Evidence for the slow reaction of hypoxia-inducible factor prolyl hydroxylase 2 with oxygen.

The response of animals to hypoxia is mediated by the hypoxia-inducible transcription factor. Human hypoxia-inducible factor is regulated by four Fe(II)- and 2-oxoglutarate-dependent oxygenases: prolyl hydroxylase domain enzymes 1-3 catalyse hydroxylation of two prolyl-residues in hypoxia-inducible factor, triggering its degradation by the proteasome. Factor inhibiting hypoxia-inducible factor catalyses the hydroxylation of an asparagine-residue in hypoxia-inducible factor, inhibiting its transcriptional activity.

SufR coordinates two [4Fe-4S]2+, 1+ clusters and functions as a transcriptional repressor of the sufBCDS operon and an autoregulator of sufR in cyanobacteria.

The sufR gene encodes a protein that functions as a transcriptional repressor of the suf regulon in cyanobacteria. It is predicted to contain an N-terminal helix loop helix DNA binding motif and a C-terminal Fe/S binding domain. Through immunoblotting assays of cell extracts, the sufR product in Synechocystis sp.

A manganese(IV)/iron(IV) intermediate in assembly of the manganese(IV)/iron(III) cofactor of Chlamydia trachomatis ribonucleotide reductase.

We recently showed that the class Ic ribonucleotide reductase from the human pathogen Chlamydia trachomatis uses a Mn(IV)/Fe(III) cofactor to generate protein and substrate radicals in its catalytic mechanism [Jiang, W., Yun, D., Saleh, L.

A manganese(IV)/iron(III) cofactor in Chlamydia trachomatis ribonucleotide reductase.

In a conventional class I ribonucleotide reductase (RNR), a diiron(II/II) cofactor in the R2 subunit reacts with oxygen to produce a diiron(III/IV) intermediate, which generates a stable tyrosyl radical (Y*). The Y* reversibly oxidizes a cysteine residue in the R1 subunit to a cysteinyl radical (C*), which abstracts the 3'-hydrogen of the substrate to initiate its reduction. The RNR from Chlamydia trachomatis lacks the Y*, and it had been proposed that the diiron(III/IV) complex in R2 directly generates the C* in R1.

Reaction of cytochrome P450BM3 and peroxynitrite yields nitrosyl complex.

Peroxynitrite has come into the spotlight in recent years. Its effects on proteins have been implicated in several diseases such as acute lung injury, rheumatoid arthritis, implant rejection, artherosclerosis, Parkinson's disease, and Alzheimer's disease. Peroxynitrite is thought to inactivate a variety of proteins including thiolate-ligated heme proteins such as cytochrome P450 2B1 and PGI2 synthase, through the nitration of tyrosine residues.

Evidence for basic ferryls in cytochromes P450.

Using a combination of Mössbauer spectroscopy and density functional calculations, we have determined that the ferryl forms of P450(BM3) and P450cam are protonated at physiological pH. Density functional calculations were performed on large active-site models of these enzymes to determine the theoretical Mössbauer parameters for the ferryl and protonated ferryl (Fe(IV)OH) species. These calculations revealed a significant enlargement of the quadrupole splitting parameter upon protonation of the ferryl unit.

Evidence for two ferryl species in chloroperoxidase compound II.

Using a combination of density functional calculations and Mössbauer spectroscopy, we have examined chloroperoxidase compound II (CPO-II). The Mössbauer spectrum of CPO-II suggests the presence of two distinct ferryl species in an approximately 70:30 ratio. Density functional calculations and cryogenic reduction and annealing experiments allow us to assign the major species as an Fe(IV)OH intermediate.

Oxygen activation by a mixed-valent, diiron(II/III) cluster in the glycol cleavage reaction catalyzed by myo-inositol oxygenase.

myo-Inositol oxygenase (MIOX) catalyzes the ring-cleaving, four-electron oxidation of its cyclohexan-(1,2,3,4,5,6-hexa)-ol substrate (myo-inositol, MI) to d-glucuronate (DG). The preceding paper [Xing, G., Hoffart, L.

A coupled dinuclear iron cluster that is perturbed by substrate binding in myo-inositol oxygenase.

myo-Inositol oxygenase (MIOX) uses iron as its cofactor and dioxygen as its cosubstrate to effect the unique, ring-cleaving, four-electron oxidation of its cyclohexan-(1,2,3,4,5,6-hexa)-ol substrate to d-glucuronate. The nature of the iron cofactor and its interaction with the substrate, myo-inositol (MI), have been probed by electron paramagnetic resonance (EPR) and Mössbauer spectroscopies. The data demonstrate the formation of an antiferromagnetically coupled, high-spin diiron(III/III) cluster upon treatment of solutions of Fe(II) and MIOX with excess O(2) or H(2)O(2) and the formation of an antiferromagnetically coupled, valence-localized, high-spin diiron(II/III) cluster upon treatment with either limiting O(2) or excess O(2) in the presence of a mild reductant (e.

Evidence for C-H cleavage by an iron-superoxide complex in the glycol cleavage reaction catalyzed by myo-inositol oxygenase.

myo-Inositol oxygenase (MIOX) activates O2 at a mixed-valent nonheme diiron(II/III) cluster to effect oxidation of its cyclohexan-(1,2,3,4,5,6-hexa)-ol substrate [myo-inositol (MI)] by four electrons to d-glucuronate. Abstraction of hydrogen from C1 by a formally (superoxo)diiron(III/III) intermediate was previously proposed. Use of deuterium-labeled substrate, 1,2,3,4,5,6-[2H]6-MI (D6-MI), has now permitted initial characterization of the C-H-cleaving intermediate.

Kinetic dissection of the catalytic mechanism of taurine:alpha-ketoglutarate dioxygenase (TauD) from Escherichia coli.

Recent studies on taurine:alpha-ketoglutarate dioxygenase (TauD) from Escherichia coli have provided evidence for a three-step, minimal kinetic mechanism involving the quaternary TauD.Fe(II).alpha-ketoglutarate.