The inhibition of caeruloplasmin by azide.

1. The inhibition of the oxidase activity of caeruloplasmin by azide was investigated at 25 degrees and 7.5 degrees . 2. The inhibition is reversible on dilution or Sephadex treatment, indicating a caeruloplasmin-azide complex. 3. The enzyme is protected against azide inhibition by chloride, acetate or EDTA, the last-named acting not by chelation but by a non-specific effect similar to that of acetate. 4. Lineweaver-Burk plots with different concentrations of azide are parallel. This may occur either when the enzyme-substrate complex or when a subsequent intermediate structure of the enzyme forms the inhibited complex. 5. At 7.5 degrees inhibition may be shown not to occur until after the initial reaction of enzyme with substrate. 6. At 7.5 degrees , the inhibition is of the mutual-depletion type, inhibitory concentrations of azide being comparable with the concentration of caeruloplasmin. It is shown that the binding of a single azide group completely inhibits a caeruloplasmin molecule. 7. An arrangement of the four valence-changing copper atoms of caeruloplasmin is proposed in which they are so close together in the cuprous form that reoxidation may occur by the simultaneous transfer of four electrons from the copper atoms to a single oxygen molecule.