Conformation-associated anomalous tyrosine fluorescence of adrenodoxin.

As reported previously (Kimura, T., Ting, J., and Huang, J. J. (1972) J Biol. Chem., 247, 4476-4479), adrenal iron-sulfur protein, which contains no tryptophan and 1 tyrosine residue at position 82, displays an anomalous fluorescence peak at 331 nm. The present study conclusively demonstrates that he emission originates from the tyrosine residue with no contribution to the fluorescence of tryptophan by excluding it as a contaminant. Upon removal of iron and labile sulfur by dialyzing the native protein against 5% (w/v) trichloroacetic acid, the apoprotein exhibited similar emission and excitation spectra as the native protein. When the protein was treated with guanidine HC1, urea, or LiCl, the normal fluorescence emission peak was observed at 305 nm. Upon the removal of denaturants by prolonged dialysis, the anomaly was restored. The treatment of the native protein at 80°C, or at pH 2.6, normalized the fluorescence emission peak to 305 nm. From these results, we came to the following conclusions: 1) The anomalous fluorescence of the tyrosine residue is independent of the iron-sulfur center; 2) the anomaly is related to the protein conformation in some manner; and 3) the fluorescence change from the anomalous state to the normal state is reversible, suggesting that conformation for the tyrosine environment in adrenodoxin is closely related to the anomaly.