100 THE BIOSYNTHESIS OF PROTEINS 



1956; Berg, 1956). But at such high concentrations, hydroxylamine reacts 

 even with common esters (Chantrenne, 1948; Raacke, 1958). The reactivity 

 towards amines of enzyme bound aminoacyl adenylates is thus much lower 

 than that of the free form. Incidentally, the high reactivity of aminoacyl 

 adenylates raised doubts as to the meaning of amino acid incorporation 

 into proteins in the presence of activation enzymes which produce amino 

 acid adenylates: could not the observed incorporation be explained by a 

 non-enzymic reaction of the mixed anhydride with — NH2 groups of pre- 

 existent proteins (Zioudrou et ah, 1958; Zioudrou and Fruton, 1959)? The 

 low reactivity of the enzyme bound anhydride with common amines 

 permits us to discard this objection ; but it would be a fundamental mistake 

 to provide an experimental system with ready made free aminoacyl 

 adenylates, for these — at least the fraction in excess to what can be bound 

 by the corresponding enzyme — would immediately react at random with 

 most available — NH2 groups (Wong et ah, 1959). 



Aminoacyl adenylates are anhydrides of the amino acids, but they cannot 

 leave the enzyme, and they are not very reactive in this bound form even 

 with as eager an acceptor as hydroxylamine. One may wonder then whether 

 such an activated form can be an intermediate in protein synthesis, and 

 what its normal acceptor is. 



These two questions are actually linked. Let us consider first the fate of 

 the activated amino acid. 



In amine acetylation (Lipmann, 1950), in the synthesis of hippuric acid 

 (Chantrenne, 1951; Schachter and Taggart, 1954), the active acyl residue 

 of the enzyme-bound acyladenylate is picked by coenzyme A, a typical 

 acyl carrier which conveys it to the condensing enzyme where the acyl 

 radical will meet the acceptor amine and condense with it in a practically 

 irreversible reaction. But aminoacyl residues are not transferred to 

 coenzyme A (Jencks and Lipmann, 1957). 



The discovery of the amino acid acceptor was a major achievement of 

 work on tissue homogenates and disrupted cells, which will now be con- 

 sidered briefly. The first experiments designed with the hope of observing 

 incorporation of labelled amino acids into homogenates gave depressingly 

 low incorporation indeed. It was established, however, that the incorpora- 

 tion was real and not due to contamination, and that it depended on 

 respiration (Winnick et al., 1948) or more precisely on oxidative phos- 

 phorylation (Frantz et ah, 1948 ; Borsook et al, 1949). The amino acids were 

 shown to be incorporated within polypeptide chains (Winnick, 1949, 1950) 

 and the incorporation of one labelled amino acid was stimulated by a mix- 

 ture of the others. The recognition of a few pitfalls (Borsook et al, 1949; 

 Winnick, 1950; Peterson and Greenberg, 1952) and their elimination, and 

 progress in the methods of preparation of homogenates (Schneider and 

 Hogeboom, 1951; Bucher, 1953) provided extracts in which amino acid 



