Magnetic circular dichroism studies on the mononuclear ferrous active site of phthalate dioxygenase from Pseudomonas cepacia show a change of ligation state on substrate binding.

Phthalate dioxygenase from Pseudomonas cepacia contains a mononuclear ferrous center that is strictly required for catalytic oxygen activation. The spectroscopic characterization of this iron site and its ligand interactions has been complicated in the past by interference from a Rieske-type binuclear (2Fe-2S) cluster in the enzyme, which dominates the absorption spectra and is superimposed in X-ray absorption spectra for the mononuclear site. We have used low-temperature, variable magnetic field circular dichroism spectroscopy to selectively detect the ligand field spectra of the paramagnetic mononuclear ferrous active site in the presence of the diamagnetic exchange-coupled Rieske center and observe spectral changes associated with substrate binding. The perturbations of the d-->d spectra for the mononuclear ferrous site reflect a decrease in coordination number from six to five on binding substrate. This structural change suggests that displacement of an iron ligand prepares the ferrous center for dioxygen activation.