BBA - Bioenergetics (v.1847, #12)

Native LH1–RC of photosynthetic purple bacteria Thermochromatium (Tch.) tepidum, B915, has an ultra-red BChl a Qy absorption. Two blue-shifted complexes obtained by chemical modification, B893 and B882, have increasing full widths at half maximum (FWHM) and decreasing transition dipole oscillator strength. 77 K Stark absorption spectroscopy studies were employed for the three complexes, trying to understand the origin of the 915 nm absorption. We found that Tr(∆α) and |∆μ| of both Qy and carotenoid (Car) bands are larger than for other purple bacterial LH complexes reported previously. Moreover, the red shifts of the Qy bands are associated with (1) increasing Tr(∆α) and |∆μ| of the Qy band, (2) the red shift of the Car Stark signal and (3) the increasing |∆μ| of the Car band. Based on the results and the crystal structure, a combined effect of exciton-charge transfer (CT) states mixing, and inhomogeneous narrowing of the BChl a site energy is proposed to be the origin of the 915 nm absorption. CT-exciton state mixing has long been found to be the origin of strong Stark signal in LH1 and special pair, and the more extent of the mixing in Tch. tepidum LH1 is mainly the consequence of the shorter BChl–BChl distances. The less flexible protein structure results in a smaller site energy disorder (inhomogeneous narrowing), which was demonstrated to be able to influence |∆μ| and absorption.
Keywords: LH1–RC; Thermochromatium (Tch.) tepidum; Stark spectroscopy; Red shift; Exciton-charge transfer state mixing; Inhomogeneous narrowing;

Interplay between the hinge region of iron sulphur protein and the Qo site in the bc 1 complex — Analysis of Plasmodium-like mutations in the yeast enzyme by Zehua Song; Jérôme Clain; Bogdan I. Iorga; Cindy Vallières; Anaïs Lalève; Nicholas Fisher; Brigitte Meunier (1487-1494).
The respiratory chain bc 1 complex is central to mitochondrial bioenergetics and the target of antiprotozoals. We characterized a modified yeast bc 1 complex that more closely resemble Plasmodium falciparum enzyme. The mutant version was generated by replacing ten cytochrome b Qo site residues by P. falciparum equivalents. The Plasmodium-like changes caused a major dysfunction of the catalytic mechanism of the bc 1 complex resulting in superoxide overproduction and respiratory growth defect. The defect was corrected by substitution of the conserved residue Y279 by a phenylalanine, or by mutations in or in the vicinity of the hinge domain of the iron–sulphur protein. It thus appears that side-reactions can be prevented by the substitution Y279F or the modification of the iron–sulphur protein hinge region. Interestingly, P. falciparum — and all the apicomplexan — contains an unusual hinge region. We replaced the yeast hinge region by the Plasmodium version and combined it with the Plasmodium-like version of the Qo site. This combination restored the respiratory growth competence. It could be suggested that, in the apicomplexan, the hinge region and the cytochrome b Qo site have co-evolved to maintain catalytic efficiency of the bc 1 complex Qo site.
Keywords: Respiratory complex III; Malaria parasite; Yeast model; Superoxide production;

Continuum electrostatic calculation of the transfer energies of anions from water into aprotic solvents gives the figures erroneous by order of magnitude. This is due to the hydrogen bond disruption that suggests the necessity to reconsider the traditional approach of the purely electrostatic calculation of the transfer energy from water into protein. In this paper, the method combining the experimental estimates of the transfer energies from water into aprotic solvent and the electrostatic calculation of the transfer energies from aprotic solvent into protein is proposed. Hydrogen bonds between aprotic solvent and solute are taken into account by introducing an imaginary aprotic medium incapable to form hydrogen bonds with the solute. Besides, a new treatment of the heterogeneous intraprotein dielectric permittivity based on the microscopic protein structure and electrometric measurements is elaborated. The method accounts semi-quantitatively for the electrostatic effect of diverse charged amino acid substitutions in the donor and acceptor parts of the photosynthetic bacterial reaction center from Rhodobacter sphaeroides. Analysis of the volatile secondary acceptor site QB revealed that in the conformation with a minimal distance between quinone QB and Glu L 212 the proton uptake upon the reduction of QB is prompted by Glu L 212 in alkaline and by Asp L 213 in slightly acidic regions. This agrees with the pH dependences of protonation degrees and the proton uptake. The method of pK calculation was applied successfully also for dissociation of Asp 26 in bacterial thioredoxin.
Keywords: Photosynthesis; Electrostatic calculations; Ion transfer energies; Hydrogen bonds and transfer energies; Dielectric permittivity distribution; Bacterial thioredoxin;

Vibrational relaxation as the driving force for wavelength conversion in the peridinin–chlorophyll a-protein by Jan P. Götze; Bora Karasulu; Mahendra Patil; Walter Thiel (1509-1517).
We present a computationally derived energy transfer model for the peridinin–chlorophyll a-protein (PCP), which invokes vibrational relaxation in the two lowest singlet excited states rather than internal conversion between them. The model allows an understanding of the photoinduced processes without assuming further electronic states or a dependence of the 2Ag state character on the vibrational sub-state. We report molecular dynamics simulations (CHARMM22 force field) and quantum mechanics/molecular mechanics (QM/MM) calculations on PCP. In the latter, the QM region containing a single peridinin (Per) chromophore or a Per-Chl a (chlorophyll a) pair is treated by density functional theory (DFT, CAM-B3LYP) for geometries and by DFT-based multireference configuration interaction (DFT/MRCI) for excitation energies.The calculations show that Per has a bright, green light absorbing 2Ag state, in addition to the blue light absorbing 1Bu state found in other carotenoids. Both states undergo a strong energy lowering upon relaxation, leading to emission in the red, while absorbing in the blue or green. The orientation of their transition dipole moments indicates that both states are capable of excited-state energy transfer to Chl a, without preference for either 1Bu or 2Ag as donor state. We propose that the commonly postulated partial intramolecular charge transfer (ICT) character of a donating Per state can be assigned to the relaxed 1Bu state, which takes on ICT character. By assuming that both 1Bu and 2Ag are able to donate to the Chl a Q band, one can explain why different chlorophyll species in PCP exhibit different acceptor capabilities.Display Omitted
Keywords: Light harvesting; Carotenoids; Excited-state energy transfer; QM/MM; Density functional; Theory;

A new group of eubacterial light-driven retinal-binding proton pumps with an unusual cytoplasmic proton donor by Andrew Harris; Milena Ljumovic; Ana-Nicoleta Bondar; Yohei Shibata; Shota Ito; Keiichi Inoue; Hideki Kandori; Leonid S. Brown (1518-1529).
One of the main functions of microbial rhodopsins is outward-directed light-driven proton transport across the plasma membrane, which can provide sources of energy alternative to respiration and chlorophyll photosynthesis. Proton-pumping rhodopsins are found in Archaea (Halobacteria), multiple groups of Bacteria, numerous fungi, and some microscopic algae. An overwhelming majority of these proton pumps share the common transport mechanism, in which a proton from the retinal Schiff base is first transferred to the primary proton acceptor (normally an Asp) on the extracellular side of retinal. Next, reprotonation of the Schiff base from the cytoplasmic side is mediated by a carboxylic proton donor (Asp or Glu), which is located on helix C and is usually hydrogen-bonded to Thr or Ser on helix B. The only notable exception from this trend was recently found in Exiguobacterium, where the carboxylic proton donor is replaced by Lys. Here we describe a new group of efficient proteobacterial retinal-binding light-driven proton pumps which lack the carboxylic proton donor on helix C (most often replaced by Gly) but possess a unique His residue on helix B. We characterize the group spectroscopically and propose that this histidine forms a proton-donating complex compensating for the loss of the carboxylic proton donor.
Keywords: Eubacterial rhodopsins; Retinal-binding proteins; Proton pumping; Photochemical cycle; FTIR spectroscopy;

In the photosynthetic electron transfer (ET) chain, two electrons transfer from photosystem I to the flavin-dependent ferredoxin-NADP+ reductase (FNR) via two sequential independent ferredoxin (Fd) electron carriers. In some algae and cyanobacteria (as Anabaena), under low iron conditions, flavodoxin (Fld) replaces Fd as single electron carrier. Extensive mutational studies have characterized the protein–protein interaction in FNR/Fd and FNR/Fld complexes. Interestingly, even though Fd and Fld share the interaction site on FNR, individual residues on FNR do not participate to the same extent in the interaction with each of the protein partners, pointing to different electron transfer mechanisms. Despite of extensive mutational studies, only FNR/Fd X-ray structures from Anabaena and maize have been solved; structural data for FNR/Fld remains elusive. Here, we present a multiscale modelling approach including coarse-grained and all-atom protein–protein docking, the QM/MM e-Pathway analysis and electronic coupling calculations, allowing for a molecular and electronic comprehensive analysis of the ET process in both complexes. Our results, consistent with experimental mutational data, reveal the ET in FNR/Fd proceeding through a bridge-mediated mechanism in a dominant protein–protein complex, where transfer of the electron is facilitated by Fd loop-residues 40–49. In FNR/Fld, however, we observe a direct transfer between redox cofactors and less complex specificity than in Fd; more than one orientation in the encounter complex can be efficient in ET.
Keywords: Protein–protein electron transfer; Protein–protein docking; FNR/Fd; FNR/Fld; QM/MM e-Pathway; Electronic coupling;

State transitions are an important photosynthetic short-term response that maintains the excitation balance between photosystems I (PSI) and II (PSII). In plants, when PSII is preferentially excited, LHCII, the main heterotrimeric light harvesting complex of PSII, is phosphorylated by the STN7 kinase, detaches from PSII and moves to PSI to equilibrate the relative absorption of the two photosystems (State II). When PSI is preferentially excited LHCII is dephosphorylated by the PPH1 (TAP38) phosphatase, and returns to PSII (State I). Phosphorylation of LHCII that remain bound to PSII has also been observed. Although the kinetics of LHCII phosphorylation are well known from a qualitative standpoint, the absolute phosphorylation levels of LHCII (and its isoforms) bound to PSI and PSII have been little studied. In this work we thoroughly investigated the phosphorylation level of the Lhcb1 and Lhcb2 isoforms that compose LHCII in PSI-LHCII and PSII-LHCII supercomplexes purified from WT and state transition mutants of Arabidopsis thaliana. We found that, at most, 40% of the monomers that make up PSI-bound LHCII trimers are phosphorylated. Phosphorylation was much lower in PSII-bound LHCII trimers reaching only 15–20%. Dephosphorylation assays using a recombinant PPH1 phosphatase allowed us to investigate the role of the two isoforms during state transitions. Our results strongly suggest that a single phosphorylated Lhcb2 is sufficient for the formation of the PSI-LHCII supercomplex. These results are a step towards a refined model of the state transition phenomenon and a better understanding of the short-term response to changes in light conditions in plants.Display Omitted
Keywords: Photosystems; Lhcb1; Lhcb2; LHCII; State transitions; Phosphorylation;

Interaction of photosystem I from Phaeodactylum tricornutum with plastocyanins as compared with its native cytochrome c 6: Reunion with a lost donor by Pilar Bernal-Bayard; Chiara Pallara; M. Carmen Castell; Fernando P. Molina-Heredia; Juan Fernández-Recio; Manuel Hervás; José A. Navarro (1549-1559).
In the Phaeodactylum tricornutum alga, as in most diatoms, cytochrome c 6 is the only electron donor to photosystem I, and thus they lack plastocyanin as an alternative electron carrier. We have investigated, by using laser-flash absorption spectroscopy, the electron transfer to Phaeodactylum photosystem I from plastocyanins from cyanobacteria, green algae and plants, as compared with its own cytochrome c 6. Diatom photosystem I is able to effectively react with eukaryotic acidic plastocyanins, although with less efficiency than with Phaeodactylum cytochrome c 6. This efficiency, however, increases in some green alga plastocyanin mutants mimicking the electrostatics of the interaction site on the diatom cytochrome. In addition, the structure of the transient electron transfer complex between cytochrome c 6 and photosystem I from Phaeodactylum has been analyzed by computational docking and compared to that of green lineage and mixed systems. Taking together, the results explain why the Phaeodactylum system shows a lower efficiency than the green systems, both in the formation of the properly arranged [cytochrome c 6-photosystem I] complex and in the electron transfer itself.Display Omitted
Keywords: Cytochrome c 6; Electron transfer; Computational docking; Laser flash photolysis; Phaeodactylum; Photosystem I; Plastocyanin;

Partition, orientation and mobility of ubiquinones in a lipid bilayer by Vanesa Viviana Galassi; Guilherme Menegon Arantes (1560-1573).
Ubiquinone is the universal mobile charge carrier involved in biological electron transfer processes. Its redox properties and biological function depend on the molecular partition and lateral diffusion over biological membranes. However, ubiquinone localization and dynamics within lipid bilayers are long debated and still uncertain. Here we present molecular dynamics simulations of several ubiquinone homologs with variable isoprenoid tail lengths complexed to phosphatidylcholine bilayers. Initially, a new force-field parametrization for ubiquinone is derived from and compared to high level quantum chemical data. Free energy profiles for ubiquinone insertion in the lipid bilayer are obtained with the new force-field. The profiles allow for the determination of the equilibrium location of ubiquinone in the membrane as well as for the validation of the simulation model by direct comparison with experimental partition coefficients. A detailed analysis of structural properties and interactions shows that the ubiquinone polar head group is localized at the water–bilayer interface at the same depth of the lipid glycerol groups and oriented normal to the membrane plane. Both the localization and orientation of ubiquinone head groups do not change significantly when increasing the number of isoprenoid units. The isoprenoid tail is extended and packed with the lipid acyl chains. For ubiquinones with long tails, the terminal isoprenoid units have high flexibility. Calculated ubiquinone diffusion coefficients are similar to that found for the phosphatidylcholine lipid. These results may have further implications for the mechanisms of ubiquinone transport and binding to respiratory and photosynthetic protein complexes.Display Omitted
Keywords: Membrane localization; Membrane insertion; Computer simulation; Free-energy profile; Molecular dynamics; Force-field parametrization;

The Carbon Monoxide Dehydrogenase from Desulfovibrio vulgaris by Jessica Hadj-Saïd; Maria-Eirini Pandelia; Christophe Léger; Vincent Fourmond; Sébastien Dementin (1574-1583).
Ni-containing Carbon Monoxide Dehydrogenases (CODHs) catalyze the reversible conversion between CO and CO2 and are involved in energy conservation and carbon fixation. These homodimeric enzymes house two NiFeS active sites (C-clusters) and three accessory [4Fe–4S] clusters. The Desulfovibrio vulgaris (Dv) genome contains a two-gene CODH operon coding for a CODH (cooS) and a maturation protein (cooC) involved in nickel insertion in the active site. According to the literature, the question of the precise function of CooC as a chaperone folding the C-cluster in a form which accommodates free nickel or as a mere nickel donor is not resolved. Here, we report the biochemical and spectroscopic characterization of two recombinant forms of the CODH, produced in the absence and in the presence of CooC, designated CooS and CooSC, respectively. CooS contains no nickel and cannot be activated, supporting the idea that the role of CooC is to fold the C-cluster so that it can bind nickel. As expected, CooSC is Ni-loaded, reversibly converts CO and CO2, displays the typical Cred1 and Cred2 EPR signatures of the C-cluster and activates in the presence of methyl viologen and CO in an autocatalytic process. However, Ni-loaded CooSC reaches maximum activity only upon reductive treatment in the presence of exogenous nickel, a phenomenon that had not been observed before. Surprisingly, the enzyme displays the Cred1 and Cred2 signatures whether it has been activated or not, showing that this activation process of the Ni-loaded Dv CODH is not associated with structural changes at the active site.
Keywords: Ni-containing Carbon Monoxide Dehydrogenase; Protein purification; Enzyme kinetics; Electron paramagnetic resonance (EPR);