In Vivo, in Vitro, and Calculated Vibrational Spectra of Plastoquinone and the Plastosemiquinone Anion Radical

Abstract

Plastoquinone (PQ-9) is active as an electron/proton transfer component in photosynthetic membranes. For example, in the photosynthetic complex, photosystem II (PSII), PQ-9 acts as Q A , a one-electron acceptor, and as Q B , a two electron, two proton accepting species. Light-minus-dark difference Fourier transform infrared (FT-IR) spectroscopy is a technique with which mechanistic information can be obtained concerning PSII. Here, we present combined experimental and computational studies designed to identify the vibrational contributions of the electron acceptor, Q A , in its oxidized and one-electron reduced states to the difference FT-IR spectrum. Infrared spectra of decyl-PQ and PQ-9 were obtained; the difference infrared spectra associated with the formation of the corresponding anion radicals were also generated in ethanol solutions. Vibrational mode assignments were made based on hybrid Hartree-Fock/density functional (HF/DF) B3LYP calculations with a 6-31G(d) basis set. Calculations were performed for hydrogen bonded models of PQ-1 and its radical anion. In addition, a methionine-tolerant strain of the cyanobacterium, Synechocystis sp. PCC 6803, was used to deuterate PQ-9 in PSII. The macrocycle and phytol tail of chlorophyll were not labeled by this procedure. Mass spectral data may be consistent with partial 13 3 methoxy labeling of chlorophyll. Lack of phytol labeling implies that carotenoids were unlabeled. Difference FT-IR spectra were then obtained by illumination at 80 K, resulting in the one-electron reduction of Q A . When spectra were obtained of PSII preparations, in which 39% of PQ was 2 H 3 labeled and 48% was 2 H 6 labeled, isotope-induced shifts were observed. Comparison of these data to vibrational spectra obtained in vitro and to mode frequencies and intensities from B3LYP/ 6-31G(d) calculations provides the basis for vibrational mode assignments.Photosystem II (PSII), a membrane-associated pigmentprotein complex, carries out the oxidation of water and reduction of PQ-9 (plastoquinone-9) in all oxygen-evolving plants, algae, and cyanobacteria. Photoexcitation of the primary electron donor, P 680 , results in electron transfer to a bound PQ-9, called Q A , via a pheophytin molecule. Reduced Q A is reoxidized by an exchangeable PQ-9, named Q B . Q A functions as a oneelectron acceptor, and the reduced form, Q A -, is an unprotonated semiquinone anion radical. Q B , on the other hand, is a twoelectron, two-proton acceptor {reviewed in ref 1}. Electron transfer events on the acceptor side of PSII resemble reactions occurring on the acceptor side of the photosynthetic bacterial reaction center. 2 This enzyme, for which high-resolution structural information is available, uses UQ (ubiquinone) or menaquinone, instead of PQ-9, as acceptor molecules {reviewed in ref 3}.On the donor side of PSII, P 680 + is reduced by a redox-active tyrosine, Z. 4-8 The tyrosine radical, Z • , is reduced by a multinuclear manganese cluster on the microsecond to millisecond time range {see ref 9 and references ther...

A hydrogen-bonding network plays a catalytic role in photosynthetic oxygen evolution

“…It has been concluded previously that this spectrum, acquired in the presence of the exogenous electron acceptor, potassium ferricyanide, is dominated by OEC contributions in the 1;800 − 1;200 cm −1 region (23). Potential acceptor side contributions from Q A − and Q A have been assigned by isotopic labeling of the quinones at 77 K (24,25). Through previous global 13 C labeling experiments, bands in the 1;650 cm −1 region were attributed to amide I modes (C ¼ O) of the polypeptide backbone on the PSII donor side (26).…”

Directly probing redox-linked quinones in photosystem II membrane fragments via UV resonance Raman scattering

“…It is noteworthy that another vibrational spectroscopy, FTIR, has been extensively used to characterize the chemical bonds in PSII. However, for plant PSII, the infrared region around 1660 cm −1 overlaps with amide I absorption, making FTIR experiments challenging in this region [21,38]. While the assignment of the Q A band has been proposed previously [21], this conclusion was based on isotope-editing and FTIR spectroscopy in a congested amide I region.…”

“…To distinguish the spectral contribution from PQs, a PQ derivative, decyl-PQ, was characterized for comparison since decyl-PQ has similar spectral features as PQ [21]. As shown in Fig.…”

“…Similarly, PQ possesses a strongest UV absorption band at around 255 nm [18,19], whereas, reduction of PQ shifts its maximum absorption to longer wavelengths as 288, 320, 412 and 436 nm [18][19][20][21]. To obtain more information concerning the chemical bonds of the redox-linked quinone cofactors of PSII, we exploited 257-nm ultraviolet resonance Raman (UVRR) spectroscopy with a sample jet technique to characterize the PSII enzyme (Fig.…”

“…1b), because 257-nm excitation is expected to resonantly enhance the spectral contribution of PQ. However, the signals of reduced PQ species (e.g., PQ − and PQH 2 ) are not expected to be resonantly enhanced because their electronic transition absorption bands are far away from the Raman excitation wavelength of 257 nm [18][19][20][21].…”