BBA - Bioenergetics (v.1847, #3)
Editorial Board (i).
Assembly of water-soluble chlorophyll-binding proteins with native hydrophobic chlorophylls in water-in-oil emulsions by Dominika Bednarczyk; Shigekazu Takahashi; Hiroyuki Satoh; Dror Noy (307-313).
The challenges involved in studying cofactor binding and assembly, as well as energy- and electron transfer mechanisms in the large and elaborate transmembrane protein complexes of photosynthesis and respiration have prompted considerable interest in constructing simplified model systems based on their water-soluble protein analogs. Such analogs are also promising templates and building blocks for artificial bioinspired energy conversion systems. Yet, development is limited by the challenge of introducing the essential cofactors of natural proteins that are highly water-insoluble into the water-soluble protein analogs. Here we introduce a new efficient method based on water-in-oil emulsions for overcoming this challenge. We demonstrate the effectiveness of the method in the assembly of native chlorophylls with four recombinant variants of the water-soluble chlorophyll-binding protein of Brassicaceae plants. We use the method to gain new insights into the protein–chlorophyll assembly process, and demonstrate its potential as a fast screening system for developing novel chlorophyll–protein complexes.Display Omitted
Keywords: Water-soluble chlorophyll binding protein (WSCP); Water-in-oil emulsion;
Monte Carlo simulations of excitation and electron transfer in grana membranes by Krzysztof Gibasiewicz; Małgorzata Adamiec; Robert Luciński; Wojciech Giera; Przemysław Chełminiak; Sebastian Szewczyk; Weronika Sipińska; Edyta Głów; Jerzy Karolczak; Rienk van Grondelle; Grzegorz Jackowski (314-327).
Time-resolved fluorescence measurements on grana membranes with instrumental response function of 3 ps reveal faster excitation dynamics (120 ps) than those reported previously. A possible reason for the faster decay may be a relatively low amount of “extra” LHCII trimers per reaction center of Photosystem II. Monte Carlo modeling of excitation dynamics in C2S2M2 form of PSII–LHCII supercomplexes has been performed using a coarse grained model of this complex, constituting a large majority of proteins in grana membranes. The main factor responsible for the fast fluorescence decay reported in this work was the deep trap constituted by the primary charge separated state in the reaction center (950–1090 cm− 1). This value is critical for a good fit, whereas typical hopping times between antenna polypeptides (from ~ 4.5 to ~ 10.5 ps) and reversible primary charge separation times (from ~ 4 to ~ 1.5 ps, respectively) are less critical. Consequently, respective mean migration times of excitation from anywhere in the PSII–LHCII supercomplexes to reaction center range from ~ 30 to ~ 80 ps. Thus 1/4–2/3 of the ~ 120-ps average excitation lifetime is necessary for the diffusion of excitation to reaction center, whereas the remaining time is due to the bottle-neck effect of the trap. Removal of 27% of the Lhcb6 apoprotein pool by mutagenesis of DEG5 gene caused the acceleration of the excitation decay from ~ 120 to ~ 100 ps. This effect may be due to the detachment of LHCII-M trimers from PSII–LHCII supercomplexes, accompanied by deepening of the reaction center trap.
Keywords: Photosystem II; Light Harvesting Complex II; Excitation energy transfer; Electron transfer; Streak camera; Monte Carlo simulation;
Polyethylenimine architecture-dependent metabolic imprints and perturbation of cellular redox homeostasis by Arnaldur Hall; Ladan Parhamifar; Marina Krarup Lange; Kathrine Damm Meyle; May Sanderhoff; Helene Andersen; Martin Roursgaard; Anna Karina Larsen; Per Bo Jensen; Claus Christensen; Jiri Bartek; Seyed Moein Moghimi (328-342).
Polyethylenimines (PEIs) are among the most efficient polycationic non-viral transfectants. PEI architecture and size not only modulate transfection efficiency, but also cytotoxicity. However, the underlying mechanisms of PEI-induced multifaceted cell damage and death are largely unknown. Here, we demonstrate that the central mechanisms of PEI architecture- and size-dependent perturbations of integrated cellular metabolomics involve destabilization of plasma membrane and mitochondrial membranes with consequences on mitochondrial oxidative phosphorylation (OXPHOS), glycolytic flux and redox homeostasis that ultimately modulate cell death. In comparison to linear PEI, the branched architectures induced greater plasma membrane destabilization and were more detrimental to glycolytic activity and OXPHOS capacity as well as being a more potent inhibitor of the cytochrome c oxidase. Accordingly, the branched architectures caused a greater lactate dehydrogenase (LDH) and ATP depletion, activated AMP kinase (AMPK) and disturbed redox homeostasis through diminished availability of nicotinamide adenine dinucleotide phosphate (NADPH), reduced antioxidant capacity of glutathione (GSH) and increased burden of reactive oxygen species (ROS). The differences in metabolic and redox imprints were further reflected in the transfection performance of the polycations, but co-treatment with the GSH precursor N-acetyl-cysteine (NAC) counteracted redox dysregulation and increased the number of viable transfected cells. Integrated biomembrane integrity and metabolomic analysis provides a rapid approach for mechanistic understanding of multifactorial polycation-mediated cytotoxicity, and could form the basis for combinatorial throughput platforms for improved design and selection of safer polymeric vectors.
Keywords: Bioenergetics; Cell death; Glycolytic flux; Mitochondrial dysfunction; Oxidative stress; Polyethylenimine;
Time-resolved visible and infrared difference spectroscopy for the study of photosystem I with different quinones incorporated into the A1 binding site by Hiroki Makita; Nan Zhao; Gary Hastings (343-354).
Room (298 K) and low (77 K) temperature time-resolved visible and infrared difference spectroscopy has been used to study photosystem I particles with phylloquinone (2-methyl-3-phytyl-1,4-naphthoquinone), menadione (2-methyl-1,4-naphthoquinone) and plastoquinone 9 (2,3-dimethyl-5-prenyl-l,4-benzoquinone), incorporated into the A1 binding site. Concentrated samples in short path-length (~ 5 μm) sample cells are typically used in FTIR experiments. Measurements were undertaken using standard “dilute” samples at 298 K, and concentrated (~ 5×) samples at both 298 and 77 K. No concentration induced alterations in the flash-induced absorption changes were observed. Concentrated samples in short path-length cells form a transparent film at 77 K, and could therefore be studied spectroscopically at 77 K without addition of a cryoprotectant. At 298 K, for photosystem I with plastoquinone 9/menadione/phylloquinone incorporated, P700+FA/B − radical pair recombination is characterized by a time constant of 3/14/80 ms, and forward electron transfer from A1A − to Fx by a time constant of 211/3.1/0.309 μs, respectively. At 77 K, for concentrated photosystem I with menadione/phylloquinone incorporated, P700+A1 − radical pair recombination is characterized by a time constant of 240/340 μs, with this process occurring in 58/39% of the PSI particles, respectively. The origin of these differences is discussed. Marcus electron transfer theory in combination with kinetic modeling is used to simulate the observed electron transfer time constants at 298 K. This simulation allows an estimate of the redox potential for the different quinones in the A1 binding site.
Keywords: Photosystem I; Time resolved; A1; Phylloquinone; Electron transfer; FTIR;
Redox changes accompanying inorganic carbon limitation in Synechocystis sp. PCC 6803 by Steven C. Holland; Anthony D. Kappell; Robert L. Burnap (355-363).
Inorganic carbon (Ci) is the major sink for photosynthetic reductant in organisms capable of oxygenic photosynthesis. In the absence of abundant Ci, the cyanobacterium Synechocystis sp. strain PCC6803 expresses a high affinity Ci acquisition system, the CO2-concentrating mechanisms (CCM), controlled by the transcriptional regulator CcmR and the metabolites NADP+ and α-ketoglutarate, which act as co-repressors of CcmR by modulating its DNA binding. The CCM thus responds to internal cellular redox changes during the transition from Ci-replete to Ci-limited conditions. However, the actual changes in the metabolic state of the NADPH/NADP+ system that occur during the transition to Ci-limited conditions remain ill-defined. Analysis of changes in the redox state of cells experiencing Ci limitation reveals systematic changes associated with physiological adjustments and a trend towards the quinone and NADP pools becoming highly reduced. A rapid and persistent increase in F0 was observed in cells reaching the Ci-limited state, as was the induction of photoprotective fluorescence quenching. Systematic changes in the fluorescence induction transients were also observed. As with Chl fluorescence, a transient reduction of the NADPH pool (‘M’ peak), is assigned to State 2→ State 1 transition associated with increased electron flow to NADP+. This was followed by a characteristic decline, which was abolished by Ci limitation or inhibition of the Calvin –Benson–Bassham (CBB) cycle and is thus assigned to the activation of the CBB cycle. The results are consistent with the proposed regulation of the CCM and provide new information on the nature of the Chl and NADPH fluorescence induction curves.
Keywords: Photosynthesis; Carbon concentrating mechanism; NADH-1; NADPH; Fluorescence; Cyanobacteria;
How cytochrome c oxidase can pump four protons per oxygen molecule at high electrochemical gradient by Margareta R.A. Blomberg; Per E.M. Siegbahn (364-376).
Experiments have shown that the A-family cytochrome c oxidases pump four protons per oxygen molecule, also at a high electrochemical gradient. This has been considered a puzzle, since two of the reduction potentials involved, Cu(II) and Fe(III), were estimated from experiments to be too low to afford proton pumping at a high gradient. The present quantum mechanical study (using hybrid density functional theory) suggests a solution to this puzzle. First, the calculations show that the charge compensated Cu(II) potential for Cu B is actually much higher than estimated from experiment, of the same order as the reduction potentials for the tyrosyl radical and the ferryl group, which are also involved in the catalytic cycle. The reason for the discrepancy between theory and experiment is the very large uncertainty in the experimental observations used to estimate the equilibrium potentials, mainly caused by the lack of methods for direct determination of reduced Cu B . Second, the calculations show that a high energy metastable state, labeled E H , is involved during catalytic turnover. The E H state mixes the low reduction potential of Fe(III) in heme a3 with another, higher potential, here suggested to be that of the tyrosyl radical, resulting in enough exergonicity to allow proton pumping at a high gradient. In contrast, the corresponding metastable oxidized state, O H , is not significantly higher in energy than the resting state, O. Finally, to secure the involvement of the high energy E H state it is suggested that only one proton is taken up via the K-channel during catalytic turnover.Display Omitted
Keywords: Cytochrome c oxidase; Proton pumping; Density functional theory; Energy profile; Reduction potential;
Corrigendum to “Fluorescence kinetics of PSII crystals containing Ca2+ or Sr2+ in the oxygen evolving complex” [Biochim. Biophys. Acta Bioenerg. 1837 (2014) 264–269] by Bart van Oort; Joanna Kargul; Karim Maghlaoui; James Barber; Herbert van Amerongen (377).