VIII. PROTEIN SYNTHESIS AND CiENE ACTION 399 



globin was formed when reticulocyte ribosomes were incubated with pH 

 5 enzymes from livers of various animals. Kruh et al. (1961) suggested 

 that this is a failure to form the peptide chain, while Lamfrom (1961) 

 postulated that species-specific "releasing" enzymes might be involved. 

 Bishop and Schweet (1962) studied this question using washed reticulo- 

 cyte ribosomes and guinea pig liver enzymes. The results with pH 5 

 enzyme fractions were similar to those noted above; little labeled, 

 soluble hemoglobin was formed. However, this failure did not involve 

 species specificity, but was related to the low incorporation observed with 

 this enzyme fraction. When a soluble fraction from liver purified by am- 

 monium sulfate fractionation was used, soluble, labeled hemoglobin was 

 formed. The soluble fraction used was prepared from a liver supernatant 

 as previously described for reticulocyte enzymes (Allen and Schweet, 

 19621. The amount of labeled hemoglobin formed with liver enzymes 

 plus reticulocyte ribosomes was similar to the amount formed with 

 reticulocyte enzymes added at a level to give the same total incorporation. 



TABLE VI 

 Pr(jperties of Escherichia coli and Reticulocyte Ribosomes" 



Reticulocyte 

 Treatment E. coli system system 



Ribonuclease (1 Mg/ml) Complete inhibition Complete inhiljition 



DNase (5-50 Mg/ml) 55% inhibition No effect 



Chloramphenicol (10-3 il/) 91% inhibition 3% inhibition 



Puromycin (10"^ M) 95% inhibition 95% inhibition 



Add TMV-RXA after DNase 80-fold stimulation No effect 



Internal ribosomal ribonuclease Present Absent 



" Data summarized from Tissieres et al. (1960); Nathans et al. (1962); Schweet et al. 

 (1961); and unpublished data. 



Lack of inhibition by DNase (Table VT) is consistent with the 

 concept that information transfer is not necessary for hemoglobin syn- 

 thesis in the cell-free system as suggested by the lack of RNA synthesis 

 in the intact cell. However, even in the absence of addition of messenger 

 RNA, the activity of the reticulocyte ribosomes is comparable to ''stimu- 

 lated" E. coli ribosomes (Table V). Further efforts along these lines in 

 several laboratories involve attempts to add various types of synthetic 

 polynucleotides or messenger RNA, such as TMV (Table VD. So far our 

 own results with TMV-RNA have been negative. However, a striking 

 stimulation of phenylalanine incorporation has been obtained with 

 poly-U, confirming the report by Arnstein." Further, hemoglobin syn- 

 thesis can be almost completely abolished by pro-incubation in the 



