| dc.description | Cytochrome c binds ATP with marked specificity at a site that contains the evolutionarily invariant residue Arginine$\sp‘$. This binding was studied using equilibrium gel filtration, equilibrium dialysis and affinity chromatography. At physiological ionic strength the affinity is such that the major change in occupancy coincides with the normal cellular ATP concentration range, and the degree of saturation is proportional to the ratio of (ATP) /(ADP). Thus under cellular conditions binding at this site is dependent upon the energy state of the cell, providing a potential mechanism for the regulation of the respiratory chain that is sensitive to cytoplasmic ATP levels. ATP analogues were covalently attached to cytochrome c in order to determine the physiological effect of binding site occupancy. 8-azido ATP and oATP were found to react specifically at this site while the site of attachment of FSBA is unclear. The modifications did not significantly affect the overall structure or stability of the protein. However, the covalent attachment of the labels to this site, and thus by inference the non-covalent interaction, profoundly decreases the activity of the protein. The results of this study therefore support the proposal that respiration is regulated through feedback inhibition of cytochrome c activity. Inhibition of cytochrome c occurs through two compounding mechanisms: a decrease in cytochrome c's affinity for the inner mitochondrial membrane and a decreased activity with complexes III and IV. | en_US |
