Immobilization of photosystem I or II complexes on electrodes for preparation of photoenergy-conversion devices

Photosystems, PSI and PSII isolated from Thermosynechococcus elongatus were successfully immobilized on a TiO₂ nanostructured film for use in dye-sensitized biosolar cells (DSBCs). The photosystem complexes were also immobilized on an ITO electrode modified with 3-aminopropyltriethoxysilane by utilizing the interactions between the electrode and the surface of the PSI or PSII polypeptide. Illumination of the PSI and PSII complexes immobilized on the ITO electrode resulted in action spectra in the presence of methyl viologen, which corresponded to the absorption spectra of the complexes. Compared with the ITO electrode, PSI or PSII complexes assembled on the TiO₂ electrode had much higher energy-conversion efficiency in the presence of an iodide/triiodide redox system of an ionic-liquid-based electrolyte. This could have interesting applications in the development of DSBCs.     

Effect of Ce<Superscript>3+</Superscript> on spectral characteristic of D1/D2/Cytb559 complex from spinach

It was studied by spectroscopy that PSII reaction center complex consisting of three polypeptides, D₁, D₂ and Cytb559, were purified from PSII particle of CeCl₃ treated spinach. The results of the experiment show that Ce³⁺ could improve the growth of spinach, and accelerate electron transport of PSII particles. Of chl-a of UV-Vis spectrum of D1/D2/Cytb559 complex, Soret band was blue-shifted by 3 nm and Q band by 2 nm, respectively, and the fluorescence emission peak was blue-shifted by 5 nm in CeCl₃-treated spinach compared with the one in control. By the extended X-ray absorption fine structure (EXAFS) spectroscopy methods, it has been found that Ce³⁺ is coordinated with 8 nitrogen atoms in the first coordination shell with Ce-N bond length of 0.253 nm, and Ce³⁺ with 6 oxygen atoms in the second coordination shell with Ce-O bond length of 0.32 nm. However, the secondary structure of D1/D2/Cytb559 complex by circular dichroism (CD) spectroscopy has no significant change after CeCl₃ treated. It might be that Ce³⁺ binds to porphyrin rings of chlorophyll and oxygen of amino acid residue of polypeptide in D1/D2/Cytb559 complex, and then accelerates the primary reaction of PSII, intensifies function of P680⁺ primary electron donor of D1/D2/Cytb559, but there is little change in conformation of PSII reaction center complex.     

LaCl3提高菠菜光系统Ⅱ活性的作用机制

The effect of LaCl3 on the K3Fe(CN)6 (FeCy) reduction rate and the oxygen-evolving rate of PSU particles of spinach, and the spectral characterization of the D1/D2/Cytb559 of a PSII reaction center complex consisting of three polypeptides from spinach were studied. The experimental results showed that LaCl3 could significantly accelerate the transformation from light energy to electric energy, the electron transport, water photolysis and oxygen evolution of PSII of spinach, which was related to the spectral characterization of the D1/D2/Cytb559 complex.Soret band and Q band of Chl-a of UV-vis spectrum of D1/D2/Cytb559 complex were blue shifted, and the fluorescence emission peak was blue shifted in LaCl3 treated spinach compared with that in the control. The EXAFS (extended X-ray absorption fine structure spectroscopy) revealed that La^3 was coordinated with 8 nitrogen or oxygen atoms in the first coordination shell with La-N or La-O bond length of 0.254 nm, and with 6 nitrogen or oxygen atoms in the second coordination shell with La-N or La-O bond length of 0.321 nm in the D1/D2/Cytb559 complex. The CD suggested that the secondary structure of D1/D2/Cytb559 complex have been litfie affected by the treatment of LaCl3.     

Function and association of CyanoP in photosystem II of Synechocystis sp. PCC 6803

The CyanoP protein is a cyanobacterial homolog of the PsbP protein, which is an extrinsic subunit of photosystem II (PSII) in green plant species. The molecular function of CyanoP has been investigated in mutant strains of Synechocystis but inconsistent results have been reported by different laboratories. In this study, we generated and characterized a Synechocystis mutant in which entire region of the CyanoP gene was eliminated. After repeated subculture in CaCl₂-depleted medium, growth retardation was clearly observed for a CyanoP knockout mutant of Synechocystis sp. PCC 6803 (?P). The PSII-mediated oxygen-evolving activity of the ?P cells was more susceptible to depletion of CaCl₂ than that of wild-type cells. The 77 K fluorescence emission spectra indicated that energy coupling between phycobilisome and PSII was perturbed in both wild-type and ?P cells under CaCl₂-depleted conditions, and was more evident for the ?P mutant. To examine the association of CyanoP with PSII complexes, we tested several detergents for solubilization of thylakoid membranes and showed that CyanoP was partly included in fractions containing large protein complexes in gel-filtration analysis. These results indicate that CyanoP constitutively stabilizes PSII functionality in vivo.     

A Blue Light-inducible Phosphodiesterase Activity in the Cyanobacterium Synechococcus elongatus

A blue light-inducible phosphodiesterase (PDE) activity, specific for the hydrolysis of cyclic di-GMP (c-di-GMP), has been identified in a recombinant protein from Synechococcus elongatus. Blue light (BL) activation is accomplished by a light, oxygen, voltage (LOV) domain, found in plant phototropins and bacterial BL photoreceptors. The genome of S. elongatus contains two genes coding for proteins with LOV domains fused to EAL domains (SL1 and SL2). In both cases, a GGDEF motif is placed in between the LOV and the EAL motifs. Such arrangement is frequently found with diguanylate-cyclase (DGC) functions that form c-di-GMP. Cyclic di-GMP acts as a second messenger molecule regulating biofilm formation in many microbial species. Both enzyme activities modulate the intracellular level of this second messenger, although in most proteins only one of the two enzyme functions is active. Both S. elongatus LOV-GGDEF-EAL proteins were expressed in full length or as truncated proteins. Only the SL2 protein, expressed as a LOV-GGDEF-EAL construct, showed an increase of PDE activity upon BL irradiation, demonstrating this activity for the first time in a LOV-domain protein. Addition of GTP or c-di-GMP did not affect the observed enzymatic activity. In none of the full-length or truncated proteins was a DGC activity detected.     

Binding Model and 3D-QSAR of 3-（2-Chloropyrid-5- ylmethylamino）-2-cyanoacrylates as PSⅡ Electron Transport Inhibitor

The binding model of 3-（2-chloropyrid-5-ylmethylamino）-2-cyanoacrylate photosystem Ⅱ （PSⅡ） electron transport inhibitors with the D 1 protein of PSII was built. The high herbicidal activity of this kind of inhibitors was explained by docking studies： in addition to usual factors, the N atom on the pyridine ring could form an H-bond with the backbone amide of Phe265 on the D1 protein. 3D-QSAR analysis on sixteen 3-（2-chloropyrid-5-yl- methylamino）-2-cyanoacrylate compounds was performed using CoMFA method to explain the nature of interactions between the compounds and D1 protein. These studies may provide useful insights for designing new PSII electron transport inhibitors.     

Force field development for cofactors in the photosystem II

We present a set of force field (FF) parameters compatible with the AMBER03 FF to describe five cofactors in photosystem II (PSII) of oxygenic photosynthetic organisms: plastoquinone‐9 (three redox forms), chlorophyll‐a, pheophytin‐a, heme‐b, and β‐carotene. The development of a reliable FF for these cofactors is an essential step for performing molecular dynamics simulations of PSII. Such simulations are important for the calculation of absorption spectrum and the further investigation of the electron and energy transfer processes. We have derived parameters for partial charges, bonds, angles, and dihedral‐angles from solid theoretical models using systematic quantum mechanics (QM) calculations. We have shown that the developed FF parameters are in good agreement with both ab initio QM and experimental structural data in small molecule crystals as well as protein complexes. © 2012 Wiley Periodicals, Inc.     

Chemical studies of marine invertebrates—XLV: The chemistry of three norsesterterpene peroxides from the sponge Sigmosceptrella laevis

The structures of sigmosceptrellin-A, -B and -C, three norsesterterpenes constituting the ichthyotoxic fraction of the sponge Sigmosceptrella laevis are represented by formulae 1a, 2a and 3a. 1a, 2a and 3a.     

Molecular dynamics characterization of the SAMHD1 Aicardi–Goutières Arg145Gln mutant: structural determinants for the impaired tetramerization

Aicardi–Goutières syndrome, a rare genetic disorder characterized by calcification of basal ganglia, results in psychomotor delays and epilepsy states from the early months of children life. This disease is caused by mutations in seven different genes encoding proteins implicated in the metabolism of nucleic acids, including SAMHD1. Twenty SAMHD1 gene variants have been discovered and in this work, a structural characterization of the SAMHD1 Aicardi–Goutières Arg145Gln mutant is reported by classical molecular dynamics simulation. Four simulations have been carried out and compared. Two concerning the wild-type SAMHD1 form in presence and absence of cofactors, in order to explain the role of cofactors in the SAMHD1 assembly/disassembly process and, two concerning the Arg145Gln mutant, also in presence and absence of cofactors, in order to have an accurate comparison with the corresponding native forms. Results show the importance of native residue Arg145 in maintaining the tetramer, interacting with GTP cofactor inside allosteric sites. Replacement of arginine in glutamine gives rise to a loosening of GTP–protein interactions, when cofactors are present in allosteric sites, whilst in absence of cofactors, the occurrence of intra and inter-chain interactions is observed in the mutant, not seen in the native enzyme, making energetically unfavourable the tetramerization process.     

Identification and in silico molecular modelling study of newly isolated Bacillus subtilis SI-18 strain against S9 protein of Rhizoctonia solani

The numerous bioactive components from Bacillus subtilis are commonly used as antimicrobial agents for reducing plant diseases caused by fungal pathovars. In this study, we isolated and identified B. subtilis SI-18 strain from twenty isolates of rhizosphere soil through morphological and molecular approaches, and explored its inhibitory activities against Rhizoctonia solani. According to morphological features and 16S rRNA and gyrB gene sequence analysis, B. subtilis SI-18 strain was identified. Additionally, the culture filtrate of B. subtilis SI-18 resulted in the suppression of R. solani mycelium growth and material leakage from the cells. Then, we have performed homology modelling and molecular docking study of S9 protein from R. solani where three potential compounds (D1, D2, and D3) were identified among 134 antimicrobial compounds derived from B. subtilis group based on higher binding energy and interaction at the active grove of the target protein. The D1 compound creates alkyl bond at Val48 whereas D2 also binds with Val48 by creating hydrogen bond. On the other hand, two hydrogen bonds were observed at Val48 and Ile52 by D3, which might be responsible for possible blocking of the target S9 protein of R. solani. To validate the docking study and understand the change in drug-ligand conformation, molecular dynamics simulation was assessed where rigid conformation was found for D1, D2 and D3 complexes. Moreover, ADMET study confirms that no toxicity and carcinogenicity were found for screened compounds. Based on our studies, we demonstrated that compounds D1, D2, and D3 derived from B. subtilis can be a potential inhibitor of S9 protein of R. solani that might be a possible strategy for fungal disease prevention.     

Assessing the Applicability of Singlet Oxygen Photosensitizers in Leaf Studies

Singlet oxygen (¹O₂) is of special interest in plant stress physiology. Studies focused on internal, chlorophyll‐mediated production are often complemented with the use of artificial ¹O₂ photosensitizers. Here, we report a comparative study on the effects of Rose Bengal (RB), Methylene Violet (MVI), Neutral Red (NR) and Indigo Carmine (IC). These were infiltrated into tobacco leaves at concentrations generating the same fluxes of ¹O₂ in solution. Following green light‐induced ¹O₂ production from these dyes, leaf photosynthesis was characterized by Photosystem (PS) II and PSI electron transport and oxidative damage was monitored as degradation of D1, a PSII core protein. Cellular localizations were identified on the basis of the dyes’ fluorescence using confocal laser scanning microscopy. We found that RB and NR were both localized in chloroplasts but the latter had very little effect, probably due to its pH‐dependent photosensitizing. Both RB and intracellular, nonplastid MVI decreased PSII electron transport, but the effect of RB was stronger than that of MVI and only RB was capable of damaging the D1 protein. Intercellularly localized IC had no significant effect. Our results also suggest caution when using RB as photosensitizer because it affects PSII electron transport.     

Protein dynamics studies on a wheat type 2 lipid transfer protein

Plant non specific lipid transfer proteins form a superfamily of related proteins composed of type 1 and type 2 LTPs. Type 2 LTPs have been less extensively studied than type 1 and differ in term of primary sequence, molecular mass and transfer efficiency. We have undertaken NMR studies of the wheat 7 kDa type 2 LTP. Whilst the 3D structure determination is in progress, we have studied the protein dynamics. Two zones have been defined within the protein. One remains unperturbed, the LTP being liganded or not; the other one exists in different states, depending on the ligandation, and on the kind of lipid interacting with the protein. S² determination showed a rigid backbone. Multifield analysis allowed the calculation of the chemical exchange rate for each residue.     

Distribution of the cationic state over the chlorophyll pair of the photosystem II reaction center

The reaction center chlorophylls a (Chla) of photosystem II (PSII) are composed of six Chla molecules including the special pair Chla P(D1)/P(D2) harbored by the D1/D2 heterodimer. They serve as the ultimate electron abstractors for water oxidation in the oxygen-evolving Mn(4)CaO(5) cluster. Using the PSII crystal structure analyzed at 1.9 ? resolution, the redox potentials of P(D1)/P(D2) for one-electron oxidation (E(m)) were calculated by considering all PSII subunits and the protonation pattern of all titratable residues. The E(m)(Chla) values were calculated to be 1015-1132 mV for P(D1) and 1141-1201 mV for P(D2), depending on the protonation state of the Mn(4)CaO(5) cluster. The results showed that E(m)(P(D1)) was lower than E(m)(P(D2)), favoring localization of the charge of the cationic state more on P(D1). The P(D1)(?+)/P(D2)(?+) charge ratio determined by the large-scale QM/MM calculations with the explicit PSII protein environment yielded a P(D1)(?+)/P(D2)(?+) ratio of ~80/~20, which was found to be due to the asymmetry in electrostatic characters of several conserved D1/D2 residue pairs that cause the E(m)(P(D1))/E(m)(P(D2)) difference, e.g., D1-Asn181/D2-Arg180, D1-Asn298/D2-Arg294, D1-Asp61/D2-His61, D1-Glu189/D2-Phe188, and D1-Asp170/D2-Phe169. The larger P(D1)(?+) population than P(D2)(?+) appears to be an inevitable fate of the intact PSII that possesses water oxidation activity.     

Durability of oxygen evolution of photosystem II incorporated into lipid bilayers

Photosystem II (PSII) has attracted a lot of attention for use in the construction of artificial photosynthetic materials due to its high activity of oxidation of water molecules. However, the robustness of PSII needs to be improved for in vitro application. In this study, we incorporated PSII (Thermosynechococcus vulcanus) into various phospholipid membranes to examine the activity and durability of oxygen evolution. PSII was incorporated into anionic 1,2-dioleoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (PSII-DOPG), zwitterionic 1,2-dioleoyl-sn-glycero-3-phosphocholine (PSII-DOPC), and cationic 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine (PSII-EDOPC). Structural integrity of PSII was examined by absorption and fluorescence spectroscopy. Compared with PSII dissolved in a micellar solution of n-dodecyl-β-d-maltoside (PSII-micelle), durability of PSII-DOPC and PSII-DOPG were enhanced by 1.3- and 1.5-fold, respectively. The activity and durability of PSII-EDOPC was significantly low. Lipid-dependent activity and durability were discussed in terms of kinetic parameters of V _max and K _m, and inhibition of the electron acceptor, phenyl-p-benzoquinone.     

A synthetic DNA and fusion PCR approach to the ectopic expression of high levels of the D1 protein of photosystem II in Synechocystis sp. PCC 6803

A hybrid approach involving synthetic DNA, fusion PCR, and ectopic expression has been used to genetically manipulate the expression of the D1 protein of photosystem II (PSII) in the model cyanobacterium Synechocystis sp. PCC6803. Due to the toxicity of the full-length psbA gene in E. coli, a chimeric psbA2 gene locus was commercially synthesised and cloned in two halves. High-fidelity fusion PCR utilizing sequence overlap between the two synthetic gene halves allowed the production of a DNA fragment that was able to recombine the full-length psbA2 gene into the Synechocystis chromosome at an ectopic (non-native) location. This was accomplished by designing the synthetic DNA/fusion PCR product to have the psbA2 gene, with control sequences, interposed between chimeric sequences corresponding to an ectopic target chromosomal location. Additionally, a recipient strain of Synechocystis lacking all three psbA genes was produced by a combination of traditional marker replacement and markerless replacement techniques. Transformation of this multiple deletion strain by the synthetic DNA/fusion PCR product faithfully restored D1 expression in terms of its expression and PSII repair capacity. The advantages and potential issues for using this approach to rapidly introduce chimeric sequence characteristics as a general tool to produce novel genetic constructs are discussed.     

Effect of Ce~(3+) on spectral characteristic of D1/D2/Cytb559 complex from spinach

In our early researches, lanthanum and cerium could enter plant and bind to porphyrin of chlorophyll to form Ln3+-chllorophyll. La and Ce greatly increase photosystem II (PSII) activity and PSII electron transport rate, and the fluorescence emission peaks of PSII are blue-shifted [1—4]. Do REEs coordinate with PSII reaction center complex in vivo? Moreover, do REEs coordinate with D1(30 kD)/D2(32 kD)/Cytb559 (~9 kD) reaction center complex of site of producing pri-mary reaction-p…     

Biocatalytic asymmetric reduction of 3-acetylisoxazoles

The reduction of the carbonyl group in 3-acetylisoxazole derivatives by algae (Cyanobacterium: Synechococcus elongatus PCC 7942 and Synechosystis sp. PCC 6803) and plant cells (Caragana chamlagu) gave the corresponding (S)-alcohols with high enantioselectivities.     

The exchange of the fast substrate water in the S2 state of photosystem II is limited by diffusion of bulk water through channels – implications for the water oxidation mechanism

The molecular oxygen we breathe is produced from water-derived oxygen species bound to the Mn₄CaO₅ cluster in photosystem II (PSII). Present research points to the central oxo-bridge O5 as the ‘slow exchanging substrate water (W_s)’, while, in the S₂ state, the terminal water ligands W2 and W3 are both discussed as the ‘fast exchanging substrate water (W_f)’. A critical point for the assignment of W_f is whether or not its exchange with bulk water is limited by barriers in the channels leading to the Mn₄CaO₅ cluster. In this study, we measured the rates of H₂¹⁶O/H₂¹⁸O substrate water exchange in the S₂ and S₃ states of PSII core complexes from wild-type (WT) Synechocystis sp. PCC 6803, and from two mutants, D1-D61A and D1-E189Q, that are expected to alter water access via the Cl1/O4 channels and the O1 channel, respectively. We found that the exchange rates of W_f and W_s were unaffected by the E189Q mutation (O1 channel), but strongly perturbed by the D61A mutation (Cl1/O4 channel). It is concluded that all channels have restrictions limiting the isotopic equilibration of the inner water pool near the Mn₄CaO₅ cluster, and that D61 participates in one such barrier. In the D61A mutant this barrier is lowered so that W_f exchange occurs more rapidly. This finding removes the main argument against Ca-bound W3 as fast substrate water in the S₂ state, namely the indifference of the rate of W_f exchange towards Ca/Sr substitution.

Access to the oxygen-evolving complex in photosynthesis is restricted by specific barriers in the channels connecting the Mn₄CaO₅ catalyst with bulk water. Together with other recent data, this finding allows assigning the two substrate waters.