The role of His117 in the redox reactions of azurin from Pseudomonas aeruginosa.

ABSTRACT

The electron-transfer properties of H117G- and wild-type azurin were compared by applying both as electron acceptors in the conversion of 4-ethylphenol by 4-ethylphenol methylenehydroxylase (4-EPMH). The reactivity of H117G-azurin was determined in the absence and presence of imidazoles, which can substitute the missing fourth ligand. In the absence of imidazoles, H117G-azurin reacted directly with 4-ethylphenol; this reaction was abolished in the presence of imidazoles. The enzymatic reduction of H117G-azurin by 4-EPMH was 40 times slower than that of wild-type azurin. The rate of this reaction was enhanced by some imidazoles, diminished by others. In all cases the reduction of H117G-azurin was irreversible. These results demonstrate that His117 is vital for electron transfer and effectively protects the copper site against aspecific reactions.