Rapid Separation of Particulate and Soluble Fractions from Isolated Cell Preparations (Digitonin and Cell Cavitation Procedures)

This chapter discusses the enzymes of metabolic pathways (such as gluconeogenesis and ureogenesis) that are located in two different cellular compartments: the mitochondria and the cytosol. Communication among these compartments is maintained by the transport of certain metabolites across the inner mitochondrial membrane. The regulation of the reaction sequences and the rates of the over/ill processes depends (1) on the activity of the individual enzymes, (2) on the concentration of different metabolites and effectors in the individual compartments, (3) on the redox state of mitochondrial and cytosolic NAD (P), and (4) on the phosphorylation potential in the two compartments. The metabolite indicator method that has been developed to estimate both the redox state and the phosphorylation potential in the mitochondrial and cytosolic compartments from measurements of the total content of metabolites in the tissue. The distributions of metabolites in the two cellular compartments have been estimated using procedures based on the metabolite indicator method. Studies with isolated mitochondria have shown that considerable concentration gradients of certain metabolites exist across the mitochondrial membrane.