The cell-free protein synthetic activity of the postmitochondrial supernatant isolated from whole brain of 6- to 32-month-old male Fischer F344 rats was compared. Protein synthesis decreased 56% from 6 to 32 months of age. The decrease in cell-free protein synthesis was not due to an age-related increase in RNase activity. Although monomeric ribosomes (ribosomes stripped of mRNA) isolated from the brains of older rats were less active in polyuridylic acid directed polyphenylalanine synthesis, the fidelity of polyuridylic acid translation by monomeric ribosomes did not decrease with increasing age.
The synthesis of various types of ribonucleic acid (RNA) isolated from 6- to 3-month-old female Fischer F344 rats was compared. The rate of RNA synthesis by freshly prepared hepatocytes was determined by dividing the amount of [3H]orotic acid incorporated into RNA as uridine-5'-monophosphate by the specific activity of the uridine-5'-triphosphate pool. The rate of total RNA synthesis by hepatocytes from 19-month-old rats was 40% less than the rate for hepatocytes from 12-month-old rats. No significant difference in the rate of total RNA synthesis was observed between 19 and 30 months of age. The percentage of [3H]orotic acid incorporated into poly(A) + RNA by 30-month-old rats was approximately 50% less than that observed for hepatocytes isolated from 6-month-old rats. The percentage of [3H]orotic acid incorporated into poly(A)-RNA as ribosomal RNA (38S, 18S, and 5S RNAs) or transfer RNA was similar for 12- and 30-month-old rats. The rate of poly(A) + RNA synthesis by hepatocytes isolated from 30-month-old rats was 65% less than that observed for hepatocytes from 6-month-old rats. In contrast to total RNA synthesis, the rate of poly(A) + RNA synthesis for the 30-month-old rats was significantly less than the rate for 19-month-old rats.
The rate of total protein synthesis in isolated hepatocytes was determined. The incorporation of L-[3H]valine into protein is linear for at least two hours of incubation and is affected by the concentration of amino acids in the medium. Uptake of valine by hepatocytes from 1.5- and 18-month-old rats was identical and appears to occur by simple passive diffusion. Within five minutes, the specific activities of the intracellular and extracellular valine pools are equivalent. The specific activities of these pools are saturated by 1.6mM valine and remain constant for 60 minutes of incubation. The rates of protein synthesis by hepatocytes from 1- to 2-month-old rats is 96.8 pmoles of valine per minute per milligram protein. This is comparable to rates of protein synthesis reported for perfused liver and liver in vivo and is approximately 64% higher than the rate of protein synthesis by hepatocytes from 18-month-old rats.
The cell-free protein synthesis by the postmitochondrial supernatant from chicken cerebrum was twofold greater than protein synthesis by the cerebellum or optic lobes. Ribosomal aggregation of mRNA and ribonuclease activity of the postmitochondrial supernatant from the three brain regions was not statistically different. The higher protein synthetic activity of the cerebral postmitochondrial supernatant was associated with both the postribosomal supernatant (cell sap) and microsomal fractions. Cerebral monomeric ribosomes were more active in polyuridylic acid directed polyphenylalanine synthesis than monomeric ribosomes from either the cerebellum or optic lobes. The ability of cerebral cell sap to support polyuridylic acid directed polyphenylalanine synthesis was 1.6 to 2 times greater than cell sap from the other two regions. Cell sap factors other than tRNAphe or phenylalanyl-tRNA synthetases appear to be responsible for the higher protein synthetic activity of the cbr cell sap.
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