Kinetics and Mechanism of Dissociation of Zinc Ion from Carbonic Anhydrase

Abstract

The kinetics of dissociation of Zn²⁺ from the metalloenzyme carbonic anhydrase was measured over a range of pH, temperature, and acetate concentration. The rate of dissociation is extremely slow at neutral pH (t_l/2 ≈ 3 years, 4°C), but increases in almost direct proportion to the hydrogen ion concentration and is enhanced in the presence of 1,10-phenanthroline or acetate. The thermodynamic stability of the zinc-apoenzyme complex was determined over a range of pH from rate data on binding and dissociation (stability constants 10⁹–10¹¹ M⁻¹, 25°C). The great stability of the complex and slow exchange of the apoenzyme ligand is attributed, at least in part, to the rigidity of the multidentate protein ligand.