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1.
Phys Chem Chem Phys ; 22(17): 9487-9493, 2020 May 07.
Artículo en Inglés | MEDLINE | ID: mdl-32314999

RESUMEN

We report on a novel approach to the study of rates and short-lived intermediates of (bio)chemical reactions that involve paramagnetic species. Temperature-cycle Electron Paramagnetic Resonance (EPR) concerns the repeated heating of a reaction mixture in the cavity of an EPR spectrometer by pulsed irradiation with a near-infrared diode laser combined with intermittent characterization of the sample by 275 GHz EPR at a lower temperature at which the reaction does not proceed. The new technique is demonstrated for the reduction of TEMPOL with sodium dithionite in aqueous solution down to the sub-second time scale. We show that a single sample suffices to obtain a complete kinetic trace. Variation of the length and power of the laser pulse offers great flexibility as regards the time scale of the experiment and the temperature at which the reaction can be studied. For water/glycerol mixtures we introduce a simple way to obtain and load an unreacted sample into the spectrometer at low temperature.

2.
Phys Chem Chem Phys ; 22(8): 4840, 2020 Feb 26.
Artículo en Inglés | MEDLINE | ID: mdl-32064478

RESUMEN

Correction for 'Conformation of bis-nitroxide polarizing agents by multi-frequency EPR spectroscopy' by Janne Soetbeer et al., Phys. Chem. Chem. Phys., 2018, 20, 25506-25517.

3.
Phys Chem Chem Phys ; 21(31): 16937-16948, 2019 Aug 21.
Artículo en Inglés | MEDLINE | ID: mdl-31339131

RESUMEN

Multi-frequency EPR spectroscopy can provide high-level structural information on high-spin Fe3+ sites in proteins and enzymes. Unfortunately, analysis of the EPR spectra of these spin systems is hindered by the presence of broad distributions in the zero-field-splitting (ZFS) parameters, which reflect conformational heterogeneity of the iron sites. We present the analysis of EPR spectra of high-spin Fe3+ bound to human serum transferrin. We apply a method termed the grid-of-errors to extract the distributions of the individual ZFS parameters from EPR spectra recorded in the high-field limit at a microwave frequency of 275 GHz. Study of a series of transferrin variants shows that the ZFS distributions are as characteristic of the structure of a high-spin Fe3+ site as the ZFS parameters themselves. Simulations based on the extracted ZFS distributions reproduce spectra recorded at 34 GHz (Q band) and 9.7 GHz (X band), including subtle variations that were previously difficult to quantify. The X-band spectrum of transferrin shows a characteristic double peak, which has puzzled researchers for decades. We show that the double peak is uniquely related to the term B4-3O4-3(S) in the spin Hamiltonian. Our method is generally applicable in the analysis of spectra that arise from a broad distribution of parameters.

4.
Phys Chem Chem Phys ; 20(39): 25506-25517, 2018 Oct 10.
Artículo en Inglés | MEDLINE | ID: mdl-30277229

RESUMEN

The chemical structure of polarizing agents critically determines the efficiency of dynamic nuclear polarization (DNP). For cross-effect DNP, biradicals are the polarizing agents of choice and the interaction and relative orientation of the two unpaired electrons should be optimal. Both parameters are affected by the molecular structure of the biradical in the frozen glassy matrix that is typically used for DNP/MAS NMR and likely differs from the structure observed with X-ray crystallography. We have determined the conformations of six bis-nitroxide polarizing agents, including the highly efficient AMUPol, in their DNP matrix with EPR spectroscopy at 9.7 GHz, 140 GHz, and 275 GHz. The multi-frequency approach in combination with an advanced fitting routine allows us to reliably extract the interaction and relative orientation of the nitroxide moieties. We compare the structures of six bis-nitroxides to their DNP performance at 500 MHz/330 GHz.

5.
Phys Chem Chem Phys ; 19(36): 24545-24554, 2017 Sep 20.
Artículo en Inglés | MEDLINE | ID: mdl-28852751

RESUMEN

X-ray diffraction (XRD) and electron paramagnetic resonance spectroscopy (EPR) were combined to study the structural transformations induced by temperature, pressure and air humidity of the "breathing" metal-organic framework (MOF) MIL-53(Al), doped with paramagnetic VIV ions, after activation. The correlation between in situ XRD and thermogravimetric analysis measurements showed that upon heating this MOF in air, starting from ambient temperature and pressure, the narrow pore framework first dehydrates and after that makes the transition to a large pore state (lp). The EPR spectra of VIV[double bond, length as m-dash]O molecular ions, replacing Al-OH in the structure, also allow to distinguish the as synthesized, hydrated (np-h) and dehydrated narrow pore (np-d), and lp states of MIL-53(Al). A careful analysis of EPR spectra recorded at microwave frequencies between 9.5 and 275 GHz demonstrates that all VIV[double bond, length as m-dash]O in the np-d and lp states are equivalent, whereas in the np-h state (at least two) slightly different VIV[double bond, length as m-dash]O sites exist. Moreover, the lp MIL-53(Al) framework is accessible to oxygen, leading to a notable broadening of the VIV[double bond, length as m-dash]O EPR spectrum at pressures of a few mbar, while such effect is absent for the np-h and np-d states for pressures up to 1 bar.

6.
Appl Magn Reson ; 47: 643-653, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27340337

RESUMEN

Rapid freeze-quench (RFQ) in combination with electron paramagnetic resonance (EPR) spectroscopy at X-band is a proven technique to trap and characterize paramagnetic intermediates of biochemical reactions. Preparation of suitable samples is still cumbersome, despite many attempts to remedy this problem, and limits the wide applicability of RFQ EPR. We present a method, which improves the collection of freeze-quench particles from isopentane and their packing in an EPR tube. The method is based on sucking the particle suspension into an EPR tube with a filter at the bottom. This procedure results in a significant reduction of the required volume of reactants, which allows the economical use of valuable reactants such as proteins. The approach also enables the successful collection of smaller frozen particles, which are generated at higher flow rates. The method provides for a reproducible, efficient and fast collection of the freeze-quench particles and can be easily adapted to RFQ EPR at higher microwave frequencies than X-band.

7.
J Biol Inorg Chem ; 20(3): 487-96, 2015 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-25537134

RESUMEN

We report 275 GHz EPR spectra of human serum transferrin. At this high microwave frequency the zero-field splitting between the magnetic sublevels of the high-spin [Formula: see text] sites can be accurately determined. We find the zero-field splitting to be a sensitive probe of the structure of the transferrin iron-binding sites. Signals arising from iron bound to the transferrin N-lobe can clearly be distinguished from signals from iron bound to the C-lobe. Moreover, our spectra show that the structure of the iron site in the N-lobe is influenced by the presence and conformation of the C-lobe. The spectra of a series of N-lobe mutants altering the second-shell interaction of Arg124 with the synergistic anion carbonate reflect conformational changes induced at the iron site.


Asunto(s)
Espectroscopía de Resonancia por Spin del Electrón , Compuestos Férricos/química , Modelos Moleculares , Transferrina/química , Sitios de Unión , Análisis Químico de la Sangre , Humanos , Transferrina/genética , Transferrina/metabolismo
8.
J Chem Phys ; 142(23): 234201, 2015 Jun 21.
Artículo en Inglés | MEDLINE | ID: mdl-26093552

RESUMEN

We develop a theoretical description of Dynamic Nuclear Polarization (DNP) in solids under Magic Angle Spinning (MAS) to describe the magnetic field dependence of the DNP effect. The treatment is based on an efficient scheme for numerical solution of the Liouville-von Neumann equation, which explicitly takes into account the variation of magnetic interactions during the sample spinning. The dependence of the cross-effect MAS-DNP on various parameters, such as the hyperfine interaction, electron-electron dipolar interaction, microwave field strength, and electron spin relaxation rates, is analyzed. Electron spin relaxation rates are determined by electron paramagnetic resonance measurements, and calculations are compared to experimental data. Our results suggest that the observed nuclear magnetic resonance signal enhancements provided by MAS-DNP can be explained by discriminating between "bulk" and "core" nuclei and by taking into account the slow DNP build-up rate for the bulk nuclei.

9.
Proc Natl Acad Sci U S A ; 106(52): 22281-6, 2009 Dec 29.
Artículo en Inglés | MEDLINE | ID: mdl-20018724

RESUMEN

Composed of the two bacteriochlorophyll cofactors, P(L) and P(M), the special pair functions as the primary electron donor in bacterial reaction centers of purple bacteria of Rhodobacter sphaeroides. Under light absorption, an electron is transferred to a bacteriopheophytin and a radical pair is produced. The occurrence of the radical pair is linked to the production of enhanced nuclear polarization called photochemically induced dynamic nuclear polarization (photo-CIDNP). This effect can be used to study the electronic structure of the special pair at atomic resolution by detection of the strongly enhanced nuclear polarization with laser-flash photo-CIDNP magic-angle spinning NMR on the carotenoid-less mutant R26. In the electronic ground state, P(L) is strongly disturbed, carrying a slightly negative charge. In the radical cation state, the ratio of total electron spin densities between P(L) and P(M) is 2:1, although it is 2.5:1 for the pyrrole carbons, 2.2:1 for all porphyrinic carbons, and 4:1 for the pyrrole nitrogen. It is shown that the symmetry break between the electronic structures in the electronic ground state and in the radical cation state is an intrinsic property of the special pair supermolecule, which is particularly attributable to a modification of the structure of P(L). The significant difference in electron density distribution between the ground and radical cation states is explained by an electric polarization effect of the nearby histidine.


Asunto(s)
Proteínas del Complejo del Centro de Reacción Fotosintética/química , Proteínas del Complejo del Centro de Reacción Fotosintética/metabolismo , Rhodobacter sphaeroides/metabolismo , Proteínas Bacterianas/química , Bacterioclorofila A/química , Fenómenos Biofísicos , Transporte de Electrón , Modelos Moleculares , Mutación , Resonancia Magnética Nuclear Biomolecular , Procesos Fotoquímicos , Fotosíntesis , Proteínas del Complejo del Centro de Reacción Fotosintética/genética , Rhodobacter sphaeroides/genética , Electricidad Estática
10.
J Phys Chem A ; 115(34): 9552-6, 2011 Sep 01.
Artículo en Inglés | MEDLINE | ID: mdl-21604722

RESUMEN

We compare the resonance Raman spectra acquired at two excitation wavelengths, 496.5 and 514.5 nm, of spheroidene in the wild-type reaction center of Rhodobacter sphaeroides and reconstituted into the reaction center of the Rhodobacter sphaeroides mutant R26. Our earlier work showed that the reconstituted R26 reaction center holds spheroidene in two configurations: 15,15'-cis and another configuration. Here we show that in the wild-type reaction center only 15,15'-cis spheroidene is present. In the resonance Raman spectra of the reconstituted R26 reaction centers, a transition is identified that arises exclusively from the second configuration. According to density-functional-theory calculations, this transition is specific for the 13,14-cis configuration.


Asunto(s)
Carotenoides/química , Fotosíntesis/fisiología , Proteínas del Complejo del Centro de Reacción Fotosintética/química , Rhodobacter sphaeroides/química , Radioisótopos de Carbono/análisis , Carotenoides/metabolismo , Conformación Molecular , Proteínas del Complejo del Centro de Reacción Fotosintética/metabolismo , Rhodobacter sphaeroides/metabolismo , Espectrometría Raman , Tritio/análisis
11.
Appl Magn Reson ; 38(1): 105-116, 2010 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-20208980

RESUMEN

Photochemically induced dynamic nuclear polarization magic-angle spinning nuclear magnetic resonance (photo-CIDNP MAS NMR) allows for the investigation of the electronic structure of the photochemical machinery of photosynthetic reaction centers (RCs) at atomic resolution. For such experiments, either continuous radiation from white xenon lamps or green laser pulses are applied to optically dense samples. In order to explore their optical properties, optically thick samples of isolated and quinone-removed RCs of the purple bacteria of Rhodobacter sphaeroides wild type are studied by nanosecond laser-flash (13)C photo-CIDNP MAS NMR using excitation wavelengths between 720 and 940 nm. Action spectra of both the transient nuclear polarization as well as the nuclear hyperpolarization, remaining in the electronic ground state at the end of the photocycle, are obtained. It is shown that the signal intensity is limited by the amount of accessible RCs and that the different mechanisms of the photo-CIDNP production rely on the same photophysical origin, which is the photocycle induced by one single photon.

12.
Appl Magn Reson ; 37(1-4): 219-227, 2010 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-19960067

RESUMEN

As part of our ongoing project that aims at the optimum characterization of the electronic structure of the blue-copper site of azurin from Pseudomonas aeruginosa, we present the complete hyperfine tensors of the protons bound to the Cbeta atom of the copper-bound cysteine 112. These tensors have been obtained from a 95 GHz pulsed electron-nuclear double resonance study of a single crystal of the protein.

13.
PLoS One ; 15(5): e0232555, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32392255

RESUMEN

We report an easy, efficient and reproducible way to prepare Rapid-Freeze-Quench samples in sub-millimeter capillaries and load these into the probe head of a 275 GHz Electron Paramagnetic Resonance spectrometer. Kinetic data obtained for the binding reaction of azide to myoglobin demonstrate the feasibility of the method for high-frequency EPR. Experiments on the same samples at 9.5 GHz show that only a single series of Rapid-Freeze-Quench samples is required for studies at multiple microwave frequencies.


Asunto(s)
Espectroscopía de Resonancia por Spin del Electrón/métodos , Animales , Espectroscopía de Resonancia por Spin del Electrón/instrumentación , Congelación , Caballos , Cinética , Microondas , Mioglobina/química
14.
Biochim Biophys Acta ; 1767(6): 610-5, 2007 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-17292850

RESUMEN

Photochemically induced dynamic nuclear polarization has been observed in reaction centres of the green sulphur bacterium Chlorobium tepidum by (13)C magic-angle spinning solid-state NMR under continuous illumination with white light. An almost complete set of chemical shifts of the aromatic ring carbons of a BChl a molecule has been obtained. All light-induced (13)C NMR signals appear to be emissive, which is similar to the pattern observed in the reaction centers of plant photosystem I and purple bacterial reaction centres of Rhodobacter sphaeroides wild type. The donor in RCs of green sulfur bacteria clearly differs from the substantially asymmetric special pair of purple bacteria and appears to be similar to the more symmetric donor of photosystem I.


Asunto(s)
Chlorobi/química , Resonancia Magnética Nuclear Biomolecular/métodos , Proteínas del Complejo del Centro de Reacción Fotosintética/química , Isótopos de Carbono , Fotoquímica , Complejo de Proteína del Fotosistema I/química , Proteobacteria/química , Rhodobacter sphaeroides/química
15.
Biochim Biophys Acta ; 1757(5-6): 369-79, 2006.
Artículo en Inglés | MEDLINE | ID: mdl-16829225

RESUMEN

Femtosecond absorption difference spectroscopy was applied to study the time and spectral evolution of low-temperature (90 K) absorbance changes in isolated reaction centers (RCs) of the HM182L mutant of Rhodobacter (Rb.) sphaeroides. In this mutant, the composition of the B-branch RC cofactors is modified with respect to that of wild-type RCs by replacing the photochemically inactive BB accessory bacteriochlorophyll (BChl) by a photoreducible bacteriopheophytin molecule (referred to as PhiB). We have examined vibrational coherence within the first 400 fs after excitation of the primary electron donor P with 20-fs pulses at 870 nm by studying the kinetics of absorbance changes at 785 nm (PhiB absorption band), 940 nm (P*-stimulated emission), and 1020 nm (BA- absorption band). The results of the femtosecond measurements are compared with those recently reported for native Rb. sphaeroides R-26 RCs containing an intact BB BChl. At delay times longer than approximately 50 fs (maximum at 120 fs), the mutant RCs exhibit a pronounced BChl radical anion (BA-) absorption band at 1020 nm, which is similar to that observed for Rb. sphaeroides R-26 RCs and represents the formation of the intermediate charge-separated state P+ BA-. Femtosecond oscillations are revealed in the kinetics of the absorption development at 1020 nm and of decay of the P*-stimulated emission at 940 nm, with the oscillatory components of both kinetics displaying a generally synchronous behavior. These data are interpreted in terms of coupling of wave packet-like nuclear motions on the potential energy surface of the P* excited state to the primary electron-transfer reaction P*-->P+ BA- in the A-branch of the RC cofactors. At very early delay times (up to 80 fs), the mutant RCs exhibit a weak absorption decrease around 785 nm that is not observed for Rb. sphaeroides R-26 RCs and can be assigned to a transient bleaching of the Qy ground-state absorption band of the PhiB molecule. In the range of 740-795 nm, encompassing the Qy optical transitions of bacteriopheophytins HA, HB, and PhiB, the absorption difference spectra collected for mutant RCs at 30-50 fs resemble the difference spectrum of the P+ PhiB- charge-separated state previously detected for this mutant in the picosecond time domain (E. Katilius, Z. Katiliene, S. Lin, A.K.W. Taguchi, N.W. Woodbury, J. Phys. Chem., B 106 (2002) 1471-1475). The dynamics of bleaching at 785 nm has a non-monotonous character, showing a single peak with a maximum at 40 fs. Based on these observations, the 785-nm bleaching is speculated to reflect reduction of 1% of PhiB in the B-branch within about 40 fs, which is earlier by approximately 80 fs than the reduction process in the A-branch, both being possibly linked to nuclear wave packet motion in the P* state.


Asunto(s)
Cromatóforos Bacterianos/fisiología , Bacterioclorofilas/fisiología , Feofitinas/fisiología , Proteínas del Complejo del Centro de Reacción Fotosintética/fisiología , Pigmentos Biológicos/metabolismo , Rhodobacter sphaeroides/fisiología , Cromatóforos Bacterianos/genética , Bacterioclorofilas/genética , Transporte de Electrón , Cinética , Mutagénesis Sitio-Dirigida , Feofitinas/genética , Proteínas del Complejo del Centro de Reacción Fotosintética/genética , Pigmentos Biológicos/genética , Rhodobacter sphaeroides/genética , Análisis Espectral
16.
J Phys Chem B ; 111(35): 10606-14, 2007 Sep 06.
Artículo en Inglés | MEDLINE | ID: mdl-17696523

RESUMEN

During the photocycle of quinone-blocked photosynthetic reaction centers (RCs), photochemically induced dynamic nuclear polarization (photo-CIDNP) is produced by polarization transfer from the initially totally electron polarized electron pair and can be observed by 13C magic-angle spinning (MAS) NMR as a strong modification of signal intensities. The same processes creating net nuclear polarization open up light-dependent channels for polarization loss. This leads to coherent and incoherent enhanced signal recovery, in addition to the recovery due to light-independent longitudinal relaxation. Coherent mixing between electron and nuclear spin states due to pseudosecular hyperfine coupling within the radical pair state provides such a coherent loss channel for nuclear polarization. Another polarization transfer mechanism called differential relaxation, which is based on the long lifetime of the triplet state of the donor, provides an efficient incoherent relaxation path. In RCs of the purple bacterium Rhodobacter sphaeroides R26, the photochemical active channels allow for accelerated signal scanning by a factor of 5. Hence, photo-CIDNP MAS NMR provides the possibility to drive the NMR technique beyond the T1 limit.

17.
Biochim Biophys Acta ; 1709(2): 105-12, 2005 Sep 05.
Artículo en Inglés | MEDLINE | ID: mdl-16095558

RESUMEN

The triplet states of photosystem II core particles from spinach were studied using time-resolved cw EPR technique at different reduction states of the iron--quinone complex of the reaction center primary electron acceptor. With doubly reduced primary acceptor, the well-known photosystem II triplet state characterised by zero-field splitting parameters |D|=0.0286 cm(-1), |E|=0.0044 cm(-1) was detected. When the primary acceptor was singly reduced either chemically or photochemically, a triplet state of a different spectral shape was observed, bearing the same D and E values and characteristic spin polarization pattern arising from RC radical pair recombination. The latter triplet state was strongly temperature dependent disappearing at T=100 K, and had a much faster decay than the former one. Based on its properties, this triplet state was also ascribed to the photosystem II reaction center. A sequence of electron-transfer events in the reaction centers is proposed that explains the dependence of the triplet state properties on the reduction state of the iron--quinone primary acceptor complex.


Asunto(s)
Complejo de Proteína del Fotosistema II/química , Spinacia oleracea/enzimología , Benzoquinonas/química , Espectroscopía de Resonancia por Spin del Electrón , Transporte de Electrón , Hierro/química , Temperatura
18.
J Magn Reson ; 180(2): 178-85, 2006 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-16515869

RESUMEN

The distance between the paramagnetic state of a native cofactor and a spin label is measured in the photosynthetic reaction centre from the bacterium Rhodobacter sphaeroides R26. A two-frequency pulsed electron paramagnetic resonance method [double-electron-electron spin resonance (DEER)] is used. A distance of 3.05 nm between the semiquinone anion state of the primary acceptor (Q(A)) and the spin label at the native cysteine at position 156 in the H-subunit is found. Molecular-dynamics (MD) simulations are performed to interpret the distance. A 6 ns run comprising the entire RC protein yields a distance distribution that is close to the experimental one. The average distance found by the MD simulation is smaller than the distance obtained by DEER by at least 0.2 nm. To better represent the experiments performed at low temperature (60K), a MD method to mimic the freezing-in of the room-temperature conformations is introduced. Both MD methods yield similar distances, but the second method has a trend towards a wider distance distribution.


Asunto(s)
Espectroscopía de Resonancia por Spin del Electrón/métodos , Proteínas del Complejo del Centro de Reacción Fotosintética/química , Rhodobacter sphaeroides/química , Marcadores de Spin , Aniones , Radicales Libres , Zinc/química
19.
Biochim Biophys Acta ; 1557(1-3): 1-12, 2003 Mar 06.
Artículo en Inglés | MEDLINE | ID: mdl-12615343

RESUMEN

The bacteriopheophytin a molecules at the H(A) and H(B) binding sites of reaction centers (RCs) of the Y(M210)W mutant of Rhodobacter sphaeroides were chemically exchanged with plant pheophytin a. The Y(M210)W mutation slows down the formation of H(A)(-), presumably by raising the free energy level of the P(+)B(A)(-) state above that of P* due to increasing the oxidation potential of the primary electron donor P and lowering the reduction potential of the accessory bacteriochlorophyll B(A). Exchange of the bacteriopheophytins with pheophytin a on the contrary lowers the redox potential of H(A), inhibiting its reduction. A combination of the mutation and pigment exchange was therefore expected to make the A-side of the RC incapable of electron transfer and cause the excited state P* to deactivate directly to the ground state or through the B-side, or both. Time-resolved absorption difference spectroscopy at 10 K on the RCs that were modified in this way showed a lifetime of P* lengthened to about 500 ps as compared to about 200 ps measured in the original Y(M210)W RCs. We show that the decay of P* in the pheophytin-exchanged preparations is accompanied by both return to the ground state and formation of a new charge-separated state, the absorption difference spectrum of which is characterized by bleachings at 811 and 890 nm. This latter state was formed with a time constant of ca. 1.7 ns and a yield of about 30%, and lasted a few nanoseconds. On the basis of spectroscopic observations these bands at 811 and 890 nm are tentatively attributed to the presence of the P(+)B(B)(-) state, where B(B) is the accessory bacteriochlorophyll in the "inactive" B-branch of the cofactors. The B(B) molecules in Y(M210)W RCs are suggested to be spectrally heterogeneous, absorbing in the Q(y) region at 813 or 806 nm. The results are discussed in terms of perturbation of the free energy level of the P(+)B(B)(-) state and absorption properties of the B(B) bacteriochlorophyll in the mutant RCs due to a long-range effect of the Y(M210)W mutation on the protein environment of the B(B) binding pocket.


Asunto(s)
Feofitinas/metabolismo , Proteínas del Complejo del Centro de Reacción Fotosintética/metabolismo , Rhodobacter sphaeroides/metabolismo , Transporte de Electrón , Complejos de Proteína Captadores de Luz , Mutación , Feofitinas/química , Proteínas del Complejo del Centro de Reacción Fotosintética/química , Plantas/metabolismo , Espectrofotometría , Temperatura , Factores de Tiempo
20.
J Phys Chem B ; 109(15): 7535-9, 2005 Apr 21.
Artículo en Inglés | MEDLINE | ID: mdl-16851865

RESUMEN

The transient radical pair P(+)Q(A)(-) in the photosynthetic reaction center from Rhodobacter sphaeroides R26 was studied over a wide temperature range using out-of-phase electron spin-echo envelope modulation (ESEEM) spectroscopy. This method is sensitive to the magnetic dipole-dipole interaction between the two electron spins of the pair and allows precise determination of the distance in the pair P(+)Q(A)(-). The out-of-phase data were complemented by normal in-phase ESEEM spectra from the two stable radicals of P(+) and Q(A)(-). The results seem to indicate that the radical pair undergoes a noticeable molecular motion around 200 K that may be characterized by a change in the distance in the pair by approximately 0.3 nm. As the two cofactors, P(+) and Q(A)(-), are held in a well-defined relative position by the reaction center protein, this means that the protein becomes flexible at 200 K. This effect may be ascribed to a dynamic glass transition around 200 K. The relation with the temperature dependence of the back reaction of P(+)Q(A)(-) is discussed.


Asunto(s)
Química Física/métodos , Proteínas del Complejo del Centro de Reacción Fotosintética/química , Rhodobacter sphaeroides/fisiología , Bioquímica/métodos , Simulación por Computador , Electrones , Análisis de Fourier , Radicales Libres , Oxidación-Reducción , Proteínas/química , Rhodobacter sphaeroides/metabolismo , Análisis Espectral , Temperatura
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