Prior work based on crystallographic data and spectroscopic studies along with computational studies 120,121 of non-heme Fe(II) a-KG dependent hydroxylases and the related nonheme Fe(II) dioxygenases has led to the proposal of the following mechanism in the catalytic cycle of non-heme hydroxylases. In its resting state, the Fe(II) hexa-coordinated active site consists of three water molecules and a 2-His-1-carboxylate (Asp/Glu) facial triad where Fe(II) is bound to two His residues and an Asp/Glu residue (Supporting Information Figure ). The first step in the catalytic cycle is the binding of the bidentate aKG co-substrate to the iron center (1) which results in the displacement of two water molecules from the active site (Figure ). When the native substrate enters the binding pocket in the proximity of Fe(II), the remaining water molecule is also displaced, resulting in a five-coordinate square pyramidal geometry around iron (Supporting Information Figure ). This is followed by the binding of molecular oxygen (2) to iron, which immediately attacks the carbonyl carbon of the bidentate aKG ligand, leading to O-O bond cleavage and oxidative decarboxylation of the bidentate aKG (Figure and Supporting Information Figure ). This catalytic step results in the formation of a highly reactive terminal Fe(IV)=O intermediate (3) along with the succinate co-substrate, which has a carboxylate group that can bind iron in a monodentate or bidentate fashion (Figure ). ). This rate-determining hydrogen atom transfer (HAT) step is followed by recombination of the substrate radical with the hydroxyl group of the Fe(III)-OH intermediate during the radical rebound step of the catalytic cycle to form the product (5) (Figure and Supporting Information Figure ). After the release of the hydroxylated product and succinate and rebinding of three water molecules to iron, the active site goes back to its resting state (Supporting Information Figure ).