INHIBITORY HIGH AFFINITY BINDING-SITE FOR ADP IN THE OLIGOMYCIN-SENSITIVE ATPASE OF BEEF-HEART SUB-MITOCHONDRIAL PARTICLES

Kinetic evidence are presented for the existence of a high affinity inhibitory site for ADP/Ki < 10-7 M/ in the oligomycin-sensitive ATPase of beef heart submitochondrial particles. The ATPase .cntdot. ADP complex is completely inactive in the ATPase reaction; it can be converted into active ATPase in a slow ATP-dependent reaction. The dependence of a 1st order rate constant for activation of the enzyme .cntdot. ADP complex on concentration of ATP gives a Km value equal to that for ATP in the ATPase reaction. The membrane-bound ATPase complex probably contains 2 kinetically distinct nucleotide-binding centers, i.e., center 1 binds ATP or ADP with a formation of enzyme-substrate or enzyme-competitive inhibitor complexes; center 2 binds ADP with a formation of a complex which is able to bind ATP in center 1 and unable to hydrolyze the bound ATP. The binding of ATP or ADP in center 1 changes the reactivity of center 2 towards ADP.