Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “FERREDOXINS”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 55 records · Page 3Linked to original sources
PubMed2000

Apicomplexan parasites possess distinct nuclear-encoded, but apicoplast-localized, plant-type ferredoxin-NADP+ reductase and ferredoxin.

In searching for nuclear-encoded, apicoplast-localized proteins we have cloned ferredoxin-NADP(+) reductase from Toxoplasma gondii and a [2Fe-2S] ferredoxin from Plasmodium falciparum. This chloroplast-localized redox system has been extensively studied in photosynthetic organisms and is responsible for the electron transfer from photosystem I to NADP+. Besides this light-dependent reaction in nonphotosynthetic plastids (e.g. from roots), electrons can also flow in the reverse direction, from NADPH to ferredoxin, which then serves as an important reductant for various plastid-localized enzymes. These plastids possess related, but distinct, ferredoxin-NADP+ reductase and ferredoxin isoforms for this purpose. We provide phylogenetic evidence that the T. gondii reductase is similar to such nonphotosynthetic isoforms. Both the P. falciparum [2Fe-2S] ferredoxin and the T. gondii ferredoxin-NADP+ reductase possess an N-terminal bipartite transit peptide domain typical for apicoplast-localized proteins. The recombinant proteins were obtained in active form, and antibodies raised against the reductase recognized two bands on Western blots of T. gondii tachyzoite lysates, indicative of the unprocessed and native form, respectively. We propose that the role of this redox system is to provide reduced ferredoxin, which might then be used for fatty acid desaturation or other biosynthetic processes yet to be defined. Thus, the interaction of these two proteins offers an attractive target for drug intervention.

Amino Acid Sequence↗
PubMed1977

On the role of ferredoxin and ferredoxin-NADP+ reductase in cyclic electron transport of spinach chloroplasts.

Antibodies prepared against purified spinach ferredoxin and ferredoxin-NADP+ reductase were used as specific inhibitors of electron-transfer reactions dependent on either ferredoxin or ferredoxin-NADP+ reductase; The possible role of both electron carriers in cyclic electron transport was checked using cytochrome b6 photoreactions as indicator. It could be demonstrated that the ferredoxin antibody inhibits cytochrome b6 photoreduction. Ferredoxin-NADP+ reductase, however, appears not to be involved in this pathway: reductase antibody in concentrations sufficient to completely inhibit electron transport to NADP+ had no effect on cytochrome b6 photoreduction. Quantitative treatment of the immunoassay data showed that osmotically shocked chloroplasts contain both bound ferredoxin and ferredoxin-NADP+ reductase in concentration approximately equal to that of cytochrome b6.

Chloroplasts↗
PubMed2000

Structural alignment of ferredoxin and flavodoxin based on electrostatic potentials: implications for their interactions with photosystem I and ferredoxin-NADP reductase.

The two proteins ferredoxin and flavodoxin can replace each other in the photosynthetic electron transfer chain of cyanobacteria and algae. However, structure, size, and composition of ferredoxin and flavodoxin are completely different. Ferredoxin is a small iron-sulfur protein (approximately 100 amino acids), whereas flavodoxin is a flavin-containing protein (approximately 170 amino acids). The crystal structure of both proteins from the cyanobacteria Anabeana PCC 7120 is known. We used these two protein structures to investigate the structural basis of their functional equivalence. We apply the Hodgkin index to quantify the similarity of their electrostatic potentials. The technique has been applied successfully in indirect drug design for the alignment of small molecule and bioisosterism elucidation. It requires no predefined atom-atom correspondences. As is known from experiments, electrostatic interactions are most important for the association of ferredoxin and flavodoxin with their reaction partners photosystem I and ferredoxin-NADP reductase. Therefore, use of electrostatic potentials for the structural alignment is well justified. Our extensive search of the alignment space reveals two alignments with a high degree of similarity in the electrostatic potential. In both alignments, ferredoxin overlaps completely with flavodoxin. The active sites of ferredoxin and flavodoxin rather than their centers of mass coincide in both alignments. This is in agreement with electron microscopy investigations on photosystem I cross-linked to ferredoxin or flavodoxin. We identify residues that may have the same function in both proteins and relate our results to previous experimental data.

Algal Proteins↗
PubMed1991

Site-specific mutations in human ferredoxin that affect binding to ferredoxin reductase and cytochrome P450scc.

Ferredoxins found in animal mitochondria function in electron transfer from NADPH-dependent ferredoxin reductase (Fd-reductase) to cytochrome P450 enzymes. To identify residues involved in binding of human ferredoxin to its electron transfer partners, neutral amino acids were introduced in a highly conserved acidic region (positions 68-86) by site-directed mutagenesis of the cDNA. Mutant ferredoxins were produced in Escherichia coli, and separate assays were used to determine the effect of substitutions on the capacity of each mutant to bind to Fd-reductase and cytochrome P450scc and to participate in the cholesterol side chain cleavage reaction. Replacements at several positions (mutants D68A, E74Q, and D86A) did not significantly affect activity, suggesting that acidic residues at these positions are not required for binding or electron transfer interactions. In contrast, substitutions at positions 76 and 79 (D76N and D79A) caused dramatic decreases in activity and in the affinity of ferredoxin for both Fd-reductase and P450scc; this suggests that the binding sites on ferredoxin for its redox partners overlap. Other substitutions (mutants D72A, D72N, E73A, E73Q, and D79N), however, caused differential effects on binding to Fd-reductase and P450scc, suggesting that the interaction sites are not identical. We propose a model in which Fd-reductase and P450scc share a requirement for ferredoxin residues Asp-76 and Asp-79 but have other determinants that differ and play an important role in binding. This model is consistent with the hypothesis that ferredoxin functions as a mobile shuttle in steroidogenic electron transfer, and it is considered unlikely that a functional ternary complex is formed.

Amino Acid Sequence↗
PubMed1984

A cross-linked complex between ferredoxin and ferredoxin-NADP+ reductase.

The water-soluble carbodiimide, N-ethyl-3-(3-dimethylaminopropyl)carbodiimide was found to effectively cross-link ferredoxin to ferredoxin-NADP+ reductase. The covalent complex has a stoichiometry of 1 mol of ferredoxin per mol of the reductase. The flavoprotein moiety of the cross-linked complex maintains most of its diaphorase activity and more interestingly has gained the capacity to catalyze the NADPH-cytochrome c reaction without addition of free ferredoxin in the assay mixture. Furthermore, the cross-linked complex binds NADP+ with a Kd = 88 microM at an ionic strength of 0.02 M. These results show that a ternary complex among the reductase and its substrates can be formed, suggesting that the binding sites for ferredoxin and the pyridine nucleotides are distinct. The bound ferredoxin can interact with cytochrome c; the iron-sulfur cluster of the cross-linked complex is shown to be reduced under anaerobic conditions by NADPH and to be required for the catalysis of the NADPH-cytochrome c reductase reaction. The cross-linked complex, added to thylakoids inhibited by the antibody against the reductase, catalyzes the H2O-cytochrome c photoreduction, which suggests that the ferredoxin moiety of the complex can interact with its electron donor in the photosynthetic chain. Restoration of NADP+ photoreduction requires the addition of free ferredoxin.

Carbodiimides↗
PubMed1981

Ferredoxin and ferredoxin reductase activities in bovine thyroid. Possible relationship to iodotyrosine deiodinase.

NADPH-ferredoxin reductase and ferredoxin activities have been identified in bovine thyroid particulate fractions (mainly mitochondria) after ultrasonication and DEAE-cellulose chromatography. The proteins were identified by their ability to reconstitute NADPH-cytochrome c reductase activity when used in combination. NADPH ferredoxin reductase and ferredoxin also catalyzed NADPH-dependent deiodination of L-diiodotyrosine; bovine adrenodoxin and adrenodoxin reductase could partially replace the thyroidal components in NADPH-dependent deiodination of L-diiodotyrosine. Both these reconstitutive activities were substantially inhibited by the iron chelators, alpha, alpha'-dipyridyl and o-phenanthroline. Deiodination by the NADPH-ferredoxin reductase-ferredoxin system was inhibited by the addition of a previously characterized dithionite-responsive flavoprotein iodotyrosine deiodinase (preparation F), isolated and purified from bovine thyroid particulate fractions after solubilization with steapsin. Ferredoxin reductase alone showed dithionite-responsive deiodinase activity and elution profiles of this activity on gel filtration before and after steapsin treatment suggest that preparation F may be a form of ferredoxin reductase modified by steapsin.

Animals↗
PubMed2002

N-isotope effects on the Raman spectra of Fe(2)S(2) ferredoxin and Rieske ferredoxin: evidence for structural rigidity of metal sites.

The diiron ferredoxins have a common diamond-core structure with two bridging sulfides, but differ in the nature of their terminal ligands: either four cysteine thiolates in the Fe(2)S(2) ferredoxins or two cysteine thiolates and two histidine imidazoles in the Rieske ferredoxins. Contributions of the bridging (b) and terminal (t) ligands to the resonance Raman spectra of the Fe(2)S(2) ferredoxins have been distinguished previously by isotopic substitution of the bridging sulfides. We now find that uniform (15)N-labeling of Anabaena Fe(2)S(2) ferredoxin results in shifts of -1 cm(-1) in the Fe-S(t) stretching modes at 282, 340, and 357 cm(-1). The (15)N dependence is ascribed to kinematic coupling of the Fe-S(Cys) stretch with deformations of the cysteine backbone, including the amide nitrogen. No (15)N dependence occurs for the nu(Fe-S(b)) modes at 395 and 426 cm(-1). Similar effects are observed for the Rieske center in T4MOC ferredoxin from the toluene-4-monooxygenase system of Pseudomonas mendocina. Upon selective (15)N-labeling of the alpha-amino group of cysteine, the vibrational modes at 321, 332, 350, and 362 cm(-1) all undergo shifts of -1 to -2 cm(-1), thereby identifying them as combinations of nu(Fe-S(t)) and delta(Cys). These same four modes undergo similar isotope shifts when T4MOC ferredoxin is selectively labeled with (15)N-histidine ((15)N in either the alpha1,delta1 or delta1,epsilon2 positions). Thus, the Fe-S(Cys) stretch must also be undergoing kinematic coupling with vibrations of the Fe-His moiety. The extensive kinematic coupling of iron ligand vibrations observed in both the Fe(2)S(2) and Rieske ferredoxins presumably arises from the rigidity of the protein framework and is reminiscent of the behavior of cupredoxins. In both cases, the structural rigidity is likely to play a role in minimizing the reorganization energy for electron transfer.

Anabaena↗
PubMed1998

Ferredoxin reduction by photosystem I from Synechocystis sp. PCC 6803: toward an understanding of the respective roles of subunits PsaD and PsaE in ferredoxin binding.

The process of ferredoxin reduction by photosystem I has been extensively investigated by flash-absorption spectroscopy in psaD and psaE deleted mutants from Synechocystis sp. PCC 6803. In both mutants, the dissociation constant for the photosystem I/ferredoxin complex at pH 8 is considerably increased as compared to the wild type: approximately 25- and 100-fold increases are found for PsaD-less and PsaE-less photosystem I, respectively. However, at high ferredoxin concentrations, submicrosecond and microsecond kinetics of electron transfer similar to that observed in the wild type are present in both mutants. The presence of these fast kinetic components indicates that the relative positions of ferredoxin and of the terminal photosystem I acceptor are not significantly disturbed by the absence of either PsaD or PsaE. The second-order rate constant of ferredoxin reduction is lowered 10- and 2-fold for PsaD-less and PsaE-less photosystem I, respectively. Assuming a simple binding equilibrium between photosystem I and ferredoxin, PsaD appears to be important for the guiding of ferredoxin to its binding site (main effect on the association rate) whereas PsaE seems to control the photosystem I/ferredoxin complex lifetime (main effect on the dissociation rate). The properties of electron transfer from photosystem I to ferredoxin were also studied at pH 5. 8. In the psaE deleted mutant as in the wild type, the change of pH from 8 to 5.8 induces a 10-fold increase in affinity of ferredoxin for photosystem I. In the absence of PsaD, this pH effect is not observed, in favor of this subunit being mostly responsible for the low pH increased affinity.

Cross-Linking Reagents↗
PubMed2006

NMR study of the electron transfer complex of plant ferredoxin and sulfite reductase: mapping the interaction sites of ferredoxin.

Plant ferredoxin serves as the physiological electron donor for sulfite reductase, which catalyzes the reduction of sulfite to sulfide. Ferredoxin and sulfite reductase form an electrostatically stabilized 1:1 complex for the intermolecular electron transfer. The protein-protein interaction between these proteins from maize leaves was analyzed by nuclear magnetic resonance spectroscopy. Chemical shift perturbation and cross-saturation experiments successfully mapped the location of two major interaction sites of ferredoxin: region 1 including Glu-29, Glu-30, and Asp-34 and region 2 including Glu-92, Glu-93, and Glu-94. The importance of these two acidic patches for interaction with sulfite reductase was confirmed by site-specific mutation of acidic ferredoxin residues in regions 1 and 2, separately and in combination, by which the ability of mutant ferredoxins to transfer electrons and bind to sulfite reductase was additively lowered. Taken together, this study gives a clear illustration of the molecular interaction between ferredoxin and sulfite reductase. We also present data showing that this interaction surface of ferredoxin significantly differs from that when ferredoxin-NADP(+) reductase is the interaction partner.

Aspartic Acid↗
PubMed1984

Ferredoxins from the photosynthetic purple non-sulfur bacterium Rhodopseudomonas palustris. Isolation and amino acid sequence of ferredoxin I.

Two ferredoxins, ferredoxins I and II, were prepared from Rhodopseudomonas palustris. They were separated on a Sephadex column after carboxymethylation and ferredoxin I, the major component, was subjected to an amino acid sequence study. The protein was composed of 63 amino acid residues and the sequence was as follows: (sequence; see text). The molecular weight was calculated to be 6,718, excluding iron and sulfur atoms. The distribution of the nine cysteine residues was similar to but clearly distinct from those of ferredoxins of other photosynthetic bacteria. Comparison of this ferredoxin with those of other bacteria suggests that the photosynthetic bacteria evolved on separate lines. Ferredoxin II was also subjected to analyses of amino acid composition and terminal sequences, but no further study was possible due to the limited material. Although the composition was different from that of ferredoxin I, the terminal sequences were exactly the same as those of ferredoxin I.

Amino Acid Sequence↗
PubMed1997

Molecular cloning of sheep and goat ferredoxin reductase messenger ribonucleic acids, and identification of an alternatively spliced form of sheep ferredoxin reductase.

Complementary DNA clones of mRNAs for sheep and goat NADPH-ferredoxin reductases (ferredoxin NADP+ oxidoreductase, EC 1.18.1.2) were isolated by the reverse transcriptase polymerase chain reaction method, and the complete nucleotide sequences of the coding and 3'-flanking regions of these cDNA clones were determined. Comparative analysis using the deduced amino acid sequences of NADPH-ferredoxin reductases clarified the interspecific conservation of the ferredoxin-binding and flavin adenine dinucleotide (FAD)-binding regions, confirming the results reported previously. During this study, we happened to identify an alternatively spliced mRNA that completely lacks exon 3, just adjacent to the FAD-binding region of the sheep NADPH-ferredoxin reductase cDNA clone. In the screening of other alternatively spliced mRNAs of NADPH-ferredoxin reductases derived from several steroidogenic organs, such as adrenocortices, testes, and ovaries of sheep and goats, only one kind of alternatively spliced mRNA as described above was detected in sheep adrenocortices. Then, we constructed Escherichia coli expression systems for these two forms of mRNA and analyzed their enzymatic properties. We found that the ability of the alternatively spliced NADPH-ferredoxin reductase protein to transfer electrons to ferredoxin is completely abolished because FAD binding is inhibited.

Alternative Splicing↗
PubMed1983

Interaction of ferredoxin and ferredoxin-NADP reductase with thylakoids.

Ferredoxin-NADP reductase accounts for about 50% of the NADPH diaphorase activity of spinach leaf homogenates. The enzyme is bound to thylakoid membranes, but can be slowly extracted by aqueous buffers. Ferredoxin-NADP reductase can be extracted from the membranes by a 1- to 2-min treatment with a low concentration of trypsin. This treatment completely inactivates NADP photoreduction but does not affect electron transport from water to ferredoxin. It is shown that the inactivation is due to solubilization of ferredoxin-NADP reductase: the activity can be restored by addition of a very large excess of soluble enzyme in pure form. When ferredoxin-NADP reductase is added as a soluble enzyme after extraction or inactivation (by a specific antibody) of the membrane-bound enzyme, NADP photoreduction requires a very large excess of this enzyme, and the apparent Km for ferredoxin is also increased. These observations are discussed as related to the interactions of thylakoids with ferredoxin-NADP reductase.

Chloroplasts↗
PubMed1985

Anthracycline antibiotic reduction by spinach ferredoxin-NADP+ reductase and ferredoxin.

Spinach NADPH:ferredoxin oxidoreductase (EC 1.6.7.1) catalyzes the NADPH-dependent reduction of the anthracyclines daunomycin, aclacinomycin A, and nogalamycin and their respective 7-deoxyanthracyclinones. Under anaerobic conditions, the endogenous rate of O2 reduction by NADPH catalyzed by ferredoxin reductase (0.12 s-1 at pH 7.4) is augmented by the anthracyclines and 7-deoxyanthracyclinones. The catalytic constants are approximately equivalent for this augmentation for all substrates (approximate V of 2 s-1 and KM of 75 microM). Both O2- and H2O2 are made. Under anaerobic conditions, anthracycline reduction catalyzed by ferredoxin reductase results in the elimination of the C-7 substituent to provide a quinone methide intermediate. Following tautomerization by C-7 protonation, 7-deoxyanthracyclinones are obtained. Under appropriate conditions these may be further reduced to the 7-deoxyanthracyclinone hydroquinones. For daunomycin, the quinone methide is formed rapidly after reduction and is easily monitored at 600 nm. It may react with electrophiles other than H+, as demonstrated by its competitive trapping by p-carboxybenzaldehyde. It may also react with nucleophiles, as demonstrated by its competitive trapping by N-acetylcysteine. For aclacinomycin, quinone methide formation is also rapid although no distinct transient near 600 nm occurs. In addition to protonation, it reacts with itself providing the 7,7'-dimer. With ethyl xanthate as a thiolate nucleophile, adducts derived from addition to C-7 are obtained. For nogalamycin, quinone methide formation is not rapid. Nogalamycin is reduced to its hydroquinone, which slowly converts in a first-order process [k = (1.2 +/- 0.2) X 10(-3) s-1, pH 8.0, 30 degrees C] to the quinone methide, which is then quenched by protonation. Spinach ferredoxin in its reduced form is chemically competent for anthracycline reduction. Its effect on both the aerobic and anaerobic reactions catalyzed by ferredoxin reductase is to increase severalfold the overall velocity for anthracycline reduction. In conclusion, the aerobic reaction pathways for the anthracyclines as mediated by ferredoxin reductase are remarkably similar, while the anaerobic reactions are remarkably different. If these anthracyclines exert their antitumor activity by a common anaerobic pathway, it is most likely that the pathway is determined by the properties of the anthracycline as complexed to its in vivo target. The behavior of ferredoxin further suggests that not only low-potential flavin centers but also iron-sulfur centers should be regarded as important loci for anthracycline reductive activation.

Aclarubicin↗
PubMed2003

Identification and molecular characterization of mitochondrial ferredoxins and ferredoxin reductase from Arabidopsis.

We have identified and characterized novel types of ferredoxin and ferredoxin reductase from Arabidopsis. Among a number of potential ferredoxin reductase genes in the Arabidopsis genome, AtMFDR was identified to encode a homologue of mitochondrial ferredoxin reductase, and AtMFDX1 and AtMFDX2 were predicted to code for proteins similar to mitochondrial ferredoxin. First, we isolated cDNAs for these proteins and expressed them in heterologous systems of insect cells and Escherichia coli, respectively. The recombinant AtMFDX1 and AtMFDR proteins exhibited spectral properties characteristic of ferredoxin and ferredoxin reductase, respectively, and a pair of recombinant AtMFDX1 and AtMFDR proteins was sufficient to transfer electrons from NAD(P)H to cytochrome c in vitro. Subcellular fractionation analyses suggested membrane association of AtMFDR protein, and protein-gel blot analyses and transient expression studies of green fluorescence protein fusions indicated mitochondrial localization of AtMFDX1 and AtMFDR. RNA-gel blot analyses revealed that the accumulation levels of AtMFDXs and AtMFDR gene transcripts were specifically high in flowers, while protein-gel blot analysis demonstrated substantial accumulation of AtMFDR protein in leaf, stem, and flower. Possible physiological roles of these mitochondrial electron transfer components are discussed in relation to redox dependent metabolic pathways in plants.

Amino Acid Sequence↗
PubMed1978

Ferredoxin-Sepharose 4B as a tool for the purification of ferredoxin-NADP+ reductase.

Ferredoxin immobilized on Sepharose 4B was prepared by reaction of CNBr-Sepharose 4B with spinach ferredoxin. The ferredoxin-Sepharose 4B conjugated ferredoxin-NADP+ reductase (NADPH: ferredoxin oxidoreductase, [EC 1.6.7.1]) in dilute buffer solution and released it in high salt concentrations. A novel method of preparation for the reductase was established by a combination of affinity adsorption on the ferredoxin-Sepharose 4B column with usual purification procedures. It was found using the new method, that there are two forms of ferredoxin-NADP+ reductase, FNR I and FNR II, in spinach. Comparative studies of the two components suggest that FNR I may be a dimer of FNR II.

Chromatography, Affinity↗
PubMed1978

Reactions of antibodies against ferredoxin, ferredoxin-NADP+ reductase and plastocyanin with spinach chloroplasts.

Purified antisera against ferredoxin, ferredoxin-NADP+ reductase and plastocyanin agglutinated osmotically shocked and washed spinach chloroplasts, prepared according to standard procedures. The monomeric antibody (immunoglobulin G fraction) of the reductase antiserum agglutinated chloroplasts specifically and directly, indicating that protruding structures (for example, the coupling factor) do not act as steric hindrances as has been suggested. With ferredoxin antiserum, the presence of a pentameric antibody (immunoglobulin M fraction) was obligatory to observe a positive agglutination reaction. Immunoglobulin G only inhibited ferredoxin-dependent reactions, like NADP+-photoreduction, but did not cause agglutination. Ferredoxin seems to be located in depressions of the membrane, possibly caused by a partial release of this protein in shocked chloroplasts. Similar results were obtained with purified immunoglobulins from a plastocyanin antiserum. Again the immunoglobulin G fraction inhibited electron transport reactions catalyzed by plastocyanin, whereas immunoglobulin M showed a positive agglutination, but had no influence on electron transport. It is concluded that ferredoxin, ferredoxin-NADP+ reductase and plastocyanin are peripheral electron transport components, located at the outer thylakoid membrane.

Agglutination Tests↗
PubMed1988

The transition metal-mediated formation of the hydroxyl free radical during the reduction of molecular oxygen by ferredoxin-ferredoxin:NADP+ oxidoreductase.

The NADPH-supported enzymatic reduction of molecular oxygen by ferredoxin-ferredoxin:NADP+ oxidoreductase was investigated. The ESR spin trapping technique was employed to identify the free radical metabolites of oxygen. The spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) was used to trap and identify the oxygen-derived free radicals. [17O]Oxygen was employed to demonstrate that the oxygen-centered radicals arose from molecular oxygen. From the data, the following scheme is proposed: (Formula:see text). The formation of the free hydroxyl radical during the reduction of oxygen was demonstrated with quantitative competition experiments. The hydroxyl radical abstracted hydrogen from ethanol or formate, and the resulting scavenger-derived free radical was trapped with known rate constants. If H2O2 was added to the enzymatic reaction, a stimulation of the production of the hydroxyl radical was obtained. This stimulation was manifested in both the concentration and the rate of formation of the DMPO/hydroxyl radical adduct. Catalase was shown to inhibit formation of the hydroxyl radical adduct, further supporting the formation of hydrogen peroxide as an intermediate during the reduction of oxygen. All three components, ferredoxin, ferredoxin:NADP+ oxidoreductase, and NADPH, were required for reduction. Ferredoxin:NADP+ oxidoreductase reduces ferredoxin, which in turn is responsible for the reduction of oxygen to hydrogen peroxide and ultimately the hydroxyl radical. The effect of transition metal chelators on the DMPO/hydroxyl radical adduct concentration suggests that the reduction of chelated iron by ferredoxin is responsible for the reduction of hydrogen peroxide to the hydroxyl radical via Fenton-type chemistry.

Algorithms↗
PubMed1984

Ferredoxin:NADP+ oxidoreductase. Equilibria in binary and ternary complexes with NADP+ and ferredoxin.

Ferredoxin:NADP+ oxidoreductase (ferredoxin: NADP+ reductase, EC 1.18.1.2) was shown to form a ternary complex with its substrates ferredoxin (Fd) and NADP(H), but the ternary complex was less stable than the separate binary complexes. Kd for oxidized binary Fd-ferredoxin NADP+ reductase complex was less than 50 nM; Kd(Fd) increased with NADP+ concentration, approaching 0.5-0.6 microM when the flavoprotein was saturated with NADP+ K(NADP+) also increased from about 14 microM to about 310 microM, on addition of excess Fd. The changes in Kd were consistent with negative cooperativity between the associations of Fd and NADP+ and with our unpublished observations which suggest that product dissociation is rate-limiting in the reaction mechanism. Similar interference in binding was observed in more reduced states; NADPH released much ferredoxin:NADP+ reductase from Fd-Sepharose whether the proteins were initially oxidized or reduced. Complexation between Fd and ferredoxin: NADP+ reductase was found to shield each center from paramagnetic probes; charge specificity suggested that the active sites of Fd and ferredoxin:NADP+ reductase were, respectively, negatively and positively charged.

Binding Sites↗