Inositol oxygenase

Protein coding gene in the species Homo sapiens

Inositol oxygenase, also commonly referred to as myo inositol oxygenase (MIOX), is a non heme di iron enzyme that oxidizes myo inositol to glucuronic acid. The enzyme employs a unique four electron transfer at its Fe(II)/Fe(III) coordination sites and the reaction proceeds through the direct binding of myo inositol followed by attack of the iron center by diatomic oxygen. This enzyme is part of the only known pathway for the catabolism of inositol in humans and is expressed primarily in the kidneys. Recent medical research regarding MIOX has focused on understanding its role in metabolic and kidney diseases such as diabetes, obesity and acute kidney injury. Industrially focused engineering efforts are centered on improving MIOX activity in order to produce glucaric acid in heterologous hosts.

Myo inositol oxygenase is a monomeric 33 kDa protein in both solution and crystal. This enzyme possesses a Fe(II)/Fe(III) atomic pair at the catalytic active site which enables its unique four electron transfer mechanism. Recent crystallization studies have elucidated the structures of the mouse MIOX in 2008. The overall structure of the mouse MIOX is primarily helical with five alpha helices forming the core of the protein. The two iron centers are coordinated by various amino acids and water molecules as shown in complex with the myo inositol substrate. The human MIOX structure superimposes closely onto the mouse MIOX structure, sharing 86% sequence identity over the structural alignment but with some differences in the residues surrounding the active site. Instead, MIOX closely resembles proteins in the HD domain superfamily based on its highly conserved metal binding strategy and the presence of the four His ligands on the iron center. A series of crystallization, spectroscopy and density functional theory experiments have revealed a putative mechanism (shown right) for the oxidation of myo inositol. ENDOR spectroscopy was used to determine that the substrate directly binds to the Fe(II)/Fe(III) di iron center of MIOX most likely through the O1 atom of myo inositol. In the kidney, MIOX converts myo inositol to glucuronic acid which is then able to enter the glucuronate xylulose pathway for conversion to xylulose 5 phosphate. Additionally, a recent study has shown that MIOX is upregulated in the diabetic state with its transcription heavily regulated by osmolarity, glucose levels and oxidative stress. This upregulation is associated with the formation of reactive oxidative species that lead to interstitial injury in the kidney. An immunoassay of MIOX expression may potentially predict these life threatening injuries earlier than the current diagnostic—detection of plasma creatinine.

The MIOX enzyme has been the object of intense metabolic engineering efforts to produce glucaric acid through biosynthetic pathways. In 2004, the U. S. Department of Energy released a list of the top value added chemicals from biomass which included glucaric acid—the direct product of the oxidation of glucuronic acid. The first biosynthetic production of glucaric acid was achieved in 2009 with use of the uronate dehydrogenase (UDH) enzyme. Since then, the MIOX enzyme has been engineered for improved glucaric acid production through numerous strategies including appendage of an N terminal SUMO tag, directed evolution and also the use of modular, synthetic scaffolds to increase its effective local concentration.