The midpoint potential (E m ) of Q A =Q −• A , the one-electron acceptor quinone of Photosystem II (PSII), provides the thermodynamic reference for calibrating PSII bioenergetics. Uncertainty exists in the literature, with two values differing by ∼80 mV. Here, we have resolved this discrepancy by using spectroelectrochemistry on plant PSII-enriched membranes. Removal of bicarbonate (HCO 3 − ) shifts the E m from ∼−145 mV to −70 mV. The higher values reported earlier are attributed to the loss of HCO 3 − during the titrations (pH 6.5, stirred under argon gassing). P hotosystem II (PSII), the water/plastoquinone photooxidoreductase, is at the heart of the major energy cycle that powers the biosphere. Chlorophyll-based photochemistry drives charge separation followed by electron transfer reactions that result in the reduction of quinone on one side of the thylakoid membrane ( Fig. 1) and the oxidation of water on the other. The photochemistry is intrinsically a one-photon/one-electron process, but the quinone reduction and water oxidation are two-and fourelectron processes, respectively. As a result, both processes involve the accumulation of intermediates that are stabilized by the protein and by coupling to protonation reactions (1-3).The electron acceptor side of PSII contains a nonheme ferrous iron (Fe 2+ ) that is flanked by the two quinones, Q A and Q B (Fig. 1) (reviewed in refs. 3 and 4). The Fe 2+ is coordinated by four histidine residues, two from D1 and two from D2, and a bicarbonate ion (HCO 3 − ) that provides a bidentate ligand (3-5). The HCO 3 − is thought to play a role in the Q B protonation pathway (reviewed in refs. 3-5). Recent EPR (6) and computation chemistry studies (7) based on the highest-resolution X-ray crystallographic model (8) implicated HCO 3 − in the second of two protonation steps that are associated with Q B reduction. Measurements of the dissociation constant of HCO 3 − (K d = 40-80 μM), compared with estimates of the HCO 3 − concentration in the stroma (reviewed in ref. 9), led to the assumption that HCO 3 − remains bound under physiological conditions (10). Recently, however, it was reported that the HCO 3 − bound to PSII in Chlamydomonas can be displaced by acetate when present in the culture medium (11).The redox potential of Q A has been the subject of research for decades (12). The current detailed thermodynamic picture of PSII redox chemistry is based on estimates of energy differences between the electron transfer components, but these estimates A , differing by ∼150 mV, depending on the nature of the PSII (12-14). The fully active enzyme showed an E m value that was ∼150 mV lower than that in PSII lacking the Mn 4 CaO 5 cluster. This shift was, in fact, due to the binding (or absence thereof) of the Ca 2+ ion involved in water splitting, but because the Mn cluster provides the Ca binding site, the potential is indirectly determined by the presence of Mn cluster (12)(13)(14). Given the high valence state on the Mn cluster even in the lowest redox state of the water splitti...

SummaryCph2 from the cyanobacterium Synechocystis sp. PCC 6803 is a hybrid photoreceptor that comprises an N-terminal module for red/far-red light reception and a C-terminal module switching between a blue-and a green-receptive state. This unusual photoreceptor exerts complex, light quality-dependent control of the motility of Synechocystis sp. PCC 6803 cells by inhibiting phototaxis towards blue light. Cph2 perceives blue light by its third GAF domain that bears all characteristics of a cyanobacteriochrome (CBCR) including photoconversion between green-and blueabsorbing states as well as formation of a bilin species simultaneously tethered to two cysteines, C994 and C1022. Upon blue light illumination the CBCR domain activates the subsequent C-terminal GGDEF domain, which catalyses formation of the second messenger c-di-GMP. Accordingly, expression of the CBCR-GGDEF module in Dcph2 mutant cells restores the blue light-dependent inhibition of motility. Additional expression of the N-terminal Cph2 fragment harbouring a red/far-red interconverting phytochrome fused to a c-di-GMP degrading EAL domain restores the complex behaviour of the intact Cph2 photosensor. c-di-GMP was shown to regulate flagellar and pilibased motility in several bacteria. Here we provide the first evidence that this universal bacterial second messenger is directly involved in the light-dependent regulation of cyanobacterial phototaxis.

Photosystem II (PSII), the light-driven water/plastoquinone photooxidoreductase, is of central importance in the planetary energy cycle. The product of the reaction, plastohydroquinone (PQH2), is released into the membrane from the QB site, where it is formed. A plastoquinone (PQ) from the membrane pool then binds into the QB site. Despite their functional importance, the thermodynamic properties of the PQ in the QB site, QB, in its different redox forms have received relatively little attention. Here we report the midpoint potentials (Em) of QB in PSII from Thermosynechococcus elongatus using electron paramagnetic resonance (EPR) spectroscopy: Em QB/QB•− ≈ 90 mV, and Em QB•−/QBH2 ≈ 40 mV. These data allow the following conclusions: 1) The semiquinone, QB•−, is stabilized thermodynamically; 2) the resulting Em QB/QBH2 (∼65 mV) is lower than the Em PQ/PQH2 (∼117 mV), and the difference (ΔE ≈ 50 meV) represents the driving force for QBH2 release into the pool; 3) PQ is ∼50× more tightly bound than PQH2; and 4) the difference between the Em QB/QB•− measured here and the Em QA/QA•− from the literature is ∼234 meV, in principle corresponding to the driving force for electron transfer from QA•− to QB. The pH dependence of the thermoluminescence associated with QB•− provided a functional estimate for this energy gap and gave a similar value (≥180 meV). These estimates are larger than the generally accepted value (∼70 meV), and this is discussed. The energetics of QB in PSII are comparable to those in the homologous purple bacterial reaction center.

No abstract