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The basis for the interdomain interaction was studied by constructing a heterologous system in which the C domain of Enteromorpha intestinalis (EiAPR) was fused to the carboxyl terminus of the APS reductase from Pseudomonas aeruginosa (PaAPR). The results suggest that the physical connection of the reductase and C domain on a single polypeptide is critical for the electron-transfer reaction.

5'-adenylyl sulfate (APS) reductase from the marine macrophytic green alga Enteromorpha intestinalis uses reduced glutathione as the electron donor for the reduction of APS to 5'-AMP and sulfite. It is proposed that the Cys342- Cys345 disulfide provides the site for entry of electrons from reduced glutathione and that the Cys166- Cys285 disulfide serves as a structural element that is essential for keeping the enzyme in the catalytically active conformation.

The structure of nitrite reductase, a key enzyme in the process of nitrogen assimilation is determined using X-ray diffraction to a resolution limit of 2.8 A. However, the structure of nitrite reductase shows a broad similarity to hemoprotein subunit of sulfite reductase but has many significant differences in the backbone positions that reflect sequence differences or could arise alterations of the sulfite reductase structure that arise from the isolation of the subunit from the native complex.

Thiol determinations and site-directed mutagenesis studies indicate that the single [4Fe-4S] cluster contains only three cysteine ligands, instead of the more typical arrangement in which clusters are bound to the protein by four cysteines. Site directed mutagenesis experiments have identified Cys139, Cys228, and Cys231 as ligands to the cluster.

Research has been conducted on sulfur bacterium Chromatium vinosum's reaction center. The role of C. vinosum growth conditions in electron donation to the reaction center has been investigated and the details are reported.

Research has been conducted on the chloroplast fructose 1,6-bisphosphatase (FBPase). The structural and functional properties of the conserved cysteines of the FBPase chloroplast loop have been investigated via the replacement of these cysteines by the site-directed mutagenesis and the analysis of their thiol content, Mg (super) 2+ dependence and redox potentials.

Research has been conducted on the 5'-adenylyl sulfate (APS) reductase that catalyzes a plant sulfate reaction. The results of the biochemical studies indicate that the enzyme product derived from one of the APS reductase encoding genes is activated by oxidation via disulfide bond formation.

The Rhodobacter capsulatus cytochrome c biogenesis system has been studied with emphasis on oxidation-reduction properties of its disulfide-containing proteins. Thermodynamic support was found for the point of view that HelX reduces Ccl2 and that reduced Cc12 serves as the reductant for production of the two thiols of the CysXxxYyyCysHis heme-binding motif of the apocytochromes.

Research has been conducted on the sorghum nicotinamide adenine dinucleotide phosphate-dependent dehydrogenase reductive activation. The measurements of the oxidation-reduction midpoint potentials are presented.

A study was conducted on the function of the ferredoxin/thioredoxin system of oxygenic photosynthetic organisms in their carbon metabolism regulation. The study reports on values derived for reduction midpoint potentials that are measured over a range of pH values for thioredoxins f from spinach and pea, for thioredoxins m from spinach and the green alga Chlamydomonas reinhardtii, for spinach ferrodoxin:thioredoxin reductase, for spinach phosphoribulokinase and for spinach and pea fructose-1,6-biphosphatases.

The role of the [Fe4S4] cluster in disulfide reduction in spinach ferredoxin: thioredoxin reductase was investigated. Preliminary freeze-quench electron paramagnetic resonance spectroscopy was conducted on ferredoxin:thioredoxin reductase reduced with stoichiometric concentrations of reduced benzyl viologen. Results show that the [Fe4S4] cluster plays a direct role in cleaving the active-site disulfide in ferredoxin:thioredoxin reductase and that this function possibly involves S-based cluster chemistry.

The roles of iron-sulfur clusters in the chloroplast ferredoxin/thioredoxin reductase (FTR) from spinach were investigated by spectroscopic experiments. A cluster that cannot be reduced but can be oxidized by ferricyanide was identified and was found to mediate electron transport. The results of parallel studies of a modified form of the FTR system confirmed the role of this cluster, suggesting that the iron-sulfur center in spinach FTR may have a biological role.

Electron transfer proteins are isolated and characterized from the marine purple-sulfur bacterium Chromatium purpuratum. Redox titrations of the flavocytochrome c hemes indicate two components with midpoint potential values of +15 and -120 mV, which is similar to that of the flavocytochrome isolated from the thermophilic Chromatium tepidum. There is also substantial homology to the main structure of the flavocytochrome c of Chromatium vinosum. In contrast, the C. purpuratum high-potential iron-sulfur protein differs from those isolated from other photosynthetic bacteria.

The effects of ionic strength on electrontransfer rates from an ubiquinol analogue to cytochrome (cyt) c2 catalyzed by Rhodobacter capsulatus and Rhodopseudomonas (Rps) viridis cyt bc1 complexes were examined. Results revealed that ionic strength is an important factor in the formation of an electrostatic complex between cyt c2 and cyt bc1 complex in the electron transfer pathway. Evidence was also obtained for a docking model where positively charged cyt c2 lysines interact with negatively charged groups on the cyt bc1 complex. A significant difference was found between the Rps viridis cyt c2 docking site on the Rps viridis reaction center and that for the cyt on the Rps viridis cyt bc1 complex.

The solution structure of Synechocystis Ga-substituted ferredoxin (Fd) was determined and compared with the structure of the native protein to establish if the Fd cluster replaced by Ga could act as suitable diamagnetic mimic. Comparison of the NMR structure of GaFd with the crystal structure of the native Fd indicated that the general structural fold found for the native, iron-containing ferredoxin is conserved in GaFd.

The paramagnetic NMR spectroscopy technique is employed to explain the redox chain comprising ferredoxin (Fd), ferredoxin:thioredoxin reductase (FTR), and thioredoxin (Trx). Two different models of the noncovalent binary complex are also presented.

Kinetic analysis of an electron transfer cascade involving photoexcited photosystem I and ferredoxin was used to characterize nitrite reductase, which reduces nitrite to ammonium in a six-electron reaction. Spectral and kinetic analyses of single reduction of the recombinant enzyme in the ferrous siroheme-NO* state by photoreduced ferredoxin indicated that the process leads to reduction of the [4Fe-4S] cluster with minor NO* reduction and the enzyme must wait for the next reduction step before NO* undergoes substantial reduction.

The thermodynamic basis of the disulfide/dithiol couples are examined in the pathway of the Yap1 oxidative stress signal transduction found in Saccharomyces cerevisiae. It is observed that the oxidized basis form of the Yap1 redox domain fragment, derived from the Yap1 transcription factor contains two disulfide bonds, one between Cys303 and Cys598 and one between Cys310 and Cys629.

Plant nitrite reductase (NiR) catalyzes the reduction of nitrite to ammonia, using reduced ferredoxin as the electron donor. Resonance Raman and absorption spectroscopy data show that the reaction of oxidized NiR ([4Fe-4S](super 2+)/Fe(super 3+)) with hydroxylamine is binding-limited, while the NH2OH conversion to nitric oxide is much faster.

The enzyme catalytic cycle with ferredoxin reduced by photosystem 1 as an electron donor is examined by electron paramagnetic resonance (EPR0 and laser flash absorption spectroscopy. A complex of ferrous siroheme with NO is the most prominent and stable intermediate observed in the enzyme catalytic cycle.

The study reveals that for wild-type thioredoxins from Escherichia coli and from green alga Chlamydomonas reinhardtii and for a number of site-directed mutants of these two thioredoxins, oxidation-reduction midpoint potential (Em) versus pH profiles were measured. It also determines that the effect of site-directed mutagenesis of two highly conserved aspartate residues plays an important structural and/or catalytic roles in both thioredoxins, and it could also play a role in proton transfer, on the pKa values of redox-linked acid dissociations for both E. coli thioredoxin and C. reinhardtii thioredoxin h.

Research has been conducted on cytochrome c (sub)1 from Rhodobacter capsulatus. The authors report that the conserved aromatic residue is important for the stabilization of native heme environment of this cytochrome.

Two novel R. capsulatus cyt c1 mutants were utilized to assess the role played by the sixth axial ligand in the case of the cyt c1 subunit of the bc1 complex. Analysis of the the two mutants, namely M183K and M183H, shows that they incorporate into the mature cyt c1 polypeptides the heme group. Such mutants produced nonfunctional bc1 complexes with unusual thermodynamics and spectroscopic properties, including decreased redox midpoint potential values and shifted optical absorption maxima.