REGULATION OF RESPIRATION AND ENERGY TRANSDUCTION IN CYTOCHROME-C-OXIDASE ISOZYMES BY ALLOSTERIC EFFECTORS

The binding of TNP-ATP (2' or 3'-O-(2,4,6-trinitrophenyl)-ATP) to cyto chrome c oxidase (COX) from bovine heart and liver and to the two-subu nit COX of Paracoccus denitrificans was measured by its change of fluo rescence. Three binding sites, two with high (dissociation constant K- d = 0.2 mu M) and one with lower affinity (K-d = 0.9 mu M), were found at COX from bovine heart and liver, while the Paracoccus enzyme showe d only one binding site (K-d = 3.6 mu M). The binding of [S-35]ATP alp ha S was measured by equilibrium dialysis and revealed seven binding s ites at the heart enzyme (K-d = 7.5 mu M) and six at the liver enzyme (K-d = 12 mu M). The Paracoccus enzyme had only one binding site (K-d = 16 mu M). The effect of variable intraliposomal ATP/ADP ratios, but at constant total concentration of [ATP + ADP] = 5 mM, on the H+/e(-)s toichiometry of reconstituted COX from bovine heart and liver were stu died. Above 98% ATP the H+/e(-)stoichiometry of the heart enzyme decre ased to about half of the value measured at 100% ATP. In contrast, the H+/e(-)stoichiometry of the liver enzyme was not influenced by the AT P/ ADP ratio. It is suggested that high intramitochondrial ATP/ADP rat ios, corresponding to low cellular work load, will decrease the effici ency of energy transduction and result in elevated thermogenesis for t he maintenance of body temperature.