368 RICHARD SCHWEET AND JOHN BISHOP 



Nathan.s and IjpiiKinii (1961) have reported that after discharging its 

 amino acid, E. coli RNA coidd be reisohited (by i)henol extraction) and 

 completely recharged. 



The original "adaptor hypothesis" (Crick, 1958) called for a rela- 

 tively small polynucleotide as the adaptor. It was suggested (Hoagland, 

 1960) that the relatively large size of the transfer RNA molecule was a 

 requirement for specific interaction with the activating enzyme and that, 

 after accepting the amino acid, the transfer RNA molecule might release 

 a small polynucleotide fragment with the amino acid attached to it. 

 The results cited above suggest that the transfer RNA does not break 

 down. Hoagland and Comly (1960) have provided careful studies which 

 point to the same conclusion. C'*-leucyl-RNA was incubated with various 

 enzyme fractions, or microsome extracts, in the presence and absence of 

 GTP, but in the absence of microsomes. At various times parallel assays 

 were made of the remaining acid-precipitable amino acid, and of the 

 competence of the complete incubation mixture in the transfer process. 

 It was found in every case that the amount of C"-leucine transferred to 

 peptide linkage was proportional to the amount of acid-jirecipitable 

 C"-leucine remaining. Thus, free, acid-soluble, amino acid polynucleo- 

 tide compounds do not appear to be significant intermediates in protein 

 synthesis. It remains to be seen whether such intermediates are foimed 

 in association with microsomes. 



a. The Transfer of Labeled, Soluble RNA to the Ribornicleoprotem 

 Particles. If, as discussed above, amino acyl-RNA fails to form a smaller 

 amino acyl polynucleotide, a direct physical association of the transfer 

 RNA and the microsomes, if only a veiy transient one, must occur 

 during transfer. A temperature- and energy-dependent transfer of labeled, 

 soluble RNA to ribosomes has been reported by von der Decken and 

 Hultin (1958), by Bosch et al. (1959), and by Hoagland and Comly 

 (1960). In all cases, a considerable non-temperaturc-dependent, non- 

 energy-dependent transfer of labeled RNA to the 105,(XX) g pellet was 

 also observed. The system studied in greatest detail is the rat liver system 

 of Bosch et al. (1959, 1960; Bloemendal et al, 1960, 1961). These authors 

 found that a P'^-'-S-RNA fi-action isolated on an Ecteola column was 

 readily incorporated into ribosonial RNA. The fraction was able to 

 accept C"-leucine in the presence of an activating system. After incuba- 

 tion with P'--S-RNA, the microsomes were sedimented and the micro- 

 somal RNA extracted with phenol and chromatographed on an Ecteola 

 column. Three fractions were recovered, only one of which was labeled. 

 Significantly, the labeled fraction had different chromatographic prop- 

 erties from those of the labeled soluble RNA. Furthermore, when micro- 

 somal RN.\ was isolated after P^--labeling in viiw, radioactivitv was 



