Julian J. Eaton-Rye

The extrinsic proteins of Photosystem II

In this review we examine the structure and function of the extrinsic proteins of Photosystem II. These proteins include PsbO, present in all oxygenic organisms, the PsbP and PsbQ proteins, which are found in higher plants and eukaryotic algae, and the PsbU, PsbV, CyanoQ, and CyanoP proteins, which are found in the cyanobacteria. These proteins serve to optimize oxygen evolution at physiological calcium and chloride concentrations. They also shield the Mn(4)CaO(5) cluster from exogenous reductants. Numerous biochemical, genetic and structural studies have been used to probe the structure and function of these proteins within the photosystem. We will discuss the most recent proposed functional roles for these components, their structures (as deduced from biochemical and X-ray crystallographic studies) and the locations of their proposed binding domains within the Photosystem II complex. This article is part of a Special Issue entitled: Photosystem II.

Manganese Compounds as Water-Oxidizing Catalysts: From the Natural Water-Oxidizing Complex to Nanosized Manganese Oxide Structures

All cyanobacteria, algae, and plants use a similar water-oxidizing catalyst for water oxidation. This catalyst is housed in Photosystem II, a membrane-protein complex that functions as a light-driven water oxidase in oxygenic photosynthesis. Water oxidation is also an important reaction in artificial photosynthesis because it has the potential to provide cheap electrons from water for hydrogen production or for the reduction of carbon dioxide on an industrial scale. The water-oxidizing complex of Photosystem II is a Mn-Ca cluster that oxidizes water with a low overpotential and high turnover frequency number of up to 25-90 molecules of O2 released per second. In this Review, we discuss the atomic structure of the Mn-Ca cluster of the Photosystem II water-oxidizing complex from the viewpoint that the underlying mechanism can be informative when designing artificial water-oxidizing catalysts. This is followed by consideration of functional Mn-based model complexes for water oxidation and the issue of Mn complexes decomposing to Mn oxide. We then provide a detailed assessment of the chemistry of Mn oxides by considering how their bulk and nanoscale properties contribute to their effectiveness as water-oxidizing catalysts.

Photosystem II and the unique role of bicarbonate: a historical perspective.

In photosynthesis, cyanobacteria, algae and plants fix carbon dioxide (CO(2)) into carbohydrates; this is necessary to support life on Earth. Over 50 years ago, Otto Heinrich Warburg discovered a unique stimulatory role of CO(2) in the Hill reaction (i.e., O(2) evolution accompanied by reduction of an artificial electron acceptor), which, obviously, does not include any carbon fixation pathway; Warburg used this discovery to support his idea that O(2) in photosynthesis originates in CO(2). During the 1960s, a large number of researchers attempted to decipher this unique phenomenon, with limited success. In the 1970s, Alan Stemler, in Govindjees lab, perfected methods to get highly reproducible results, and observed, among other things, that the turnover of Photosystem II (PSII) was stimulated by bicarbonate ions (hydrogen carbonate): the effect would be on the donor or the acceptor, or both sides of PSII. In 1975, Thomas Wydrzynski, also in Govindjees lab, discovered that there was a definite bicarbonate effect on the electron acceptor (the plastoquinone) side of PSII. The most recent 1.9Å crystal structure of PSII, unequivocally shows HCO(3)(-) bound to the non-heme iron that sits in-between the bound primary quinone electron acceptor, Q(A), and the secondary quinone electron acceptor Q(B). In this review, we focus on the historical development of our understanding of this unique bicarbonate effect on the electron acceptor side of PSII, and its mechanism as obtained by biochemical, biophysical and molecular biological approaches in many laboratories around the World. We suggest an atomic level model in which HCO(3)(-)/CO(3)(2-) plays a key role in the protonation of the reduced Q(B). In addition, we make comments on the role of bicarbonate on the donor side of PSII, as has been extensively studied in the labs of Alan Stemler (USA) and Vyacheslav Klimov (Russia). We end this review by discussing the uniqueness of bicarbonates role in oxygenic photosynthesis and its role in the evolutionary development of O(2)-evolving PSII. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial.

Effects of inactivating psbM and psbT on photodamage and assembly of photosystem II in Synechocystis sp. PCC 6803.

PsbM and PsbT have been assigned to electron densities on both photosystem II (PSII) monomers at the PSII dimer interface in X-ray crystallographic structures from Thermosynechoccocus elongatus and T. vulcanus. Our results show that removal of either or both proteins from Synechocystis sp. PCC 6803 resulted in photoautotrophic strains but the DeltaPsbM:DeltaPsbT mutant did not form stable dimers. A CP43-less PSII monomer accumulated in both single mutants, although absence of PsbT destabilized PSII to a greater extent than removing PsbM. Additionally, DeltaPsbT cells exhibited slowed electron transfer between the plastoquinone electron acceptors, Q(A) and Q(B); however, S-state cycling in both mutants was similar to wild type. Oxygen evolution in these mutants rapidly inactivated following exposure to high light where recovery required protein synthesis and could proceed in the dark in DeltaPsbM cells but required light in DeltaPsbT cells. Interestingly, the extent of recovery of oxygen-evolving activity was greatest in the DeltaPsbM:DeltaPsbT strain. We also found recovery required Psb27 in DeltaPsbT cells although, under our conditions, the DeltaPsb27 strain remained similar to wild type. In contrast, the DeltaPsbM:DeltaPsb27 mutant could not assemble PSII beyond a CP43-minus intermediate. Our results suggest essential roles for Psb27 in biogenesis in the DeltaPsbM strain and for repair from photodamage in cells lacking PsbT.

Construction of gene interruptions and gene deletions in the cyanobacterium Synechocystis sp. strain PCC 6803.

A series of protocols are presented for the storage, growth, transformation, and characterization of wild type and mutant strains of Synechocystis sp. strain PCC 6803. These protocols include the isolation of genomic DNA and the strategies required for the construction of specific gene interruptions or deletions in this organism. This cyanobacterium has been used widely as a model for photosynthesis research, and the sequence of its genome is available at CyanoBase (http://genome.kazusa.or.jp/cyanobase/). The details provided in this chapter do not assume any previous experience in working with cyanobacteria and are intended to enable new investigators to take advantage of a wide range of gene modification and mutation mapping techniques that have been adapted for use in this system.

Solution Structure of Psb27 from Cyanobacterial Photosystem II

Psb27 is a highly conserved component of photosystem II. The three-dimensional structure has a well-defined helical core, composed of four helices arranged in a right-handed up-down-up-down fold, with a less ordered region of the structure located at the N-terminus. The position of conserved residues on the surface suggests conserved functional roles for distinct interconnected features encompassing a P-phi-P loop, a polar patch spanning helices alpha3 and alpha4, and the N-terminal sequence.

Investigation of a requirement for the PsbP-like protein in Synechocystis sp. PCC 6803

The sll1418 gene encodes a PsbP-like protein in Synechocystis sp. PCC 6803. Expression of sll1418 was similar in BG-11 and in Cl−- or Ca2+-limiting media, and inactivation of sll1418 did not prevent photoautotrophic growth in normal or nutrient-limiting conditions. Also the wild-type and ΔPsbP strains exhibited similar oxygen evolution and assembly of Photosystem II (PS II) centers. Inactivation of sll1418 in the ΔPsbO: ΔPsbP, ΔPsbQ:ΔPsbP, ΔPsbU:ΔPsbP and ΔPsbV:ΔPsbP mutants did not prevent photoautotrophy or alter PS II assembly and oxygen evolution in these strains. Moreover, the absence of PsbP did not affect the ability of alkaline pH to restore photoautotrophic growth in the ΔPsbO:ΔPsbU strain. The PsbO, PsbU and PsbV proteins are required for thermostability of PS II and thermal acclimation in Synechocystis sp. PCC 6803 [Kimura et al. (2002) Plant Cell Physiol 43: 932–938]. However, thermostability and thermal acclimation in ΔPsbP cells were similar to wild type. These results are consistent with the conclusion that PsbP is associated with ∼3 of PS II centers, and may play a regulatory role in PS II [Thornton et al. (2004) Plant Cell 16: 2164–2175].

The construction of gene knockouts in the cyanobacterium Synechocystis sp. PCC 6803.

A series of protocols are presented for the storage, growth, transformation, and characterization of wild-type (wt) and mutant strains of Synechocystis PCC 6803. These protocols include the isolation of genomic DNA and the strategies required for the construction of specific gene knockouts in this organism. This cyanobacterium has been used widely as a model for photosynthesis research and the sequence of its genome, together with a database of mutants that have already been constructed, is available at CyanoBase (http://kazusa.or.jp/cyano/). The details provided in this chapter do not assume any previous experience in working with cyanobacteria and are intended to enable new investigators to take advantage of a wide-range of gene modification and mutation mapping techniques that have been adapted for use in this system.

The effects of bicarbonate depletion and formate incubation on the kinetics of oxidation-reduction reactions of the photosystem II quinones acceptor complex

Chloroplast thylakoid membranes depleted of bicarbonate exhibit a slowed oxidation of the primary quinone acceptor (Qᴀ) by the secondary quinone acceptor (Qв) of photosystem II. The kinetics of these slowed reactions have been followed by using short xenon flashes of light both to excite photosystem II and to probe the redox state of Qᴀ. Thylakoids incubated with formate but not depleted of bicarbonate showed the same pattern of slowed reaction kinetics of the quinone acceptors as seen in bicarbonate-depleted| thylakoids. This led us to conclude that there was a simple competition between bicarbonate and formate at this site; however, steady-state electron transfer measured with an oxygen electrode showed that although the bicarbonate- depleted thylakoids were indeed inhibited, rates in the formate-incubated thylakoids were only slightly slowed. We suggest that the inhibition seen at the quinone acceptor site of photosystem II depends in a subtle way upon the rate of exchange of bicarbonate and formate at this site.

Phosphate sensing in Synechocystis sp. PCC 6803: SphU and the SphS–SphR two-component regulatory system

The Pho regulon is controlled by the histidine kinase-response regulator pair SphS–SphR in many cyanobacteria and up-regulation of the Pho regulon can be monitored by measuring alkaline phosphatase activity. However, the mechanism regulating signal transduction between SphS and SphR has not been described. We have created a cyanobacterial strain allowing the introduction of mutations into the transmitter domain of SphS. Mutations at Thr-167, adjacent to the H motif of SphS, introduce elevated alkaline phosphatase activity in the presence of phosphate and an enhancement of alkaline phosphatase activity, when compared to the control strain, in phosphate-limiting media. SphU acts as a negative regulator of the SphS–SphR system in Synechocystis sp. PCC 6803 and we show that constitutive alkaline phosphatase activity in the absence of SphU requires signal transduction through SphS and SphR. However, constitutive activity in the absence of SphU is severely attenuated in the ΔSphU:SphS-T167N mutant. Our data suggest that Thr-167 contributes to the mechanism underlying regulation by SphU. We have also assembled a deletion mutant system allowing the introduction of mutations into SphR and show that Gly-225 and Trp-236, which are both conserved in SphR from cyanobacteria, are essential for activation of the Pho regulon under phosphate-limiting conditions.