Cyanogen-induced gamma-glutamyl to imidazole cross-link in carbonic anhydrase. A unique mode of inhibition.

The irreversible inhibition of carbonic anhydrase by cyanogen occurs by a unique mechanism. Cyanogen is an affinity label: it behaves like a carbodiimide and produces an intra-molecular cross-link without being incorporated. The nucleophile-labile cross-link is formed between a gamma-COOH of a Glu and an imidazole of a His with a 1:1:1 stoichiometry with the enzyme. The deletion of approximately 1 Glu and approximately 1 His was noted by amino acid analysis of enzymatically hydrolyzed carbonic anhydrase. The modified Glu was converted to 2,4-diaminobutanoic acid and quantitated by amino acid analysis. The presence and quantity of modified His was supported through high-resolution proton NMR analysis.