The interaction of Cr-II with taurine/alpha-ketoglutarate (alpha KG) dioxygenase (TauD) was examined. Cr-II replaces Fe-II and binds stoichiometrically with alpha KG to the Fe-II/alpha KG binding site of the protein, with additional Cr-II used to generate a chromophore attributed to a Cr-III-semiquinone in a small percentage of the sample. Formation of the latter oxygen-sensitive species requires the dihydroxyphenylalanine (DOPA) quinone form of Tyr-73. This preformed side chain is generated by intracellular self-hydroxylation of Tyr-73 to form DOPA, which is subsequently oxidized to the quinone. No chromophore is generated when using NaBH4-treated sample, protein isolated from anaerobically grown cells, inactive TauD variants that are incapable of self-hydroxylation, or the Y73F active mutant of TauD. A Cr-III-DOPA semiquinone also was observed in the herbicide hydroxylase SdpA.