Net light-induced oxygen evolution in photosystem I deletion mutants of the cyanobacterium Synechocystis sp. PCC 6803.

Oxygenic photosynthesis in cyanobacteria, algae, and plants requires photosystem II (PSII) to extract electrons from H(2)O and depends on photosystem I (PSI) to reduce NADP(+). Here we demonstrate that mixotrophically-grown mutants of the cyanobacterium Synechocystis sp. PCC 6803 that lack PSI (ΔPSI) are capable of net light-induced O(2) evolution in vivo. The net light-induced O(2) evolution requires glucose and can be sustained for more than 30 min. Utilizing electron transport inhibitors and chlorophyll a fluorescence measurements, we show that in these mutants PSII is the source of the light-induced O(2) evolution, and that the plastoquinone pool is reduced by PSII and subsequently oxidized by an unidentified electron acceptor that does not involve the plastoquinol oxidase site of the cytochrome b(6)f complex. Moreover, both O(2) evolution and chlorophyll a fluorescence kinetics of the ΔPSI mutants are highly sensitive to KCN, indicating the involvement of a KCN-sensitive enzyme(s). Experiments using (14)C-labeled bicarbonate show that the ΔPSI mutants assimilate more CO(2) in the light compared to the dark. However, the rate of the light-minus-dark CO(2) assimilation accounts for just over half of the net light-induced O(2) evolution rate, indicating the involvement of unidentified terminal electron acceptors. Based on these results we suggest that O(2) evolution in ΔPSI cells can be sustained by an alternative electron transport pathway that results in CO(2) assimilation and that includes PSII, the platoquinone pool, and a KCN-sensitive enzyme.

Digital biology: an emerging and promising discipline.

This article examines the role of computation and quantitative methods in modern biomedical research to identify emerging scientific, technical, policy and organizational trends. It identifies common concerns and practices in the emerging community of computationally-oriented bio-scientists by reviewing a national symposium, Digital Biology: the Emerging Paradigm, held at the National Institutes of Health in Bethesda, Maryland, November 6th and 7th 2003. This meeting showed how biomedical computing promises scientific breakthroughs that will yield significant health benefits. Three key areas that define the emerging discipline of digital biology are: scientific data integration, multi-scale modeling and networked science. Each area faces unique technical challenges and information policy issues that must be addressed as the field matures. Here we summarize the emergent challenges and offer suggestions to academia, industry and government on how best to expand the role of computation in their scientific activities.

Cytochrome b6 arginine 214 of Synechococcus sp. PCC 7002, a key residue for quinone-reductase site function and turnover of the cytochrome bf complex.

Quinone-reductase (Q(i)) domains of cyanobacterial/chloroplast cytochrome bf and bacterial/mitochondrial bc complexes differ markedly, and the cytochrome bf Q(i) site mechanism remains largely enigmatic. To investigate the bf Q(i) domain, we constructed the mutation R214H, which substitutes histidine for a conserved arginine in the cytochrome b(6) polypeptide of the cyanobacterium Synechococcus sp. SPCC 7002. At high light intensity, the R214H mutant grew approximately 2.5-fold more slowly than the wild type. Slower growth arose from correspondingly slower overall turnover of the bf complex. Specifically, as shown in single flash turnover experiments of cytochrome b(6) reduction and oxidation, the R214H mutation partially blocked electron transfer to the Q(i) site, mimicking the effect of the Q(i) site inhibitor 2-N-4-hydroxyquinoline-N-oxide. The kinetics of cytochrome b(6) oxidation were largely unaffected by hydrogen-deuterium exchange in the mutant but were slowed considerably in the wild type. This suggests that although protonation events influenced the kinetics of cytochrome b(6) oxidation at the Q(i) site in the wild type, electron flow limited this reaction in the R214H mutant. Redox titration of membranes revealed midpoint potentials (E(m,7)) of the two b hemes similar to those in the wild type. Our data define cytochrome b(6) Arg(214) as a key residue for Q(i) site catalysis and turnover of the cytochrome bf complex. In the recent cytochrome bf structures, Arg(214) lies near the Q(i) pocket and the newly discovered c(i) or x heme. We propose a model for Q(i) site function and a role for Arg(214) in plastoquinone binding.

The slow reversibility of photosystem II thermal energy dissipation on transfer from high to low light may cause large losses in carbon gain by crop canopies: a theoretical analysis.

Regulated thermal dissipation of absorbed light energy within the photosystem II antenna system helps protect photosystem II from damage in excess light. This reversible photoprotective process decreases the maximum quantum yield of photosystem II (Fv)/Fm) and CO2 assimilation (phiCO2), and decreases the convexity of the non-rectangular hyperbola describing the response of leaf CO2 assimilation to photon flux (theta). At high light, a decrease in phiCO2 has minimal impact on carbon gain, while high thermal energy dissipation protects PSII against oxidative damage. Light in leaf canopies in the field is continually fluctuating and a finite period of time is required for recovery of phiCO2 and when light drops below excess levels. Low phiCO2) and can limit the rate of photosynthetic carbon assimilation on transfer to low light, an effect prolonged by low temperature. What is the cost of this delayed reversal of thermal energy dissipation and phiCO2 recovery to potential CO2 uptake by a canopy in the field? To address this question a reverse ray-tracing algorithm for predicting the light dynamics of 120 randomly selected individual points in a model canopy was used to describe the discontinuity and heterogeneity of light flux within the canopy. Because photoprotection is at the level of the cell, not the leaf, light was simulated for small points of 10(4) micro m rather than as an average for a leaf. The predicted light dynamics were combined with empirical equations simulating the dynamics of the light-dependent decrease and recovery of phiCO2 and and their effects on the integrated daily canopy carbon uptake (A'c). The simulation was for a model canopy of leaf area index 3 with random inclination and orientation of foliage, on a clear sky day (latitude 44 degrees N, 120th day of the year). The delay in recovery of photoprotection was predicted to decrease A'c by 17% at 30 degrees C and 32% at 10 degrees C for a chilling-susceptible species, and by 12.8% at 30 degrees C and 24% at 10 degrees C for a chilling-tolerant species. These predictions suggest that the selection, or engineering, of genotypes capable of more rapid recovery from the photoprotected state would substantially increase carbon uptake by crop canopies in the field.

Proteomic analysis of a highly active photosystem II preparation from the cyanobacterium Synechocystis sp. PCC 6803 reveals the presence of novel polypeptides.

A highly active oxygen-evolving photosystem II (PSII) complex was purified from the HT-3 strain of the widely used cyanobacterium Synechocystis sp. PCC 6803, in which the CP47 polypeptide has been genetically engineered to contain a polyhistidine tag at its carboxyl terminus [Bricker, T. M., Morvant, J., Masri, N., Sutton, H. M., and Frankel, L. K. (1998) Biochim. Biophys. Acta 1409, 50-57]. These purified PSII centers had four manganese atoms, one calcium atom, and two cytochrome b(559) hemes each. Optical absorption and fluorescence emission spectroscopy as well as western immunoblot analysis demonstrated that the purified PSII preparation was devoid of any contamination with photosystem I and phycobiliproteins. A comprehensive proteomic analysis using a system designed to enhance resolution of low-molecular-weight polypeptides, followed by MALDI mass spectrometry and N-terminal amino acid sequencing, identified 31 distinct polypeptides in this PSII preparation. We propose a new nomenclature for the polypeptide components of PSII identified after PsbZ, which proceeds sequentially from Psb27. During this study, the polypeptides PsbJ, PsbM, PsbX, PsbY, PsbZ, Psb27, and Psb28 proteins were detected for the first time in a purified PSII complex from Synechocystis 6803. Five novel polypeptides were also identified in this preparation. They included the Sll1638 protein, which shares significant sequence similarity to PsbQ, a peripheral protein of PSII that was previously thought to be present only in chloroplasts. This work describes newly identified proteins in a highly purified cyanobacterial PSII preparation that is being widely used to investigate the structure, function, and biogenesis of this photosystem.