Further study of factors affecting amino acid incorporation into protein by isolated mitochondria.

1. The incorporation of amino acids into protein in isolated mitochondria has now been studied in more detail, and with mitochondria from a range of tissues. 2. Liver mitochondria from newborn rats are twice as active as those from adult rats. 3. The mitochondria are inactivated by excessive homogenization and repeated freezing and thawing. 4. The incorporation is sensitive to conditions of incubation and in particular to the rate of oxygenation, shape of vessel and depth of fluid. Best results are obtained by incubation in flat-bottomed vessels containing suspensions with less than 3mm. depth of fluid. 5. The requirement for oxidizable substrates has been examined with a range of substances, and most of the common energy-yielding metabolites of the mitochondrion are effective. Their activity is greatly influenced by concentration, and some, but not all, of the substrates show optimum concentrations for incorporation with decreased activity at higher concentrations. 6. Some amino acids can act as energy sources for the incorporation. 7. The effect of increasing the concentration of labelled amino acid is different for different amino acids, and complex effects occur on the addition of amino acid mixtures. 8. It is concluded that the amino acid incorporation into protein finally obtained is the result of many interacting factors related to the structural and metabolic state of the mitochondrion.