136 H. GOBIND KHORANA 



c. Incorporation of Ribonucleoside-5' -Triphosphates into the Interior of Ribo- 

 polynucleotide Chains 



Goldwasser 64 presented evidence that in the pigeon liver homogenates, la- 

 beled adenosine-5 '-phosphate was incorporated intact into the interior of 

 RNA chains. The incorporation of labeled pyrimidine nucleotide precursors 

 into the nuclear fractions of rat liver homogenates was demonstrated by 

 Herbert et al. u Edmunds and Abrams 72 working with soluble fractions of 

 Ehrlich ascites tumor cells presented evidence for the utilization of C 14 - 

 labeled ATP for the inner labeling of RNA and indicated that different 

 enzyme systems are involved in the terminal and inner incorporation of 

 nucleotides. Herbert 73 ' 75 has demonstrated that the enzymes responsible 

 for the appearance of adenine nucleotides into the interior of RNA are 

 present in the particulate fractions of tissue homogenates. 



Using a nuclear fraction of rat liver homogenates, Weiss and Gladstone 83 

 have recently described the incorporation of labeled CTP into the interior 

 of RNA. The incorporation is dependent on the presence of the other three 

 ribonucleoside triphosphates and magnesium. The triphosphates are more 

 than twice as effective as the corresponding diphosphates. In these proper- 

 ties, the system resembles the polymerases for deoxyribonucleotides de- 

 scribed in the following section. 



It should be pointed out that in the above studies the evidence for in- 

 corporation of ribonucleotides into the interior of ribopoly nucleotide chains 

 has been derived only from the alkaline hydrolysis which results in the 

 transfer of the P 32 -label in the nucleoside-5'-phosphoryl group incorporated 

 to the adjacent nucleotide. It is highly desirable to obtain additional evi- 

 dence for "extensive" internal incorporation of ribopolynucleotide precur- 

 sors. A promising tool in this field is provided by the venom phosphodies- 

 terase. 



Finally, while the evidence that is accumulating points to ribonucleoside- 

 5 '-triphosphates as being the precursors of polynucleotides it is clear that 

 the realization of well-defined enzymic systems from mammalian sources 

 for extensive ribopolynucleotide synthesis has yet to come. 



IV. Enzymic Synthesis of Deoxyribopolynucleotides 



Deoxyribonucleoside-5 '-triphosphates have so far been found to be 

 unique precursors for the enzymic synthesis of deoxyribopolynucleotides. 

 This is in contrast with the synthesis of ribopolynucleotides, which utilizes 

 both the 5'-di- and triphosphates of ribonucleosides as the monomeric 

 units. 



83 S. B. Weiss and L. Gladstone, J. Am. Chem. Soc. 81, 4118 (1959). 



