Iron-sulfur cluster redox chemistry and dimer dissociation in the outer mitochondrial membrane protein, mitoNEET.

The outer mitochondrial membrane protein known as mitoNEET was discovered when it was labeled by a photoaffinity derivative of the anti-diabetes medication, pioglitazone. The biological role for mitoNEET and its specific mechanism for achieving this remains an active subject for research. There is accumulating evidence suggesting that mitoNEET could be a component of mitochondrial FeS cofactor biogenesis.

Crystal structure of 12-lipoxygenase catalytic-domain-inhibitor complex identifies a substrate-binding channel for catalysis.

Lipoxygenases are critical enzymes in the biosynthesis of families of bioactive lipids including compounds with important roles in the initiation and resolution of inflammation and in associated diseases such as diabetes, cardiovascular disease, and cancer. Crystals diffracting to high resolution (1.9 Å) were obtained for a complex between the catalytic domain of leukocyte 12-lipoxygenase and the isoform-specific inhibitor, 4-(2-oxapentadeca-4-yne)phenylpropanoic acid (OPP).

A scaffold hopping approach to identify novel monoamine oxidase B inhibitors.

Monoamine oxidase B (MAO-B) inhibitors are used to treat Parkinson's disease. In this study, we searched for novel MAO-B inhibitors using a scaffold hopping approach based on our experience with the thiazolidinedione (TZD) class of compounds as MAO-B inhibitors. Several novel compounds were identified, with potencies in the low nanomolar and low micromolar range.

A novel binding assay identifies high affinity ligands to the rosiglitazone binding site of mitoNEET.

A novel outer mitochondrial membrane protein containing [2Fe-2S] clusters, mitoNEET was first identified through its binding to the anti-diabetic drug pioglitazone. Pioglitazone belongs to a family of drugs that are peroxisome proliferator-activated receptor (PPAR) gamma agonists, collectively known as glitazones. With the lack of pharmacological tools available to fully elucidate mitoNEET's function, we developed a binding assay to probe the glitazone binding site with the aim of developing selective and high affinity compounds.

Structure-activity relationship and docking studies of thiazolidinedione-type compounds with monoamine oxidase B.

The neuroprotective activity of pioglitazone and rosiglitazone in the MPTP parkinsonian mouse prompted us to evaluate a set of thiazolidinedione (TZD) type compounds for monoamine oxidase A and B inhibition activity. These compounds were able to inhibit MAO-B over several log units of magnitude (82 nM to 600 μM). Initial structure-activity relationship studies identified key areas to modify the aromatic substituted TZD compounds.

Complexes of the outer mitochondrial membrane protein mitoNEET with resveratrol-3-sulfate.

Binding of the thiazolidinedione antidiabetic drug pioglitazone led to the discovery of a novel outer mitochondrial membrane protein of unknown function called mitoNEET. The protein is homodimeric and contains a uniquely ligated two iron-two sulfur cluster in each of its two cytosolic domains. Electrospray ionization mass spectrometry was employed to characterize solutions of the soluble cytosolic domain (amino acids 32--108) of the protein.

Identification of novel monoamine oxidase B inhibitors by structure-based virtual screening.

Parkinson's disease is a severe debilitating neurodegenerative disorder. Recently, it was shown that the peroxisome proliferating-activator receptor-gamma agonist pioglitazone protected mice from 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine neurotoxicity due to its ability to inhibit monoamine oxidase B (MAO-B). Docking studies were initiated to investigate pioglitazone's interactions within the substrate cavity of MAO-B.

Structure-based design of a thiazolidinedione which targets the mitochondrial protein mitoNEET.

Several PPAR-gamma agonists containing a thiazolidinedione moiety (referred to as glitazones) have been proposed to be neuroprotective and appear to alter mitochondrial function. Recently, a search for mitochondrial proteins that bind pioglitazone identified a novel protein, mitoNEET, which was later shown to regulate the oxidative capacity of the mitochondria. This identified an alternative target for the glitazones suggesting a possible new drug target for the treatment of neurodegenerative diseases.

EPR spectroscopy and electrospray ionization mass spectrometry reveal distinctive features of the iron site in leukocyte 12-lipoxygenase.

The procedure for the expression and purification of recombinant porcine leukocyte 12-lipoxygenase using Escherichia coli [K.M. Richards, L.

Secondary alkyl hydroperoxides as inhibitors and alternate substrates for lipoxygenase.

Lipoxygenase plays a central role in polyunsaturated fatty acid metabolism, inaugurating the biosynthesis of eicosanoids in animals and phytooxylipins in plants. Redox cycling of the non-heme iron cofactor represents a critical element of the catalytic mechanism. Paradoxically, the isolated enzyme contains Fe(II), but the catalytically active form contains Fe(III), and the natural oxidant for the iron is the hydroperoxide product of the catalyzed reaction.

Electrospray ionization mass spectrometry of soybean lipoxygenases: N-terminal acetylation, chemical modification, and solution conformation.

Electrospray ionization mass spectrometry was used to examine both the covalent structure and solution conformation of the soybean lipoxygenases. The post-translational modifications of two lipoxgyenases were identified as N-terminal acetylations by tandem mass spectrometry of peptides generated by trypsin digestion. The N-terminal sequence suggests that the proteins were substrates for the plant homolog of the N-terminal acetyltransferase complex C in yeast.

Interaction between non-heme iron of lipoxygenases and cumene hydroperoxide: basis for enzyme activation, inactivation, and inhibition.

Lipoxygenase catalysis depends in a critical fashion on the redox properties of a unique mononuclear non-heme iron cofactor. The isolated enzyme contains predominantly, if not exclusively, iron(II), but the catalytically active form of the enzyme has iron(III). The activating oxidation of the iron takes place in a reaction with the hydroperoxide product of the catalyzed reaction.

Protein micelles from lipoxygenase 3.

Heat-induced conformational changes in lipoxygenase 3 were characterized by differential scanning calorimetry. The positions of the observed transitions were sensitive to the composition of the buffer. In particular, lipoxygenase 3 heated in carbonate buffer at pH 8.