I. V. Borovykh

Selective excitation in pulsed EPR of spin-correlated radical pairs: electron–electron interactions, zero-, single-, and double-quantum relaxation and spectral diffusion

Experiments are described in which a low-amplitude microwave pulse excites only one out of four allowed transitions of a spin-correlated radical pair (SCRP). A second high-amplitude pulse produces an FID whose temporal shape is strongly modulated with frequencies determined by electron–electron spin-exchange and dipolar interactions. The dependence of the FID intensity on the delay between the two pulses is determined by the relaxation of populations of energy levels connected by allowed single-quantum transitions and by forbidden zero- and double-quantum transitions. The first direct measurement of the zero-quantum population relaxation rate for P+QA− radical pairs in bacterial photosynthetic reaction centers shows it is temperature independent.

Light-induced structural changes in photosynthetic reaction centres studied by ESEEM of spin-correlated D+QA− radical pairs

Zn-substituted Rhodobacter sphaeroides R26 reaction centres (RCs) frozen in the dark and under illumination exhibit quite different recombination kinetics of the D+QA- radical pairs [Kleinfeld et al., Biochemistry, 23 (1984) 5780]. We have applied electron spin echo envelope modulation (ESEEM) of the spin-correlated D+QA- radical pairs to assess a possible light-induced change in the distance between the D and QA cofactors. The recombination kinetics and the field-swept spin-polarized EPR signal for the two preparations have been monitored by time-resolved EPR spectroscopy. For the samples frozen under illumination, a slight increase in the distance, 0.4+/-0.2 A, has been detected.

The dynamical transition in proteins of bacterial photosynthetic reaction centers observed by echo-detected EPR of specific spin labels

An echo-detected electron paramagnetic resonance (ED EPR) approach was used to study molecular dynamics in photosynthetic reaction centers (RCs) fromRhodobacter sphaeroides R26, employing the specific spin label methanethiosulfonate and 3-maliemido proxyl. ED EPR has recently been shown to be sensitive to so-called dynamical transition in disordered media, which is characterized by the transition from a harmonic-like librational motion of a molecule to an anharmonic one or to a stochastic wobbling motion. ED EPR line shapes studied over a wide temperature range reveal a sharp transition occurring above 180 K. The possible relation of the found transition to the temperature dependence of electron transfer reactions in RC is discussed.

Conformation transition in the protein of a photosynthetic reaction center observed at the nanometer range of distances at cryogenic temperatures

Abstract Electron spin echo (ESE) spectroscopy is applied to study magnetic dipolar interaction between electron spins in the transient P + Q A − radical pairs (P is the primary donor, a bacteriochlorophyll dimer, and Q A is the primary quinone acceptor) and the 3 PQ A − triplet-radical pairs in bacterial photosynthetic reaction centers of Rhodobacter sphaeroides R26. Distance separation in both pairs is about 29 A. A well-resolved reversible conformational transition of the reaction center protein holding the P and Q A cofactors was observed between 13 and 20 K. This transition results in a narrowing of the distribution of protein conformations with decreasing temperature, with the width of distance distribution between P and Q A dropping from ca. 4 A at 20 K to ca. 1 A at 13 K. This transition implies the existence of low barriers in the protein energy landscape and the presence of cooperatively rearranging domains of the size of several nm both in the protein and in the surrounding glassy environment.

Selective excitation in pulsed EPR of a spin-correlated triplet-radical pair.

Experiments are described in which a low-amplitude microwave pulse excites only one out of three allowed transitions of the quinone radical (Q(A)(-)) in a spin-correlated triplet-radical pair 3PQ(A)(-) of the bacterial photosynthetic reaction center. A second high-amplitude pulse produces a FID whose temporal shape is strongly modulated with frequencies determined by electron-electron dipolar interaction in the pair. The FID is detected in both the in-phase and the out-of-phase channels. The out-of-phase FID is a result of switching off the magnetic dipolar interaction between 3P and Q(A)(-) due to decay of 3P during the time interval between the two pulses. Refocusing of FID by an additional non-selective pulse allows a dead-time free measurement of this modulation. The influence of the dead-time problem on the distance determination is discussed.

PHOTOSELECTION EFFECTS IN LASER FLASH-INDUCED SPIN-POLARISED EPR SPECTRA OF THE RADICAL PAIR STATE P+QA- IN RHODOBACTER SPHAEROIDES REACTION CENTERS

Abstract The spin-polarised EPR signal of the radical pair state [P + Q A − ] in Zn-substituted Rhodobacter sphaeroides R26 reaction centers demonstrates a strong dependence on the wavelength and polarisation of the excitation light. This dependence is attributed to photoselection by the plane-polarised laser light utilised in the experiments. The model of spin-correlated radical pairs describes the experimental results well if the effect of photoselection is taken into account.

Photoselection in ESP Spectra of Photosynthetic Reaction Centers

The first relatively stable stage of photoinduced electron transfer in reaction centers (RCs) of many photosynthetic bacteria is P+Q A - , where P is the so — called primary donor (dimer of bacteriochlorophyll molecules), and Q A - primary acceptor, a molecule of quinone. As early as 1977 [1], it was reported that when the normally present magnetic interaction between Q A - and paramagnetic Fe2+ — ion is disrupted, an electron spin — polarised (ESP) photoinduced signal may be detected using time — resolved EPR. This signal was subsequently attributed to the state P+Q A - , which since 1987 [2] is considered as a spin — correlated radical pair (SCRP). Simulation of experimental ESP spectra of P+Q A - using the SCRP model, will provide information on the structural organisation of RC and on interactions between its cofactors. Recently we observed that the shape of the ESP signal of P+Q A - in Zn2+ — substituted RCs of Rb. sphaeroides R26 strongly depends on the wavelength of the excitation laser flash. This dependence arises due to the effect of photoselection, i.e., selective excitation of certain orientations of RCs in the sample by plane — polarised laser light. When taking photoselection into account, the SCRP model satisfactorily describes the quite different ESP signals for light polarised parallel and perpendicular to the magnetic field of the spectrometer.

Photoselection Effects in Epr-Detected Triplet States of Photosynthetic Pigment Molecules

Photosynthetic reaction centers (RCs) transform the energy of light absorbed by pigment molecules into chemical energy. RCs of bacteria are best characterised both functionally and structurally. In them, excitation of the primary donor P (dimer of bacteriochlorophylls) leads to a sequence of electron-transfer reactions to intermediate acceptor (bacteriopheophytin) and then to a quinone primary acceptor. When electron transfer past the intermediate acceptor is blocked, reverse electron transfer to oxidised P occurs, populating the triplet state of P (at 1<77 K with almost-unity yield). Though the triplet states are the side products, their properties make them excellent probes of the RC photochemistry (for review see [1, 2]). Magnetophotoselection, MPS, or EPR detection of triplet states excited with polarised light, proved to be a powerful technique for the studies of the triplet states. Usually MPS is performed utilising cw EPR detection [1]. First application of time-resolved direct-detection EPR (DD-EPR) to the study of a synthetic aromatic molecule was reported in [3]. DD-EPR is advantageous in such studies, for it is essentially free from a drawback of cw EPR MPS, the latter’s strong dependence on spin-lattice relaxation. This enables investigations in a wide temperature range. Here we report on the first observation of DD-EPR MPS of the primary donor triplet state of Rb. sphaeroides R26 RCs and D1/D2-cyt b-559 complexes of plant photosystem II.

Electron Spin Polarisation in Quinone Reconstituted Photosynthetic Reaction Centers of RB. Sphaeroides R26.

Light excitation of the primary bacteriochlorophyll dimer, P, in photosynthetic reaction centers (RCs) of Rhodobacter (Rb.) sphaeroides R26 triggers a series of electron transfer events in the reaction center: Open image in new window in which I, QA and QB are a bacteriopheophytin, a primary and a secondary electron acceptor quinone, respectively. Each state is characterised by a lifetime k-1. The secondary radical pair P+·QA-·which is generated in this process has been studied extensively by time resolved EPR spectroscopy. The transient EPR spectrum of P·Q-·A shows a high degree of electron spin polarisation (ESP) due to magnetic interactions between the two spins. Simulation of experimental ESP spectra, using the spin-correlated radical pair model [1], have yielded valuable information about the mutual orientation, and magnetic and electronic interactions of both radicals. However, in most of these studies, the lifetime of the primary radical pair, P+·I -·, was assumed to be much shorter compared to the timescale of its magnetic interactions. Thus, the observed secondary radical pair was considered to be initially in a pure singlet state and any admixture of triplet character was neglected. We have investigated the effect of the intermediate radical pair P+·I-· on the ESP spectrum of P·QA-· in Zn-substituted RCs of Rb. sphaeroides R26 by prolongation of its lifetime by replacing the native ubiquinone-10 with different quinones [2]. To simulate the observed X-band ESP spectra we used the theoretical description of the transfer of electron spin-correlation from one radical pair to another presented by Hore [3]. Furthermore, the recently discovered effect magnetophotoselection on the laser flash-induced ESP spectra was taken into account [4].