Anion inhibition of firefly luciferase

The activity of firefly luciferase was shown to be very sensitive to the presence of salts in the assay mix. The general ionic strength effect was expressed by an increase in the Km for MgATP as the ionic strength was increased. When this effect was controlled by using concentrations of Mg2+ and ATP 10 times that normally saturating, a specific anion inhibition was observed. The order of effectiveness of inhibition by the anions, SCN− > I− ~ NO3− > Br− > Cl−, followed their position in the Hofmeister series. Only one anion was bound per active site and all the anions bind at this same site. Every reaction catalyzed by luciferase, even those proceeding by apparently different mechanisms, was affected in a similar manner by the anions. It was assumed that a small localized conformational change was occurring in the area of the active site upon the binding of the anions.