Amino Acid Transport and Early Stages in Protein 

 Synthesis in Isolated Cell Nuclei 



Vincent G. Allfrey 

 The Rockefeller Institute, New York, U.S.A. 



The application of tracer techniques to the study of nuclear protein 

 synthesis dates back to 1948, to the exciting experiments of Bergstrand, 

 Eliasson, Hammarsten, Norberg, Reichard and von Ubisch ; they showed 

 that, after the injection of [^^N]-glycine, the ^'N concentration in the pro- 

 teins of the liver cell nucleus was just as high as that of proteins in the 

 cytoplasm [i]. This great disclosure of the dynamic state of protein meta- 

 bolism in the interphase nucleus is now a classic observation in nuclear 

 biochemistry and it has become a point of departure for many subsequent 

 experiments on protein synthesis in the nuclei of intact organisms. 



Until 1954, virtually all tracer work on nuclear protein synthesis was 

 based on similar experiments carried out in vivo, in which nuclei or nuclear 

 protein fractions w^ere isolated from the tissues of animals previously 

 injected with isotopic amino acids. (This procedure is still clearly the 

 method ot choice for studying many aspects of the physiology of the 

 nuclear proteins (e.g. [2, 3].) 



However, in 1954 the surprising synthetic potentialities of the isolated 

 thymocyte nucleus came to light [4], and it became possible to study the 

 chemical mechanisms employed by the isolated nucleus in incorporating 

 free amino acids into polypeptide chains. This aspect of nuclear protein 

 synthesis has held our interest for the past five years, and some of the re- 

 actions and sites involved in amino acid incorporation are now known. 

 Part of this work, done in collaboration with Drs. A. E. Mirsky, J. W. 

 Hopkins, and J. H. Frenster, will now be described. It presents our current 

 views of the protein synthetic pathway in the cell nucleus. The treatment 

 of previously published work will omit details in order to permit the des- 

 cription of some newer aspects of nuclear amino acid and protein 

 metabolism. 



Pathways in nuclear protein synthesis 



AMINO ACID UPTAKE BY ISOLATED CELL NUCLEI 



Suspensions of nuclei possessing a high degree of purity can be ob- 

 tained by differential centrifugation of homogenates of calf thymus tissue. 



