Amino acid sequences around the pyridoxal-5'-phosphate-binding sites of phenol hydroxylase

Phenol hydroxylase was labelled with pyridoxal 5'-phosphate. A radioactive label was introduced by using sodium boro[3H]hydride to reduce the initially formed Schiff's base. The labelled enzyme was digested with Staphylococcus V8 protease. Labelled peptides were isolated and their sequences were determined. The label could be located to three different lysyl residues. Sequence similarities with the known structures of p-hydroxybenzoate hydroxylase and glutathione reductase are discussed. The positions of the labelled sequences, relative to the bound ligands at the active site, are proposed on the basis of such sequence similarities.