Escherichia coli phosphofructokinase: inhibition by 8-anilino-1-naphthalenesulfonate.

Phosphofructokinase was purified 1200-fold from extracts of Escherichia coli B. Kinetic studies of the enzyme were carried out in the presence of the fluorescent dye 8-anilino-1-naphthalenesulfonate (1,8-ANS). 1,8-ANS was competitive with ATP and an uncompetitive inhibitor with respect to fructose-6-P. These parabolic inhibitions were accounted for by assuming that at least two molecules of the inhibitor were responsible for decreasing the affinity of the enzyme for ATP. ADP and GDP are both positive effectors for E. coli Phosphofructokinase. Evidence is presented to indicate that 1,8-ANS binding decreases the affinity of a regulatory site for ADP but not the binding site for regulation by GDP.