INDUCED ENZYME FORMATION 



labels. Thus, the induction of enzyme synthesis in uniformly 

 labeled cells suspended in unlabeled medium should provide 

 the necessary data, providing the enzyme synthesized can be 

 isolated in a pure state and its iso topic content determined. 

 Rotman and Spiegelman (66), and Hogness, Cohn, and Monod 

 (38) independently undertook to provide data relevant to this 

 issue, using the beta-galactosidase system E. coli. 



Rotman and Spiegelman (66) secured uniformly labeled 

 cells by growth in C^^ lactate. Enzyme was induced for short 

 periods while the cells were suspended in nonradioactive me- 

 dium. The beta-galactosidase synthesized was isolated and puri- 

 fied by means of zone ionophoresis through starch columns. 

 Further purification was achieved with the aid of specific 

 precipitation with purified antiserum. The results obtained 

 revealed that less than 1 per cent carbon of the newly formed 

 enzyme molecules could have been derived from any cellular 

 components existing prior to the moment of the addition of the 

 inducer. In the experiments of Cohn, Hogness, and Monod, 

 S^^ was employed as the isotopic label and similar methods were 

 used for the isolation of the enzyme. Identical results and con- 

 clusions were obtained. 



These findings virtually eliminate any hypothesis which 

 assumes the preexistence of a complex precursor material which 

 is convertible into enzyme. It is evident that a mechanism 

 suggesting the de novo formation of enzyme from amino acids is at 

 present the only one which has received experimental support. 



A consequence of considerable importance issuing from this 

 last conclusion is that induced enzyme synthesis is thereby 

 equated to the process of protein synthesis. It follows that data 

 derived from the study of enzyme induction are pertinent and 

 relevant to the more general problem of protein formation. 

 Further, the use of inducible enzymes as model systems of pro- 

 tein formation can, in principle and fact, confer two significant 

 operative advantages. In the first place, one is assured that the 

 synthesis of a protein is being examined — a certainty not avail- 

 able to experiments dependent solely on incorporation studies. 



125 



