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Kinetic and structural investigations of the flavohemoglobin-type NO dioxygenase have suggested critical roles for transient Fe(III)O 2 complex formation and O 2 -forced movements affecting hydride transfer to the FAD cofactor and electron-transfer to the Fe(III)O 2 complex. Stark-effect theory together with structural models and dipole and internal electrostatic field determinations provided a semi-quantitative spectroscopic method for investigating the proposed Fe(III)O 2 complex and O 2 -forced movements. Deoxygenation of the enzyme causes Stark effects on the ferric heme Soret and charge-transfer bands revealing the Fe(III)O 2 complex. Deoxygenation also elicits Stark effects on the FAD that expose forces and motions that create a more restricted NADH access to FAD for hydride transfer and switch electron-transfer off. Glucose also forces the enzyme toward an off state. Amino acid substitutions at the B10, E7, E11, G8, D5, and F7 positions influence the Stark effects of O 2 on resting heme spin states and FAD consistent with the proposed roles of the side chains in the enzyme mechanism. Deoxygenation of ferric myoglobin and hemoglobin A also induces Stark effects on the hemes suggesting a common 'oxy-met' state. The ferric myoglobin and hemoglobin heme spectra are also glucose-responsive. A conserved glucose or glucose-6-phosphate binding site is found bridging the BC-corner and G-helix in flavohemoglobin and myoglobin suggesting novel allosteric effector roles for glucose or glucose-6-phosphate in the NO dioxygenase and O 2 storage functions. The results support the proposed roles of a ferric O 2 intermediate and protein motions in regulating electron-transfer during NO dioxygenase turnover. [Display omitted] • O 2 binds Fe(III) heme in flavohemoglobin as detected by Stark-effect spectroscopy. • O 2 also forces motion of the CD loop, FAD and electron-transfer switch. • Glucose binding opposes the O 2 -driven motions in flavohemoglobin. • O 2 binds Fe(III) heme in myoglobin and hemoglobin. • Glucose or glucose-6-phosphate bind myoglobin and presumably other globins. [ABSTRACT FROM AUTHOR]

Aqueous solutions of the four highest vanadium oxidation states exhibit four diverse colors, which only hint at the diverse reactions that these ions can undergo. Cationic vanadium ions form complexes with ligands; anionic vanadium ions form complexes with ligands and self-react to form isopolyanions. All vanadium species undergo oxidation –reduction reactions. With a few exceptions, elucidation of the dynamics of these reactions awaited the development of fast reaction techniques before the kinetics of elementary ligation, condensation, reduction, and oxidation of the aqueous vanadium ions could be investigated. As the biological roles played by endogenous and therapeutic vanadium expand, it is appropriate to bring the results of the diverse kinetics studies under one umbrella. To achieve this goal this review presents a systematic examination of elementary aqueous vanadium ion dynamics. [ABSTRACT FROM AUTHOR]

Abstract: Combined quantum mechanical and molecular mechanical (QM/MM) calculations were used to explore the electron pathway involved in the suicide inactivation of cytochrome P450BM3 from Bacillus megaterium. The suicide inactivation is a common phenomenon observed for heme peroxidases, in which the enzyme is inactivated as a result of self-oxidation mediated by highly oxidizing enzyme intermediates formed during the catalytic cycle. The selected model was a mutant comprising only the heme domain (CYPBM3 21B3) that had been previously evolved to efficiently catalyze hydroxylation reactions with hydrogen peroxide (H2O2) as electron acceptor. An extensive mapping of residues involved in electron transfer routes was obtained from density functional calculations on activated heme (i.e. Compound I) and selected amino acid residues. Identification of oxidizable residues (electron donors) was performed by selectively activating/deactivating different quantum regions. This method allowed a rational identification of key oxidizable targets in order to replace them for less oxidizable residues by site-directed mutagenesis. The residues W96 and F405 were consistently predicted by the QM/MM electron pathway to hold high spin density; single and double mutants of P450BM3 on these positions (W96A, F405L, W96A/F405L) resulted in a more stable variants in the presence of hydrogen peroxide, displaying a similar reaction rate than P450BM3 21B3. Furthermore, mass spectrometry confirmed these oxidation sites and corroborated the possible routes described by QM/MM electron transfer (ET) pathways. [Copyright &y& Elsevier]

Abstract: Schizosaccharomyces pombe (Sp) ferredoxin contains a C-terminal electron transfer protein ferredoxin domain (etpFd) that is homologous to adrenodoxin. The ferredoxin has been characterized by spectroelectrochemical methods, and Mössbauer, UV–Vis and circular dichroism spectroscopies. The Mössbauer spectrum is consistent with a standard diferric [2Fe–2S]2+ cluster. While showing sequence homology to vertebrate ferredoxins, the E°'' and the reduction thermodynamics for etpFd (−0.392V) are similar to plant-type ferredoxins. Relatively stable Cys to Ser derivatives were made for each of the four bound Cys residues and variations in the visible spectrum in the 380–450nm range were observed that are characteristic of oxygen ligated clusters, including members of the [2Fe–2S] cluster IscU/ISU scaffold proteins. Circular dichroism spectra were similar and consistent with no significant structural change accompanying these mutations. All derivatives were active in an NADPH-Fd reductase cytochrome c assay. The binding affinity of Fd to the reductase was similar, however, Vmax reflecting rate limiting electron transfer was found to decrease ~13-fold. The data are consistent with relatively minor perturbations of both the electronic properties of the cluster following substitution of the Fe-bond S atom with O, and the electronic coupling of the cluster to the protein. [Copyright &y& Elsevier]