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G Feher

Publications and source records attributed to G Feher.

At least 55 records · Page 3Linked to original sources
PubMed1989

ENDOR studies of the intermediate electron acceptor radical anion I-. in Photosystem II reaction centers.

The EPR and ENDOR characteristics of the intermediate electron acceptor radical anion I-. in Photosystem II (PS II) are shown to be identical in membrane particles and in the D1D2 cytochrome b-559 complex (Nanba, O. and Satoh, K. (1987) Proc. Natl. Acad. Sci. USA 84, 109-112). These findings provide further evidence that the D1D2 complex is the reaction center of PS II and show that the pheophytin binding site is intact. A hydrogen bond between I-. and the protein (GLU D1-130) is postulated on the basis of D2O exchange experiments. The ENDOR data of I-. and of the pheophytin a radical anion in different organic solvents are compared and the observed differences are related to structural changes of the molecule on the basis of molecular orbital calculations (RHF-INDO/SP). The importance of the orientation of the vinyl group (attached to ring I) on electron transfer is discussed.

Anions↗
PubMed1989

Pathway of proton transfer in bacterial reaction centers: replacement of glutamic acid 212 in the L subunit by glutamine inhibits quinone (secondary acceptor) turnover.

The mechanism of proton transfer in the reaction centers (RCs) from Rhodobacter sphaeroides was investigated by site-directed mutagenesis. Replacement of Glu-212 of the L subunit, a protonatable residue located near the secondary acceptor (QB) binding site, by glutamine reduced the in vitro electron turnover from cytochrome c to 2,3-dimethoxy-5-methylbenzoquinone (UQ0) by a factor of 25. The electron transfer rate to QB remained essentially unimpaired. Consequently, it is postulated that the reduced turnover in the mutant is due to a reduced rate of proton transfer to QB2-. The lack of pH dependence of the forward electron transfer rate DQA-QB----DQAQB- and the back reaction rate D+QAQB- ----DQAQB (where D = primary donor and QA = primary acceptor) in the mutant RC indicate that the observed pH dependence in the native RC is due to Glu-212, which has an anomalously high pKa value of 9.5 +/- 0.3. These results support the involvement of Glu-212 as a proton donor to reduced QB.

Bacterial Proteins↗
PubMed1988

Structure of the reaction center from Rhodobacter sphaeroides R-26 and 2.4.1: protein-cofactor (bacteriochlorophyll, bacteriopheophytin, and carotenoid) interactions.

The three-dimensional structures of the cofactors and protein subunits of the reaction center (RC) from the carotenoidless mutant strain of Rhodobacter sphaeroides R-26 and the wild-type strain 2.4.1 have been determined by x-ray diffraction to resolutions of 2.8 A and 3.0 A with R values of 24% and 26%, respectively. The bacteriochlorophyll dimer (D), bacteriochlorophyll monomers (B), and bacteriopheophytin monomers (phi) form two branches, A and B, that are approximately related by a twofold symmetry axis. The cofactors are located in hydrophobic environments formed by the L and M subunits. Differences in the cofactor-protein interactions between the A and B cofactors, as well as between the corresponding cofactors of Rb, sphaeroides and Rhodopseudomonas viridis [Michel, H., Epp, O. & Deisenhofer, J. (1986) EMBO J. 3, 2445-2451], are delineated. The roles of several structural features in the preferential electron transfer along the A branch are discussed. Two bound detergent molecules of beta-octyl glucoside have been located near BA and BB. The environment of the carotenoid, C, that is present in RCs from Rb. sphaeroides 2.4.1 consists largely of aromatic residues of the M subunit. A role of BB in the triplet energy transfer from D to C and the reason for the preferential ease of removal of BB from the RC is proposed.

Bacteriochlorophylls↗
PubMed1988

Structure of the reaction center from Rhodobacter sphaeroides R-26: protein-cofactor (quinones and Fe2+) interactions.

The three-dimensional structure of the reaction center (RC) from Rhodobacter sphaeroides has been determined by x-ray diffraction to a resolution of 2.8 A with an R value of 24%. The interactions of the protein with the primary quinone, QA, secondary quinone, QB, and the nonheme iron are described and compared to those of RCs from Rhodopseudomonas viridis. Structural differences between the QA and QB environments that contribute to the function of the quinones (the electron transfer from QA- to QB and the charge recombination of QA-, QB- with the primary donor) are delineated. The protein residues that may be involved in the protonation of QB are identified. A pathway for the doubly reduced QB to dissociate from the RC is proposed. The interactions between QB and the residues that have been changed in herbicide-resistant mutants are described. The environment of the nonheme iron is compared to the environments of metal ions in other proteins.

Bacterial Proteins↗
PubMed1988

Structure of the reaction center from Rhodobacter sphaeroides R-26 and 2.4.1: symmetry relations and sequence comparisons between different species.

Photosynthetic reaction centers from purple bacteria exhibit an approximate twofold symmetry axis, which relates both the cofactors and the L and M subunits. For the reaction center from Rhodobacter sphaeroides, deviations from this twofold symmetry axis have been quantitated by superposing, by a 180 degrees rotation, the cofactors of the B branch onto the A branch and the M subunit onto the L subunit. An alignment of the sequences of the L and M subunits from four purple bacteria, one green bacterium, and the D1 and D2 subunits of a photosystem II-containing green alga is presented. The residues that are conserved in all six species are shown in relation to the structure of Rb. sphaeroides and their possible role in the function of the reaction center is discussed. A method is presented for characterizing the exposure of alpha-helices to the membrane based on the periodicity of conserved residues. This method may prove useful for modeling the three-dimensional structures of membrane proteins.

Amino Acid Sequence↗
PubMed1987

Structure of the reaction center from Rhodobacter sphaeroides R-26: the cofactors.

The three-dimensional structure of the cofactors of the reaction center of Rhodobacter sphaeroides R-26 has been determined by x-ray diffraction and refined at a resolution of 2.8 A with an R value of 26%. The main features of the structure are similar to the ones determined for Rhodopseudomonas viridis [Michel, H., Epp, O. & Deisenhofer, J. (1986) EMBO J. 5, 2445-2451]. The cofactors are arranged along two branches, which are approximately related to each other by a 2-fold symmetry axis. The structure is well suited to produce light-induced charge separation across the membrane. Most of the structural features predicted from physical and biochemical measurements are confirmed by the x-ray structure.

Bacterial Proteins↗
PubMed1987

Structure of the reaction center from Rhodobacter sphaeroides R-26: the protein subunits.

The three-dimensional structure of the protein subunits of the reaction center (RC) of Rhodobacter sphaeroides has been determined by x-ray diffraction at a resolution of 2.8 A with an R factor of 26%. The L and M subunits each contain five transmembrane helices and several helices that do not span the membrane. The L and M subunits are related to each other by a 2-fold rotational symmetry axis that is approximately the same as that determined for the cofactors. The H subunit has one transmembrane helix and a globular domain on the cytoplasmic side, which contains a helix that does not span the membrane and several beta-sheets. The structural homology with RCs from other purple bacteria is discussed. A structure of the complex formed between the water soluble cytochrome c2 and the RC from Rb. sphaeroides is proposed.

Bacterial Proteins↗
PubMed1987

Structure of the reaction center from Rhodobacter sphaeroides R-26: membrane-protein interactions.

The energetics of membrane-protein interactions are analyzed with the three-dimensional model of the photosynthetic reaction center (RC) from Rhodobacter sphaeroides. The position of the RC in the membrane and the thickness of the membrane were obtained by minimizing the hydrophobic energy with the energy function of Eisenberg and McLachlan. The 2-fold symmetry axis that relates the L and M subunits is, within the accuracy of 5 degrees, parallel to the normal of the membrane. The thickness of the membrane is estimated to be 40-45 A. Residues that are exposed to the membrane are relatively poorly conserved in the sequences of homologous RC proteins. The surface area of the RC is comparable to the surface areas of water-soluble proteins of similar molecular weight. The volumes of interior atoms in the RC are also similar to those of water-soluble proteins, indicating the same compact packing for both types of proteins. The electrostatic potential of the cofactors was calculated. The results show an asymmetry in the potential between the two possible pathways of electron transfer, with the A branch being preferred electrostatically.

Bacteria↗
PubMed1987

The Stark effect in reaction centers from Rhodobacter sphaeroides R-26 and Rhodopseudomonas viridis.

The effect of an electric field on the optical absorption (Stark effect) of reaction centers (RCs) from Rhodobacter sphaeroides and Rhodopseudomonas viridis embedded in films of poly(vinyl alcohol) was measured. The infrared bands were investigated at 295 K and 77 K. In RCs from Rp. viridis at 77 K six peaks (at 982, 849, 835, 818, 803, and 787 nm), associated with the Qy transitions of the six pigments, were resolved; in addition, a small broad band at 865 nm was resolved. In RCs from Rb. sphaeroides only five bands (at 877, 817, 802, 761, and 754 nm) assigned to the Qy transitions were resolved; in addition, two small bands at 697 and 683 nm were observed. The additional bands have been tentatively assigned to vibrational side bands, although the contribution from charge-transfer states cannot be excluded. The Stark spectra had line shapes similar to the second derivative of the absorption spectra and were interpreted in terms of the interaction between the applied electric field and the dipole moments of the ground and excited states. Analyses of the spectra yielded the apparent change in dipole moment delta mu app = f delta mu (where the factor f corrects for the difference between the local field and the applied field) and delta, the angle between delta mu---- and the transition moment mu trans. At 77 K the values of delta mu----app and delta for the peaks at 877, 802, and 761 nm in Rb. sphaeroides were 6.5 debye, 38 degrees; 2.1 debye, 23 degrees; and 3.5 debye, 8 degrees. In Rp. viridis the debye values for the peaks at 982, 835, 818, and 787 were 8.2, 40 degrees; 1.8, 50 degrees; 3.4, 14 degrees; and 2.7, 0 degrees. The large values of delta mu app associated with the long-wavelength peak of the bacteriochlorophyll dimers are consistent with a significant charge-transfer contribution to the excited state of the primary donor.

Bacterial Proteins↗
PubMed1986

Iron-depleted reaction centers from Rhodopseudomonas sphaeroides R-26.1: characterization and reconstitution with Fe2+, Mn2+, Co2+, Ni2+, Cu2+, and Zn2+.

Reaction centers (RCs) from the photosynthetic bacterium Rhodopseudomonas sphaeroides R-26.1 were depleted of Fe by a simple procedure involving reversible dissociation of the H subunit. The resulting intact Fe-depleted RCs contained 0.1-0.2 Fe per RC as determined from atomic absorption and electron paramagnetic resonance (EPR) spectroscopy. Fe-depleted RCs that have no metal ion occupying the Fe site differed from native RCs in the following respects: (1) the rate of electron transfer from QA- to QB exhibited nonexponential kinetics with the majority of RCs having a rate constant slower by only a factor of approximately 2, (2) the efficiency of light-induced charge separation (DQA----D+QA-) produced by a saturating flash decreased to 63%, and (3) QA appeared readily reducible to QA2-. Various divalent metal ions were subsequently incorporated into the Fe site. The electron transfer characteristics of Fe-depleted RCs reconstituted with Fe2+, Mn2+, Co2+, Ni2+, Cu2+, and Zn2+ were essentially the same as those of native RCs. These results demonstrate that neither Fe2+ nor any divalent metal ion is required for rapid electron transfer from QA- to QB. However, the presence of a metal ion in the Fe site is necessary to establish the characteristic, native, electron-transfer properties of QA. The lack of a dominant role of Fe2+ or other divalent metals in the observed rate of electron transfer from QA- to QB suggests that a rate-limiting step (for example, a protonation event or a light-induced structural change) precedes electron transfer.

Bacterial Proteins↗
PubMed1986

Primary structure of the reaction center from Rhodopseudomonas sphaeroides.

The reaction center is a pigment-protein complex that mediates the initial photochemical steps of photosynthesis. The amino-terminal sequences of the L, M, and H subunits and the nucleotide and derived amino acid sequences of the L and M structural genes from Rhodopseudomonas sphaeroides have previously been determined. We report here the sequence of the H subunit, completing the primary structure determination of the reaction center from R. sphaeroides. The nucleotide sequence of the gene encoding the H subunit was determined by the dideoxy method after subcloning fragments into single-stranded M13 phage vectors. This information was used to derive the amino acid sequence of the corresponding polypeptide. The termini of the primary structure of the H subunit were established by means of the amino and carboxy terminal sequences of the polypeptide. The data showed that the H subunit is composed of 260 residues, corresponding to a molecular weight of 28,003. A molecular weight of 100,858 for the reaction center was calculated from the primary structures of the subunits and the cofactors. Examination of the genes encoding the reaction center shows that the codon usage is strongly biased towards codons ending in G and C. Hydropathy analysis of the H subunit sequence reveals one stretch of hydrophobic residues near the amino terminus; the L and M subunits contain five such stretches. From a comparison of the sequences of homologous proteins found in bacterial reaction centers and photosystem II of plants, an evolutionary tree was constructed. The analysis of evolutionary relationships showed that the L and M subunits of reaction centers and the D1 and D2 proteins of photosystem II are descended from a common ancestor, and that the rate of change in these proteins was much higher in the first billion years after the divergence of the reaction center and photosystem II than in the subsequent billion years represented by the divergence of the species containing these proteins.

Amino Acid Sequence↗
PubMed1986

Primary photochemistry of iron-depleted and zinc-reconstituted reaction centers from Rhodopseudomonas sphaeroides.

The primary photochemistry of Fe-depleted and Zn-reconstituted reaction centers from Rhodopseudomonas sphaeroides R-26.1 was studied by transient absorption spectroscopy and compared with native, Fe(2+)-containing reaction centers. Excitation of metal-free reaction centers with 30-ps flashes produced the initial charge-separated state P(+)I(-) (P(+)BPh(-), where P is the primary donor and BPh is bacteriopheophytin) with a yield and visible/near-infrared absorption difference spectrum indistinguishable from that observed in native reaction centers. However, the lifetime of P(+)I(-) was found to increase approximately 20-fold to 4.2 +/- 0.3 ns (compared to 205 ps in native reaction centers), and the yield of formation of the subsequent state P(+)Q(A) (-) (Q(A) is the primary quinone acceptor) was reduced to 47 +/- 5% (compared to essentially 100% in native reaction centers). The remaining 53% of the metal-free reaction centers were found to undergo charge recombination during the P(+)I(-) lifetime to yield both the ground state (28 +/- 5%) and the triplet state P(R) (25 +/- 5%). Reconstitution of Fe-depleted reaction centers with Zn(2+) restored the "native" photochemistry. Possible mechanisms responsible for the reduced decay rate of P(+)I(-) in metal-free reaction centers are discussed.

Journal Article↗
PubMed1986

Isotope effect on electron transfer in reaction centers from Rhodopseudomonas sphaeroides.

Previous ENDOR studies on reaction centers from Rhodopseudomonas sphaeroides have shown the presence of two hydrogen-bonded protons associated with the primary, ubiquinone, acceptor Q(A). These protons exchange with deuterons from solvent (2)H(2)O. The effect of this deuterium substitution on the charge-recombination kinetics (BChl)(2) (+)Q(A) (-) --> (BChl)(2)Q(A) has been studied with a sensitive kinetic difference technique. The electron-transfer rate was found to increase with deuterium exchange up to a maximum Deltak/k of 5.7 +/- 0.3%. The change in rate was found to have an exchange time of 2 hr, which matched the disappearance of the ENDOR lines due to the exchangeable protons. These results indicate that these protons play a role in the vibronic coupling associated with electron transfer. A simple model for the isotope effect on electron transfer predicts a maximum rate increase of 20%, which is consistent with the experimental results.

Journal Article↗
PubMed1986

Structural homology of reaction centers from Rhodopseudomonas sphaeroides and Rhodopseudomonas viridis as determined by x-ray diffraction.

Crystals of the reaction center (RC) from Rhodopseudomonas sphaeroides with the space group P2(1)2(1)2(1), have been studied by x-ray diffraction. The Patterson search (molecular replacement) technique was used to analyze the data, with the structure of the reaction center from Rhodopseudomonas viridis as a model system. A preliminary electron density map of the reaction center from R. sphaeroides has been obtained. Comparison of the structure of the RC from R. sphaeroides with that from R. viridis showed the following conserved features: five membrane-spanning helices in each of the L and M subunits, a single membrane-spanning helix in the H subunit, a 2-fold symmetry axis, and similar positions and orientations of the cofactors. Unlike the RCs from R. viridis, both quinones are retained in the RCs from R. sphaeroides. The secondary quinone is located near the position related by the 2-fold symmetry axis to the primary quinone.

Bacterial Proteins↗
PubMed1985

Electron nuclear double resonance of semiquinones in reaction centers of Rhodopseudomonas sphaeroides.

Replacement of Fe2+ by Zn2+ in reaction centers of Rhodopseudomonas sphaeroides enabled us to perform ENDOR (electron nuclear double resonance) experiments on the anion radicals of the primary and secondary ubiquinone acceptors (QA- and QB-. Differences between the QA and QB sites, hydrogen bonding to the oxygens, interactions with the protons of the proteins and some symmetry properties of the binding sites were deduced from an analysis of the ENDOR spectra.

Bacterial Proteins↗
PubMed1985

The effect of an applied electric field on the charge recombination kinetics in reaction centers reconstituted in planar lipid bilayers.

Reaction Centers (RCs) from the photosynthetic bacterium Rhodopseudomonas sphaeroides were incorporated in planar bilayers made from monolayers derived from liposomes reconstituted with purified RCs. The photocurrents associated with the charge recombination process between the reduced primary quinone (QA-) and the oxidized bacteriochlorophyll donor (D+) were measured as a function of voltage (-150 mV less than V less than 150 mV) applied across the bilayer. When QA was the native ubiquinone (UQ) the charge recombination was voltage independent. However, when UQ was replaced by anthraquinone (AQ), the recombination time depended on the applied voltage V according to the relation tau = 8.5 X 10(-3) eV/0.175S. These results were explained by a simple model in which the charge recombination from UQ- proceeds directly to D+ while that from AQ occurs via a thermally activated intermediate state, D+I-QA, where I is the intermediate acceptor. The voltage dependence arises from an electric field induced change in the energy gap, delta G0, between the states D+I-QA and D+IQA-. This model is supported by the measured temperature dependence of the charge recombination time, which for RCs with AQ gave a value of delta G0 = 340 +/- 20 meV. In contrast, delta G0 for RCs with UQ as the primary acceptor, is sufficiently large (approximately 550 meV) so that even in the presence of the field, the direct pathway dominates. The voltage dependence shows that the electron transfer from I- to QA is electrogenic. From a quantitative analysis of the voltage dependence on the recombination rate it was concluded that the component of the distance between I and QA along the normal to the membrane is about one-seventh of the thickness of the membrane. This implies that the electron transfer from I to Q contributes at least one-seventh to the potential generated by the charge separation between D+ and QA-.

Bacterial Proteins↗