Kinetics of the allosteric interactions of phosphofructokinase from Escherichia coli

Phosphofructokinase has been partially purified from Escherichia coli, and its kinetic properties investigated. It shows co-operative interactions with respect to one of its substrates, fructose-6-phosphate, but not towards the second, namely ATP. ADP and other diphosphonucleosides act as activators and phosphoenolpyruvate as an inhibitor. Both effectors decrease the homotropic interactions between fructose-6-phosphate molecules; but, whereas the activators increase the affinity of the enzyme for this substrate, the inhibitor decreases it. These ligands have no effect on the maximum velocity of the reaction, except in the case of ADP which is a competitive inhibitor of ATP. These homotropic and heterotropic interactions are qualitatively and quantitatively accounted for by the concerted transition theory proposed by Monod, Wyman & Changeux (1965), assuming the enzyme to be in equilibrium between two conformational states which differ in their dissociation constants for fructose-6-phosphate, activators and inhibitor. A convenient method of obtaining these intrinsic dissociation constants has been derived from the equations of the theory. From the kinetic data, it is also possible to obtain the value of the equilibrium constant between the two states, if it is assumed that the enzyme is a tetramer made up of four identical subunits and that the transition is perfectly concerted.