1 66 



SUBCELLULAR PARTICLES 



o£ 37°C. Ascites cells were incubated with C"-leucine, and at various time inter- 

 vals cells were lysed and fractionated by differential centrifugation. The S-RNA, 

 RNA and protein of the ribonucleoprotein particles and the remaining soluble pro- 

 teins were isolated. Figure 6 indicates that S-RNA was maximally labeled before 

 the other fractions had reached their highest specific activity, thus indicating 

 that this unique type of RNA might be an intermediate in the process. 



Dr. Hoagland has added amino acid-labeled S-RNA to microsomes plus a 

 small amount of supernatant enzyme, and has shown that 20 per cent of the amino 

 acid appeared in the protein (22). This reaction was dependent upon GTP and 

 ATP as well as a small amount of the supernatant fraction, whereas GTP was 

 not necessary for amino acid activation or binding to S-RNA. Addition of C^-- 

 amino acid to the reaction mixture did not change the amount of C^*-amino 

 acid incorporated into protein. This indicated that the amino acids bound to the 

 S-RNA do not et]uilibrate with free amino acids before they become incorporated 

 into protein. Hultin and Beskow have also presented evidence for a bound amino 

 acid intermediate which does not equilibrate with free amino acids (26, 27). 



Figure 7 shows a time curve for the transfer of this type of S-RNA-bound 

 amino acid to microsome protein (22). In this case, a liver pH 5 fraction was 

 incubated with C^^-amino acid and ATP, and the free amino acids and ATP 

 were removed by reprecipitation at pH 5.2 from dilute solution. This reprecipi- 

 tated and redissolved pH 5 fraction contained C^^-amino acids presumably 

 bound to the S-RNA therein. This labeled enzyme fraction was subsequently 

 incubated with liver microsomes and CjTP. The figure indicates the rapid loss 

 of amino acid from this fraction with the concomitant incorporation of the 

 amino acid into microsomal protein. 



il]3i000 



o 



on 

 a. 



<3 



2P00- 



Q. 

 O 



1.000 



^RNP PARTICLES 



20 



MINUTES 



< 

 1,000^ 



500 o 



Fig. 6. Time curxc dI' incuiporation of L-leucine-C" into tlic RNA and protein of the 

 ribonucleoprotein particles ami the solulile fraction in intact ascites cells. [From (22).] 



