98 MICROSOMAL PARTICLES 



not very informative, and the measured leucine-arginine ratio, which lies almost 

 exactly half way between microsomal protein and hemoglobin, is very ambigu- 

 ous. These results are compatible with the assumption that the transient ma- 

 terial in the microsome is largely pre-hemoglobin. More evidence is needed to 

 confirm this assumption, however. 



Boiling microsomes with 66 per cent ethanol extracted about 25 per cent of the 

 radioactive material, whose free amino nitrogen increased greatly on renuxing 

 with 6 N HC1, suggesting that this material is peptide in nature. The specific 

 activity of the material was 5 to 10 times that of the unextracted microsomal 

 protein. Both the extractable and nonextractable radioactive materials were 

 transient; i.e., the counts were removed on incubating cells in nonlabeled amino 

 acid for 15 minutes. 



One would like to conclude from the above extraction data that the extracted 

 material is richer than the whole microsome in growing peptide chains of short 

 length, whereas the unextractable material represents growing peptide chains 

 which are too long to dissolve in 66 per cent ethanol. The short chains would 

 presumably represent the earliest stages of hemoglobin formation in the micro- 

 some. Further purification and characterization are necessary to prove this 

 point. 



SOME NUMEROLOGY AND CONCLUSIONS 



The above data lead to some interesting results if the following assumptions 

 (or approximations) are made: (1) hemoglobin is the only protein being made 

 in rabbit reticulocytes; (2) all microsomal particles are equally active in syn- 

 thesizing hemoglobin; (3) all the transient label in the microsome is pre-hemo- 

 globin. The steady-state label level of the microsomal particle, the specific ac- 

 tivity of leucine used, together with the facts that 12 per cent of the protein is 

 leucine and one-half of the microsome is protein, lead to the conclusion that 

 0.05 per cent of the mass of the microsomal particle is pre-hemoglobin, i.e., grow- 

 ing peptide chain. Since the molecular weight of the microsomal particle is 

 4,000,000 as shown above, this means that the average weight of growing chain 

 per particle is 2000. In a random population of growing chains the average 

 weight might be expected to be about one-half of the finished chain weight. If 

 all the growing chain per particle is in one piece, this leads to a value of 4000 

 for the finished weight of polypeptide chain made per microsomal particle, a 

 value reasonably close to the weight of one-fourth of a hemoglobin molecule, 



