Alpha-ketoglutarate-dependent hydroxylases

Alpha-ketoglutarate-dependent hydroxylases are a major class of non-heme iron proteins that catalyse a wide range of reactions.

[4][5] In microorganisms such as bacteria, αKG-dependent dioxygenases are involved in many biosynthetic and metabolic pathways;[6][7][8] for example, in E. coli, the AlkB enzyme is associated with the repair of damaged DNA.

[18] Many αKG-dependent dioxygenase also catalyse uncoupled turnover, in which oxidative decarboxylation of αKG into succinate and carbon dioxide proceeds in the absence of substrate.

αKG coordinates as a bidentate ligand to Fe(II), while the substrate is held by noncovalent forces in close proximity.

The uncoordinated end of the superoxide ligand attacks the keto carbon, inducing release of CO2 and forming an Fe(IV)-oxo intermediate.

[36][37][38] Finally, as αKG-dependent dioxygenases require molecular oxygen as a co-substrate, it has also been shown that gaseous molecules such as carbon monoxide[39] and nitric oxide[40][41] are inhibitors of αKG-dependent dioxygenases, presumably by competing with molecular oxygen for the binding at the active site Fe(II) ion.

[42] For example, assays were developed to study ligand binding,[43][44][45] enzyme kinetics,[46] modes of inhibition[47] as well as protein conformational change.

[56] Other biophysical techniques including (but not limited to) isothermal titration calorimetry (ITC)[57] and electron paramagnetic resonance (EPR) were also applied.

Consensus catalytic mechanism of the αKG-dependent dioxygenase superfamily.
Simplified view of the active site of prolyl hydroxylase isoform 2 (PHD2), a human αKG-dependent dioxygenase. The Fe(II) is coordinated by two imidazoles and one carboxylate provided by the protein. Other ligands on iron, which are transiently occupied αKG and O 2 , are omitted for clarity.
Common inhibitors of αKG-dependent dioxygenases. They compete against the cosubstrate αKG for binding to the active site Fe(II).