Escherichia coli succinic thiolinase. Stoichiometry of phosphorylation and coenzyme A binding.

Equilibrium and covalent binding studies of succinic thiokinase from Escherichia coli indicates that there can be a stoichiometric relationship between coenzyme A binding and the phosphoyrlation capacity of the enzyme. A comparison of homogeneous enzyme preparations has revealed that enzyme of high specific activity exhibits greater binding capacity and that this property is proportional to enzyme activity. Phosphorylation capacity was related to specific activity in highly active enzyme preparations, but leveled off at 1 mol of phosphorus/mol of thiokinase. These studies show that the "dimer of dimers" structure of succinic thiokinase contains the expected two active sites and that this enzyme does not demonstrate "half-the-sites" reactivity. A coenzyme A binding site of lower affinity can be detected in some enzyme samples of lower specific activity. Binding of coenzyme A to the higher affinity sites may involve positive cooperativity. ATP, unlike ADP, does not bind to phosphorylated enzyme.