotide metabolism and have 

 given hints about the existence of reactions which have so far not been detected in more 

 fractionated systems. Consequently we initiated studies on more or less intact or un- 

 fractionated liver preparations. Bennett and I injected S-^^C adenine into a perused 

 rabbit liver and found that the adenine disappeared largely from the blood within 5 

 minutes. A large proportion was found in the adenylpyrophosphate (ADP,ATP) fraction*'^. 

 Goldwasser studied the purine metabolism of liver slices and homogenates and found 

 that a marked incorporation of adenine into nucleotide took place'*^. Homogenates of 

 pigeon liver turned out as on so many previous occasions to be the most reproducible 

 cell free system. The supernatant obtained from spinning at 100,000 g was practically 

 free of microsomes and possessed all the activity. The adenine was incorporated into 

 5-adenylic acid or adenosine diphosphate*^. Saffran and Scarano'^^ have found that 

 the particle free extract is completely inactivated by dialysis but can be completeh' 

 reactivated by adding ribose-5-phosphate and ATP. The ribose-5-phosphate can be 

 References p. 263I264. 



