CHEMICAL PATHWAYS 93 



amines which also results in the formation of a peptide-like linkage, is 

 blocked by dinitrophenol, indicating that the energy required for peptide 

 bond formation is provided by oxidative phosphorylation. With a system 

 of soluble enzymes from pigeon liver, the requirement for ATP was 

 established, and it was found that one mole of ATP is split for each mole of 

 amine acetylated. Similar conclusions were rapidly extended to the synthesis 

 of other small peptides: hippuric acid (Borsook and Dubnoff, 1947), 

 /)-aminohippuric acid (Cohen and McGilvery, 1946, 1947) and glutathione 

 (Johnston and Bloch, 1951 ; Webster, 1953). Evidence that the synthesis of 

 enzymes in bacteria and in yeast depends on phosphorylation had mean- 

 while been obtained (Monod, 1944; Spiegelman, 1946). 



Peptidic bond formation in small peptides was thus regarded as a model 

 of possible mechanisms of polypeptide biosynthesis and these studies had 

 indeed a fundamental influence on the discovery of what is known at 

 present about the pathways of protein formation. The advent of i^C- 

 labelled amino acids was the other element which made possible the rapid 

 progresses of the last decade in this field. 



As soon as the requirement for energy coupling was discovered for small 

 peptidic compounds, it was readily established that the incorporation of 

 labelled amino acids into protein of tissue slices or of homogenates also 

 depends on respiration (Frantz et al, 1947, 1948; Winnick et al., 1947; 

 Melchior et al, 1948; Peterson and Greenberg, 1952). Amino acid in- 

 corporation, like the synthesis of small peptides, is driven by oxidative 

 phosphorylation since the inhibition of the incorporation process by dinitro- 

 phenol, runs parallel to the inhibition of phosphorylation (Frantz et al., 

 1948). Finally, it was clearly established that ATP or systems able to 

 regenerate it are required for protein synthesis in homogenates (Peterson 

 and Greenberg, 1952; Siekevitz, 1952; Zamecnik and Keller, 1954). It 

 was probable therefore that the condensation of amino acids into polypep- 

 tides involved — at one stage at least — a coupling with ATP splitting. A 

 great many works have now confirmed these data and extended them to 

 many various tissues and organisms. Coupling with ATP utilization is a 

 quite general requirement for amino acid incorporation into protein. 



Most workers in the field were long reluctant to equate amino acid in- 

 corporation to de novo protein synthesis from amino acids. The possibility 

 that incorporation corresponds to replacement of an old amino acid unit 

 by a new one within polypeptides has been a constant source of worry. 

 This question had already been raised by Schoenheimer et al. (1939) who 

 were the first to observe amino acid incorporation into protein in living 

 animals, and it became more acute in works with tissue slices or homo- 

 genates where incorporation is always very low and where no measurable 

 net increase in protein was observed. Exchange incorporation found 

 apparent support in certain facts. For instance the amino acid 'analogue' 



