Origin of the ammonia found in protein-free extracts of rat-liver mitochondria and rat hepatocytes.
1. Protein-free extracts of isolated rat-liver mitochondria contain 5.17 +/- 0.19 nmol ammonia/mg protein [cf. Harris, E. J. and Bassett, D. J. (1971) FEBS Lett. 19, 214-217]. 2. The ammonia found in the protein-free extracts does not originate from lysosomes contaminating the mitochondrial preparation. 3. When isolated mitochondria are incubated with ornithine, 14CO2 and a source of ATP a small amount of citrulline is formed. This amount is stoichiometrically equivalent to the ammonia that disappears from the extramitochondrial space, whereas the amount of ammonia found in the protein-free extracts of the mitochondria remains unchanged. Similar results were obtained when the reductive amination of 2-oxoglutarate was used as an ammonia-consuming reaction. 4. When isolated mitochondria are incubated under conditions such that the glutamate dehydrogenase and 3-hydroxybutyrate dehydrogenase reactions reach equilibrium, the thermodynamically active concentration of ammonia is not equal to the concentration measured in the protein-free extracts. 5. About 80% of the ammonia found in protein-free extracts of rat-liver mitochondria is derived from a component or components with a molecular weight of greater than or equal to 50,000. 6. Protein-free extracts of isolated rat-liver cells contain considerable amounts of ammonia. After digitonin fractionation this ammonia is found in the protein-free extract of the particulate fraction. 7. It is concluded that the ammonia found in protein-free extracts of rat-liver tissue is derived from a component or components in the mitochondria and is released during deproteinization.
D000641:Ammonia / Q000378:metabolism*; D000818:Animals; D002222:Carbamoyl-Phosphate Synthase (Ammonia) / Q000378:metabolism; D002956:Citrulline / Q000378:metabolism; D005969:Glutamate Dehydrogenase / Q000378:metabolism; D006884:Hydroxybutyrate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism; D008297:Male; D008930:Mitochondria, Liver / Q000378:metabolism*; D051381:Rats; D013347:Subcellular Fractions / Q000378:metabolism
D002956:Citrulline; D000641:Ammonia; D006884:Hydroxybutyrate Dehydrogenase; D005969:Glutamate Dehydrogenase; D002222:Carbamoyl-Phosphate Synthase (Ammonia)