37. NUCLEIC ACID AND PROTEIN SYNTHESIS 



387 



this view. To establish the fact that RNA-amino acid compounds are 

 true intermediates in protein synthesis, however, it is necessary to show, 

 in vivo, that they satisfy the kinetic requirements for an intermediate 

 and to show, in vitro, that they may serve directly as the source of amino 

 acid in specific newly synthesized protein molecules. Some approaches to 

 satisfying these criteria have been made. 



Because of the rapidity of labeling of ribosomal protein in in vivo systems 

 it was necessary to slow down the process in order to investigate events 

 which might precede the appearance of amino acid in protein. This was 

 accomplished by incubating ascites tumor cells in ascitic fluid at a reduced 

 temperature. By this means, it was shown that following the exposure 

 of the cells to a C 14 -amino acid, the radioactivity appeared initially in 

 the sRNA fraction, rising rapidly to a plateau value. At a slower rate 

 the amino acid found its way into particle RNA where it also quickly 

 reached a low but definite plateau value. The particle protein accumulated 

 label even more slowly but progressively and finally label appeared in 

 soluble protein. 117 Furthermore, as we see in Fig. 4, from the recent work 

 of Stephenson and Zamecnik (cf. Zamecnik 96 ), when maximal activity has 

 been introduced into the RNA fractions, the addition of a quenching 

 dose of C 12 -amino acid resulted in a rapid loss of label from sRNA. Con- 



3600 



3000 



PROTEIN 



RNA 



quenching 



Soluble RNA 



700 



- 500 



300 



100 



tr 



E 



E 

 b. 

 6 



40 10 



minutes 



Fig. 4. The incorporation of valine-C 14 into.the RNA and protein of whole Ehrlich 

 ascites tumor cells incubated in their own ascitic fluid at 20°C. At 20 minutes valine- 

 C 12 was added to the incubation medium such that the intracellular valine-C 14 specific 

 activity was reduced to about a third of its previous activity. [From P. C. Zamecnik, 

 Harvey Lectures 54, 256 (I960).] 



