VIII. PROTEIN SYNTHESIS AND GENE ACTION 371 



prelabeled ribosomes (Table III). This formation of soluble protein 

 occurs in the absence of incorporation of free C^'*-amino acids into ribo- 

 somes. This result dcmon.><trates a role for GTP at a later stage than 

 the transfer of amino acid from amino acyl-RNA. One interpretation of 

 this finding is that GTP acts to convert intraribosomal intermediates 

 into peptide linkage, thus producing completed or almost completed 

 chains which are then released. It is postulated that a GTP is split in 



TABLE III 

 Requirements for Incorporation and Release" 



Incorporation Release 



Assay components (%) (%) 



Complete system 100 100 

 No energy 1.5 4.1 



Plus 1 Mmole ATP only 51.1 50 . 2 



Plus 1 nmole GTP only 4.9 35.2 



" A. Morris and R. Schweet (unpublished data). 



the process of forming each peptide bond and that GTP does not act in 

 the initial transfer step from amino acyl-RNA. However, since ribo- 

 somes as isolated are filled with intermediates, no transfer from amino 

 acyl-RNA is observed in the absence of GTP, since no ribosomal sites 

 are available. Thus, an intraribosomal reaction requiring GTP is needed 

 in order for an earlier step, the transfer from amino acyl-RNA, to occur. 

 This mechanism is advanced as a working hypothesis to guide future 

 studies of the role of GTP. 



4. Specificity in the Transfer Reaction 



A lack of specificity for the transfer RNA species is the most strik- 

 ing aspect of this reaction. Von Ehrenstein and Lipmann (1961) demon- 

 strated that E. coli C"-amino acyl-RNA could form hemoglobin when 

 incubated with rabbit reticulocyte ribosomes. Bishop et al. (1961) showed 

 that amino acyl-RNA from guinea pig liver was competent in hemo- 

 globin synthesis with reticulocyte ribosomes. In these experiments, the 

 labeled hemoglobin was digested with trypsin and fingerprinted. The 

 correspondence of leucine-labeling in these peptides with whole-cell 

 labeled hemoglobin (Table IV) provides good evidence for the synthesis 

 of hemoglobin from this heterologous amino acyl-RNA. 



Despite the ability of guinea pig liver amino acyl-RNA to act as a 

 precursor of hemoglobin synthesis, it was observed (Bishop et al, 1961) 

 that amino acids were transferred more readily from amino acyl-RNA 

 to homologous ribosomes. Guinea pig liver transfer RNA was labeled 



