Cell Structure and Protein Synthesis 461 



The total fraction of cytoplasmic granules was divided by differential centri- 

 fuging into mitochondria, microsomes and intermediate granules, termed 'light 

 large granules', which are different from mitochondria or microsomes. Upon 

 separate incubation of each of these three fractions of granules, no synthesis of 

 amylase occurred. It was further found that amylase can be synthesized from 

 amino acids by the 'light large granules' if they are supplemented with mito- 

 chondria or with a medium in which mitochondria were preliminarily incubated. 

 The mediimi is activated and capable of stimulating amylase synthesis in the 

 'light large granules' only if the mitochondria were incubated in aerobic condi- 

 tions in the presence of ATP and of a respiratory substrate. Hence, mitochondria 

 form some substances which in addition to amino acids are required by other 

 granules for protein synthesis. These substances ('mitochondrial factor'), are, 

 possibly, identical to the 'Siekevitz factor' required for incorporation of labelled 

 alanine into the proteins during incubation of microsomes. The 'mitochondrial 

 factor' may be formed in the absence of added amino acids. But since mitochondria 

 always contain some quantity of free amino acids, the possibility that amino 

 adds form part of the 'factor' cannot be excluded. 



As regards these experiments the objection was brought forward that there 

 might have occurred in the course of incubation of the granules, an activation of 

 some precursor rather than a synthesis of amylase. Therefore experiments were 

 undertaken to determine the total amoimt of protein before and after incubation 

 of granules in different conditions. It was shown that incubation of 'hght large 

 granules' in the presence of amino acids and of the medium preincubated with 

 mitochondria results in an increase of the total protein content of the system. 

 The overall synthesis of total protein, Uke the formation of individual proteins, 

 requires the simultaneous presence of a full set of amino acids. If the medium is 

 lacking even one of the essential or non-essential amino acids, the rate of syn- 

 thesis of total protein is greatly lowered. Apparently no synthesis of proteins, 

 with changed amino acid composition, occurs when individual amino acids are 

 absent. These data and those of the experiments on amylase formation show that 

 upon incubation of granules there occurs sjmthesis of the specific proteins 

 characteristic of the granules of the utilized tissue. 



'Light large granules' were obtained not only from the pigeon pancreas, but 

 also from regenerating rat liver. The liver granules synthesize proteins in the 

 same conditions as the pancreatic granules. In these experiments the total amount 

 of protein formed during 30 minutes' incubation occasionally reached 10% of 

 the initial quantity of protein in the granules. 



Thus, synthesis of many proteins proceeds in cytoplasmic granules of a definite 

 type, different from the mitochondria and microsomes. The 'hght large granules' 

 exhibit a high content of ribonucleic acid; its concentration can reach 260 /xg/mg 

 of dry protein, i.e. not less than in the microsomes which were hitherto considered 

 as the cytoplasmic units containing the largest quantity of ribonucleic acid. A 

 study of the incorporation of labelled amino acids injected into the living 

 organism has shown that our conclusion concerning the role of 'light large 

 granules' in protein synthesis, which was based on experiments in vitro, also 

 applies to the processes which take place in the intact cells of a living organism. 



