共查询到20条相似文献,搜索用时 9 毫秒
1.
《FEBS letters》1987,221(2):343-348
Spinach sulfite reductase has been shown to co-migrate during gel filtration chromatography at low ionic strength with spinach ferredoxin. No co-migration was observed at high ionic strength. These results indicate that the two proteins form a high-affinity, electrostatically stabilized complex, as had previously been demonstrated for three other ferredoxin-dependent, plant enzymes. Modification of 3–4 ferredoxin carboxyl groups had little detectable effect on the ferredoxin-sulfite reductase interaction. 相似文献
2.
Xu X Kim SK Schürmann P Hirasawa M Tripathy JN Smith J Knaff DB Ubbink M 《FEBS letters》2006,580(28-29):6714-6720
The reduction of ferredoxin-thioredoxin reductase (FTR) by plant-type ferredoxin plays an important role in redox regulation in plants and cyanobacteria. Nuclear magnetic resonance (NMR) was used to map the binding sites on Synechocystis ferredoxin for FTR. A gallium-substituted structural analog of this [2Fe-2S] ferredoxin was obtained by reconstituting the apoprotein in a refolding buffer containing gallium. For the first time, the complete interaction interface of a [2Fe-2S] ferredoxin with a target enzyme has been mapped by NMR chemical shift perturbation with this diamagnetic structural analog. 相似文献
3.
Kurisu G Kusunoki M Katoh E Yamazaki T Teshima K Onda Y Kimata-Ariga Y Hase T 《Nature structural biology》2001,8(2):117-121
All oxygenic photosynthetically derived reducing equivalents are utilized by combinations of a single multifuctional electron carrier protein, ferredoxin (Fd), and several Fd-dependent oxidoreductases. We report the first crystal structure of the complex between maize leaf Fd and Fd-NADP(+) oxidoreductase (FNR). The redox centers in the complex--the 2Fe-2S cluster of Fd and flavin adenine dinucleotide (FAD) of FNR--are in close proximity; the shortest distance is 6.0 A. The intermolecular interactions in the complex are mainly electrostatic, occurring through salt bridges, and the interface near the prosthetic groups is hydrophobic. NMR experiments on the complex in solution confirmed the FNR recognition sites on Fd that are identified in the crystal structure. Interestingly, the structures of Fd and FNR in the complex and in the free state differ in several ways. For example, in the active site of FNR, Fd binding induces the formation of a new hydrogen bond between side chains of Glu 312 and Ser 96 of FNR. We propose that this type of molecular communication not only determines the optimal orientation of the two proteins for electron transfer, but also contributes to the modulation of the enzymatic properties of FNR. 相似文献
4.
Plant sulfite reductase: molecular structure, catalytic function and interaction with ferredoxin 总被引:6,自引:0,他引:6
Plant sulfite reductase contains the siroheme and the [4Fe-4S] cluster as catalytically active redox centers and catalyzes the six-electron reductions of sulfite and nitrite using electrons donated from ferredoxin. A heterologous expression of a cDNA for maize sulfite reductase in E. coli has enabled us to produce the wild-type and mutant enzymes. Putative substrate-binding basic residues, located at the siroheme distal side, have been substituted for other residues with neutral or negatively charged side chains. Kinetic studies of the resulting mutant enzymes have demonstrated that substrate specificity for the two anions is remarkably changed by amino acid substitutions at a single site. We have also produced two classes of ferredoxin mutants with less ability to donate electrons to sulfite reductase: one with a defect in the recognition of the partner enzyme and the other with an unfavorable redox property. This article summarizes our knowledge about the structure function relationships of plant sulfite reductase. 相似文献
5.
Teshima K Fujita S Hirose S Nishiyama D Kurisu G Kusunoki M Kimata-Ariga Y Hase T 《FEBS letters》2003,546(2-3):189-194
In order to elucidate the importance of a ferredoxin (Fd) Arg-Glu pair involved in dynamic exchange from intra- to intermolecular salt bridges upon complex formation with ferredoxin-NADP(+) oxidoreductase (FNR), Equisetum arvense FdI and FdII were investigated as normal and the pair-lacking Fd, respectively. The FdI mutant lacking this pair was unstable and rapidly lost the [2Fe-2S] cluster. The catalytic constant (k(cat)) of the electron transfer for FdI is 5.5 times that for FdII and the introduction of this pair into FdII resulted in the increase of k(cat) to a level comparable to that for FdI, demonstrating directly that the Arg-Glu pair is important for efficient electron transfer between Fd and FNR. 相似文献
6.
7.
The effects of complex formation with flavodoxin on the proton NMR spectrum of cytochrome c are to change the resonance frequencies and to increase the bandwidths of most of the low and high field heme, Met-80, and His-18 protons. These effects are, in general, more pronounced than has been reported for other cytochrome c complexes. The degree of line broadening for many heme related resonances suggests that complex formation induces changes in the cytochrome structure. These results provide the first spectroscopic evidence which corroborates the proposed model for the cytochrome c: flavodoxin complex (1-3). 相似文献
8.
T Akashi T Matsumura T Ideguchi K Iwakiri T Kawakatsu I Taniguchi T Hase 《The Journal of biological chemistry》1999,274(41):29399-29405
Plant-type ferredoxin (Fd), a [2Fe-2S] iron-sulfur protein, functions as an one-electron donor to Fd-NADP(+) reductase (FNR) or sulfite reductase (SiR), interacting electrostatically with them. In order to understand the protein-protein interaction between Fd and these two different enzymes, 10 acidic surface residues in maize Fd (isoform III), Asp-27, Glu-30, Asp-58, Asp-61, Asp-66/Asp-67, Glu-71/Glu-72, Asp-85, and Glu-93, were substituted with the corresponding amide residues by site-directed mutagenesis. The redox potentials of the mutated Fds were not markedly changed, except for E93Q, the redox potential of which was more positive by 67 mV than that of the wild type. Kinetic experiments showed that the mutations at Asp-66/Asp-67 and Glu-93 significantly affected electron transfer to the two enzymes. Interestingly, D66N/D67N was less efficient in the reaction with FNR than E93Q, whereas this relationship was reversed in the reaction with SiR. The static interaction of the mutant Fds with each the two enzymes was analyzed by gel filtration of a mixture of Fd and each enzyme, and by affinity chromatography on Fd-immobilized resins. The contributions of Asp-66/Asp-67 and Glu-93 were found to be most important for the binding to FNR and SiR, respectively, in accordance with the kinetic data. These results allowed us to map the acidic regions of Fd required for electron transfer and for binding to FNR and SiR and demonstrate that the interaction sites for the two enzymes are at least partly distinct. 相似文献
9.
Ferredoxin and the flavoprotein, ferredoxin: NADP reductase, have been covalently linked by incubation in the presence of a water soluble carbodiimide. The cross-linking reaction yields an adduct having a 1:1 stoichiometry. The adduct has depressed levels of diaphorase and NADPH oxidase activity and is inactive in reduction of cytochrome c using NADPH as an electron donor. Thus, although similar to an adduct described by Zanetti and coworkers [J Biol Chem 259: 6153–6157 (1984)] in its stoichiometry, the adduct described herein has significantly different enzymatic properties. It is suggested that this may be a reflection of differences in the interaction between the two proteins resulting from differences in experimental conditions in which the two adducts were prepared.Abbreviations Fd
ferredoxin
- Fp
ferredoxin: NADP reductase
- Fd
Fp covalently linked Fd-Fp adduct
- Fd:Fp
noncovalently linked complex between Fd and Fp
- EDC
1-ethyl-3-(dimethylaminopropyl) carbodiimide
- Tris
tris-hydroxymethylaminomethane
- MOPS
3-(N-morpholino)propane sulfonic acid
- DCIP
2,6-dichloropenolindophenol 相似文献
10.
11.
A comparison was made of graphical and subtractive methods for the determination of the dissociation constant of a complex between ferredoxin:NADP reductase and NADP. The subtractive method gave Kd values near 10 μm which are consistent with recently determined values for Km,NADP in assays of NADP photoreduction by chloroplast membranes. The graphical method gave values which were considerably higher. The difference between the two methods is due to the failure of the graphical method to correct for the amount of each component present in the complex at the low NADP/ flavoprotein ratios necessary for binding studies. A second NADP binding site of much lower affinity (Kd approx 1 mm) was also detected. 相似文献
12.
Vlasie MD Fernández-Busnadiego R Prudêncio M Ubbink M 《Journal of molecular biology》2008,375(5):1405-1415
Copper-containing nitrite reductase is able to catalyze the reduction of nitrite with a turnover rate of several hundreds per second. Electrons for the reaction are donated by the electron transfer protein pseudoazurin. The process of protein complex formation, electron transfer and dissociation must occur on the millisecond timescale to enable the fast turnover of the enzyme. The structure of this transient protein complex has been studied using paramagnetic NMR spectroscopy. Gadolinium complexes were attached specifically through two engineered Cys residues on three sites on the surface of nitrite reductase, causing strong distance-dependent relaxation effects on the residues of pseudoazurin. Docking of the two proteins based on these NMR-derived distance restraints and the chemical shift perturbation data shows convergence to a cluster of structures with an average root-mean-square deviation of 1.5 Å. The binding interface consists of polar and non-polar residues surrounded by charges. The interprotein distance between the two type-1 copper sites is 15.5(± 0.5) Å, enabling fast interprotein electron transfer. The NMR-based lower limit estimate of 600 s−1 for the dissociation rate constant and the fast electron transfer are consistent with the transient nature of the complex. 相似文献
13.
The ferredoxin:thioredoxin reductase is an essential enzyme of the light dependent regulatory system in oxygenic photosynthesis. It is composed of two dissimilar subunits and contains a 4Fe-4S cluster and a redox-active disulfide bridge. Artificial electron donors of redox potentials below –300 mV are capable of reducing the disulfide bridge. Based on our results we speculate that a group of more negative potential than the disulfide bridge is the first acceptor of the electrons in FTR. The chemical reduction of FTR has been used successfully for the detection of the enzyme during its purification.Abbreviation FBPase
fructose 1,6-bisphosphatase
- FTR
ferredoxin:thioredoxin reductase
- MV
methyl viologen
Dedicated to Prof. D.I. Arnon. 相似文献
14.
Simone Dell’Acqua Isabel Moura José J. G. Moura Sofia R. Pauleta 《Journal of biological inorganic chemistry》2011,16(8):1241-1254
Identifying redox partners and the interaction surfaces is crucial for fully understanding electron flow in a respiratory
chain. In this study, we focused on the interaction of nitrous oxide reductase (N2OR), which catalyzes the final step in bacterial denitrification, with its physiological electron donor, either a c-type cytochrome or a type 1 copper protein. The comparison between the interaction of N2OR from three different microorganisms, Pseudomonas nautica, Paracoccus denitrificans, and Achromobacter cycloclastes, with their physiological electron donors was performed through the analysis of the primary sequence alignment, electrostatic
surface, and molecular docking simulations, using the bimolecular complex generation with global evaluation and ranking algorithm.
The docking results were analyzed taking into account the experimental data, since the interaction is suggested to have either
a hydrophobic nature, in the case of P. nautica N2OR, or an electrostatic nature, in the case of P. denitrificans N2OR and A. cycloclastes N2OR. A set of well-conserved residues on the N2OR surface were identified as being part of the electron transfer pathway from the redox partner to N2OR (Ala495, Asp519, Val524, His566 and Leu568 numbered according to the P. nautica N2OR sequence). Moreover, we built a model for Wolinella succinogenes N2OR, an enzyme that has an additional c-type-heme-containing domain. The structures of the N2OR domain and the c-type-heme-containing domain were modeled and the full-length structure was obtained by molecular docking simulation of these
two domains. The orientation of the c-type-heme-containing domain relative to the N2OR domain is similar to that found in the other electron transfer complexes. 相似文献
15.
Charon MH Volbeda A Chabriere E Pieulle L Fontecilla-Camps JC 《Current opinion in structural biology》1999,9(6):663-669
The first crystal structure of pyruvate:ferredoxin oxidoreductase to be determined has provided significant new information on its structural organization and redox chemistry. Spectroscopic analyses of a radical reaction intermediate have shed more light on its thiamin-based mechanism of catalysis. Different approaches have been used to study the interaction between the enzyme and ferredoxin, its redox partner. 相似文献
16.
Hurley JK Morales R Martínez-Júlvez M Brodie TB Medina M Gómez-Moreno C Tollin G 《Biochimica et biophysica acta》2002,1554(1-2):5-21
The interaction between reduced Anabaena ferredoxin and oxidized ferredoxin:NADP(+) reductase (FNR), which occurs during photosynthetic electron transfer (ET), has been investigated extensively in the authors' laboratories using transient and steady-state kinetic measurements and X-ray crystallography. The effect of a large number of site-specific mutations in both proteins has been assessed. Many of the mutations had little or no effect on ET kinetics. However, non-conservative mutations at three highly conserved surface sites in ferredoxin (F65, E94 and S47) caused ET rate constants to decrease by four orders of magnitude, and non-conservative mutations at three highly conserved surface sites in FNR (L76, K75 and E301) caused ET rate constants to decrease by factors of 25-150. These residues were deemed to be critical for ET. Similar mutations at several other conserved sites in the two proteins (D67 in Fd; E139, L78, K72, and R16 in FNR) caused smaller but still appreciable effects on ET rate constants. A strong correlation exists between these results and the X-ray crystal structure of an Anabaena ferredoxin/FNR complex. Thus, mutations at sites that are within the protein-protein interface or are directly involved in interprotein contacts generally show the largest kinetic effects. The implications of these results for the ET mechanism are discussed. 相似文献
17.
《FEBS letters》1985,186(2):149-152
The interaction of ferredoxin with thylakoids is shown to occur at two distinct sites: at the reducing end of photosystem I, and at the site where ferredoxin-NADP reductase (FNR) is located on the membrane. The evidence is based on the lack of inhibition of ferredoxin photoreduction by the extraction of FNR or its inactivation by an antibody, and on the difference between Km values for ferredoxin in reactions requiring FNR as compared to those only requiring ferredoxin. 相似文献
18.
The trinitrophenylation of a single amino group of spinach ferredoxin abolishes its ability to inhibit the diaphorase activity of the flavoprotein, ferredoxin:NADP oxidoreductase (EC 1.6.7.1); in contrast, the ability of ferredoxin to participate in the ferredoxin-linked cytochrome c reductase activity catalyzed by the flavoprotein is unaffected. Comparison with previously published results [Davis, D. J., and San Pietro, A. (1977) Biochem. Biophys. Res. Commun.74, 33–40]indicates that the site of interaction between ferredoxin and the flavoprotein resulting in inhibition if diaphorase activity is responsible for the spectrally observable 1:1 complex between the two proteins and is identical to the site of ferredoxin involvement in NADP photoreduction. The role of ferredoxin in the ferredoxin-linked cytochrome c reductase activity of the flavoprotein has been reexamined under conditions were the entire electron-accepting system (rather than just the ferredoxin component) is rate limiting. The data support a mechanism by which ferredoxin can bind either to the flavoprotein or to cytochrome c, and the ferredoxin:cytochrome c complex serves as the true substrate for reduction by the flavoprotein. Furthermore, Chromatographic evidence is presented for the formation of complexes between ferredoxin and cytochrome c. 相似文献
19.
Electron transfer by ferredoxin:NADP+ reductase. Rapid-reaction evidence for participation of a ternary complex 总被引:1,自引:0,他引:1
Rapid reaction studies presented herein show that ferredoxin:NADP+ oxidoreductase (FNR, EC 1.18.1.2) catalyzes electron transfer from spinach ferredoxin (Fd) to NADP+ via a ternary complex, Fd X FNR X NADP+. In the absence of NADP+, reduction of ferredoxin:NADP+ reductase by Fd was much slower than the catalytic rate: 37-80 s-1 versus at least 445 e-s-1; dissociation of oxidized spinach ferredoxin (Fdox) from one-electron reduced ferredoxin:NADP+ reductase (FNRsq) limited the reduction of FNR. This confirms the steady-state kinetic analysis of Masaki et al. (Masaki, R., Yoshikaya, S., and Matsubara, H. (1982) Biochim. Biophys. Acta 700, 101-109). Occupation of the NADP+ binding site of FNR by NADP+ or by 2',5'-ADP (a nonreducible NADP+ analogue) greatly increased the rate of electron transfer from Fd to FNR, releiving inhibition by Fdox. NADP+ (and 2',5'-ADP) probably facilitate the dissociation of Fdox; equilibrium studies have shown that nucleotide binding decreases the association of Fd with FNR (Batie, C. J. (1983) Ph.D. dissertation, Duke University; Batie, C. J., and Kamin, H. (1982) in Flavins and Flavoproteins VII (Massey, V., and Williams, C. H., Jr., eds) pp. 679-683, Elsevier, New York; Batie, C.J., and Kamin, H. (1982) Fed. Proc. 41, 888; and Batie, C.J., and Kamin, H. (1984) J. Biol. Chem. 259, 8832-8839). Premixing Fd with FNR was found to inhibit the reaction of the flavoprotein with NADP+ and with NADPH; thus, substrate binding may be ordered, NADP+ first, then Fd. FNRred and NADP+ very rapidly formed an FNRred X NADP+ complex with flavin to nicotinamide charge transfer bands. The Fdred X NADP+ complex then relaxed to an equilibrium species; the spectrum indicated a predominance of FNRox X NADPH charge-transfer complex. However, charge-transfer species were not observed during turnover; thus, their participation in catalysis of electron transfer from Fd to NADP+ remains uncertain. The catalytic rate of Fd to NADP+ electron transfer, as well as the rates of electron transfer from Fd to FNR, and from FNR to NADP+ were decreased when the reactants were in D2O; diaphorase activity was unaffected by solvent. On the basis of the data presented, a scheme for the catalytic mechanism of catalysis by FNR is presented. 相似文献
20.
R Cammack J Neumann N Nelson D O Hall 《Biochemical and biophysical research communications》1971,42(2):292-297
Circular dichroism (CD) spectra are presented of ferredoxin, ferredoxin-NADP reductase and their complex. A change in CD occurs on complex formation which is consistent with a decrease in the Cotton effects due to the ferredoxin. This change is interpreted as due to a decrease in interaction in ferredoxin between the iron-sulphur chromophore group and the protein. 相似文献