X-ray Absorption Spectroscopic Investigation of the Resting Ferrous and Cosubstrate-Bound Active Sites of Phenylalanine Hydroxylase

Previous studies of ferrous wild-type phenylalanine hydroxylase, {Fe2+}PAHT[], have shown the active site to be a six-coordinate distorted octahedral site. After the substrate and cofactor bind to the enzyme ({Fe2+}PAHR[l-Phe,5-deaza-6-MPH4]), the active site converts to a five-coordinate square pyramidal structure in which the identity of the missing ligand had not been previously determined. X-ray absorption spectroscopy (XAS) at the Fe K-edge further supports this coordination number change with the binding of both cosubstrates to the enzyme, and determines this to be due to the loss of a water ligand.