BBA - Bioenergetics (v.1817, #3)
Editorial Board (i).
Plastoquinol is more active than α-tocopherol in singlet oxygen scavenging during high light stress of Chlamydomonas reinhardtii by Beatrycze Nowicka; Jerzy Kruk (389-394).
In the present study, we have performed comparative analysis of different prenyllipids in Chlamydomonas reinhardtii cultures during high light stress under variety of conditions (presence of inhibitors, an uncoupler, heavy water). The obtained results indicate that plastoquinol is more active than α-tocopherol in scavenging of singlet oxygen generated in photosystem II. Besides plastoquinol, also its oxidized form, plastoquinone shows antioxidant action during the stress conditions, resulting in formation of plastoquinone-C, whose level can be regarded as an indicator of singlet oxygen oxidative stress in vivo. The pronounced stimulation of α-tocopherol consumption and α-tocopherolquinone formation by an uncoupler, FCCP, together with the results of additional model system studies, led to the suggestion that α-tocopherol can be recycled in thylakoid membranes under high light conditions from 8a-hydroperoxy-α-tocopherone, the primary oxidation product of α-tocopherol by singlet oxygen.► Plastoquinol is more active than α-tocopherol in scavenging of singlet oxygen. ► Plastoquinone also shows antioxidant action during high-light stress. ► Oxidation of plastoquinone by singlet oxygen results in formation of plastoquinone-C. ► Uncoupling of proton gradient results in enhanced α-tocopherolquinone production.
Keywords: Chlamydomonas reinhardtii; Oxidative stress; Plastoquinone; Plastoquinol; Singlet oxygen; α-tocopherol;
Isolation and pigment composition of the reaction centers from purple photosynthetic bacterium Rhodopseudomonas palustris species by Tadashi Mizoguchi; Megumi Isaji; Jiro Harada; Hitoshi Tamiaki (395-400).
The reaction centers (RCs) from several species of a purple photosynthetic bacterium, Rhodopseudomonas palustris, were first isolated by ammonium-sulfate fractionation of the isolated core complexes, and were successfully purified by anion-exchange and gel-filtration chromatography as well as sucrose-density gradient centrifugation. The RCs were characterized by spectroscopic and biochemical analyses, indicating that they were sufficiently pure and had conserved their redox activity. The pigment composition of the purified RCs was carefully analyzed by LCMS. Significant accumulation of both bacteriochlorophyll(BChl)-a and bacteriopheophytin(BPhe)-a esterified with various isoprenoid alcohols in the 17-propionate groups was shown in RCs for the first time. Moreover, a drastic decrease in BPhe-a with the most dehydrogenated and rigid geranylgeranyl(GG) ester was observed, indicating that BPhe-a in RC preferably took partially hydrogenated and flexible ester groups, i.e. dihydro-GG and tetrahydro-GG in addition to phytyl. Based on the reported X-ray crystal structures of purple bacterial RCs, the meaning of flexibility of the ester groups in BChl-a and BPhe-a as the cofactors of RCs is proposed.► Reaction centers (RCs) from Rhodopseudomonas palustris species were first isolated. ► Their bacteriochlorophyll-a & bacteriopheophytin-a had various 17-propionate esters. ► The bacteriopheophytin-a takes preferably more hydrogenated geranylgeranyl groups. ► The flexibility of the 17-propionates is discussed using crystal structures of RCs.
Keywords: Reaction center; Rhodopseudomonas palustris; Bacteriochlorophyll-a; Bacteriopheophytin-a; 17-Propionate;
Silencing of nicotinamide nucleotide transhydrogenase impairs cellular redox homeostasis and energy metabolism in PC12 cells by Fei Yin; Harsh Sancheti; Enrique Cadenas (401-409).
Mitochondrial NADPH generation is largely dependent on the inner-membrane nicotinamide nucleotide transhydrogenase (NNT), which catalyzes the reduction of NADP+ to NADPH utilizing the proton gradient as the driving force and NADH as the electron donor. Small interfering RNA (siRNA) silencing of NNT in PC12 cells results in decreased cellular NADPH levels, altered redox status of the cell in terms of decreased GSH/GSSG ratios and increased H2O2 levels, thus leading to an increased redox potential (a more oxidized redox state). NNT knockdown results in a decrease of oxidative phosphorylation while anaerobic glycolysis levels remain unchanged. Decreased oxidative phosphorylation was associated with a) inhibition of mitochondrial pyruvate dehydrogenase (PDH) and succinyl-CoA:3-oxoacid CoA transferase (SCOT) activity; b) reduction of NADH availability, c) decline of mitochondrial membrane potential, and d) decrease of ATP levels. Moreover, the alteration of redox status actually precedes the impairment of mitochondrial bioenergetics. A possible mechanism could be that the activation of the redox-sensitive c-Jun N-terminal kinase (JNK) and its translocation to the mitochondrion leads to the inhibition of PDH (upon phosphorylation) and induction of intrinsic apoptosis, resulting in decreased cell viability. This study supports the notion that oxidized cellular redox state and decline in cellular bioenergetics – as a consequence of NNT knockdown – cannot be viewed as independent events, but rather as an interdependent relationship coordinated by the mitochondrial energy-redox axis. Disruption of electron flux from fuel substrates to redox components due to NNT suppression induces not only mitochondrial dysfunction but also cellular disorders through redox-sensitive signaling.► NNT plays a critical role in maintaining cellular redox status. ► NNT silencing impairs mitochondrial bioenergetics. ► Inter-dependence of mitochondrial energy metabolism and redox status. ► Regulation of redox signaling and apoptosis by mitochondrial H2O2.
Keywords: Nicotinamide nucleotide transhydrogenase; Mitochondrion; Bioenergetics; Redox status; JNK; Apoptosis;
Brown adipose tissue mitochondria oxidizing fatty acids generate high levels of reactive oxygen species irrespective of the uncoupling protein-1 activity state by Peter Schönfeld; Lech Wojtczak (410-418).
Mitochondria from brown adipose tissue (BATM) have a high enzymatic capacity for fatty acid oxidation and therefore are an ideal model to examine the sites of reactive oxygen species (ROS) generation during fatty acid oxidation. ROS generation by BATM (isolated from 3-week-old rats) was measured during acylcarnitine oxidation as release of H2O2 into the medium and as inactivation of the matrix enzyme aconitase. The following results were obtained: (1) BATM release large amounts of H2O2 in the coupled as well as in the uncoupled states, several times more than skeletal muscle mitochondria. (2) H2O2 release is especially large with acylcarnitines of medium-chain fatty acids (e.g. octanoylcarnitine). (3) Reverse electron transport does not contribute in a significant extent to the overall ROS generation. (4) Despite the large release of H2O2, the ROS-sensitive matrix enzyme aconitase is not inactivated during acylcarnitine oxidation. (5) In contrast to acylcarnitines, oxidation of α-glycerophosphate by BATM is characterized by large H2O2 release and a pronounced aconitase inactivation. We hypothesize that acylcarnitine-supported ROS generation in BATM may be mainly associated with acyl-CoA dehydrogenase and electron transferring flavoprotein-ubiquinone reductase rather than with complexes of the respiratory chain.► BAT mitochondria oxidizing acylcarnitine esters generate high amounts of ROS. ► These ROS are predominantly released at the external face of the inner membrane. ► Reverse electron transport does not contribute significantly to this process. ► It may be associated with β-oxidation enzymes rather than with electron transfer chain.
Keywords: Brown adipose tissue; Mitochondria; Fatty acid oxidation; Reactive oxygen species (ROS); Reverse electron transfer;
Damage to mitochondrial complex I during cardiac ischemia reperfusion injury is reduced indirectly by anti-anginal drug ranolazine by Ashish K. Gadicherla; David F. Stowe; William E. Antholine; Meiying Yang; Amadou K.S. Camara (419-429).
Ranolazine, an anti-anginal drug, is a late Na+ channel current blocker that is also believed to attenuate fatty acid oxidation and mitochondrial respiratory complex I activity, especially during ischemia. In this study, we investigated if ranolazine's protective effect against cardiac ischemia/reperfusion (IR) injury is mediated at the mitochondrial level and specifically if respiratory complex I (NADH Ubiquinone oxidoreductase) function is protected. We treated isolated and perfused guinea pig hearts with ranolazine just before 30 min ischemia and then isolated cardiac mitochondria at the end of 30 min ischemia and/or 30 min ischemia followed by 10 min reperfusion. We utilized spectrophotometric and histochemical techniques to assay complex I activity, Western blot analysis for complex I subunit NDUFA9, electron paramagnetic resonance for activity of complex I Fe–S clusters, enzyme linked immuno sorbent assay (ELISA) for determination of protein acetylation, native gel histochemical staining for respiratory supercomplex assemblies, and high pressure liquid chromatography for cardiolipin integrity; cardiac function was measured during IR. Ranolazine treated hearts showed higher complex I activity and greater detectable complex I protein levels compared to untreated IR hearts. Ranolazine treatment also led to more normalized electron transfer via Fe–S centers, supercomplex assembly and cardiolipin integrity. These improvements in complex I structure and function with ranolazine were associated with improved cardiac function after IR. However, these protective effects of ranolazine are not mediated by a direct action on mitochondria, but rather indirectly via cytosolic mechanisms that lead to less oxidation and better structural integrity of complex I.► Mitochondrial complex I is a major target of cardiac ischemia/reperfusion (IR) injury. ► IR injury caused specific biophysical, biochemical and molecular changes in complex I. ► A cardio-protective drug, ranolazine, was found to indirectly reduce complex I damage. ► Cardiac function after IR injury can be improved by indirectly reducing complex I dysfunction.
Keywords: Complex I; Mitochondrion; IR injury; Ranolazine; EPR; Heart;
High-valent [MnFe] and [FeFe] cofactors in ribonucleotide reductases by Nils Leidel; Ana Popović-Bijelić; Kajsa G.V. Havelius; Petko Chernev; Nina Voevodskaya; Astrid Gräslund; Michael Haumann (430-444).
Ribonucleotide reductases (RNRs) are essential for DNA synthesis in most organisms. In class-Ic RNR from Chlamydia trachomatis (Ct), a MnFe cofactor in subunit R2 forms the site required for enzyme activity, instead of an FeFe cofactor plus a redox-active tyrosine in class-Ia RNRs, for example in mouse (Mus musculus, Mm). For R2 proteins from Ct and Mm, either grown in the presence of, or reconstituted with Mn and Fe ions, structural and electronic properties of higher valence MnFe and FeFe sites were determined by X-ray absorption spectroscopy and complementary techniques, in combination with bond-valence-sum and density functional theory calculations. At least ten different cofactor species could be tentatively distinguished. In Ct R2, two different Mn(IV)Fe(III) site configurations were assigned either L4MnIV(μO)2FeIIIL4 (metal–metal distance of ~ 2.75 Å, L = ligand) prevailing in metal-grown R2, or L4MnIV(μO)(μOH)FeIIIL4 (~ 2.90 Å) dominating in metal-reconstituted R2. Specific spectroscopic features were attributed to an Fe(IV)Fe(III) site (~ 2.55 Å) with a L4FeIV(μO)2FeIIIL3 core structure. Several Mn,Fe(III)Fe(III) (~ 2.9–3.1 Å) and Mn,Fe(III)Fe(II) species (~ 3.3–3.4 Å) likely showed 5-coordinated Mn(III) or Fe(III). Rapid X-ray photoreduction of iron and shorter metal–metal distances in the high-valent states suggested radiation-induced modifications in most crystal structures of R2. The actual configuration of the MnFe and FeFe cofactors seems to depend on assembly sequences, bound metal type, valence state, and previous catalytic activity involving subunit R1. In Ct R2, the protonation of a bridging oxide in the MnIV(μO)(μOH)FeIII core may be important for preventing premature site reduction and initiation of the radical chemistry in R1.Display Omitted► We studied high-valent MnFe and FeFe sites in ribonucleotide reductase from Chlamydia. ► More than 8 cofactor configurations were identified using, e.g., X-ray spectroscopy. ► The active site properties depend on assembly, metal type and valence, and catalysis. ► Two Mn(IV)Fe(III) species seem to differ in the protonation state of bridging oxides. ► Protonation may determine cofactor redox potentials and radical chemistry initiation.
Keywords: Ribonucleotide reductase; Chlamydia; MnFe cofactor; Redox intermediate; X-ray absorption spectroscopy;
Red antenna states of Photosystem I trimers from Arthrospira platensis revealed by single-molecule spectroscopy by Marc Brecht; Martin Hussels; Eberhard Schlodder; Navassard V. Karapetyan (445-452).
Single-molecule fluorescence spectroscopy at 1.4 K was used to investigate the spectral properties of red (long-wavelength) chlorophylls in trimeric Photosystem I (PSI) complexes from the cyanobacterium Arthrospira platensis. Three distinct red antenna states could be identified in the fluorescence spectra of single PSI trimers from A. platensis in the presence of oxidized P700. Two of them are responsible for broad emission bands centered at 726 and 760 nm. These bands are similar to those found in bulk fluorescence spectra measured at cryogenic temperatures. The broad fluorescence bands at ≅ 726 and ≅ 760 nm belong to individual emitters that are broadened by strong electron–phonon coupling giving rise to a large Stokes-shift of about 20 nm and rapid spectral diffusion. An almost perpendicular orientation of the transition dipole moments of F726 and F760 has to be assumed because direct excitation energy transfer does not occur between F726 and F760. For the first time a third red state assigned to the pool absorbing around 708 nm could be detected by its zero-phonon lines. The center of the zero-phonon line distribution is found at ≅ 714 nm. The spectral properties of the three red antenna states show a high similarity to the red antenna states found in trimeric PSI of Thermosynechoccocus elongatus. Based on these findings a similar organization of the red antenna states in PSI of these two cyanobacteria is discussed.► Single-molecule spectroscopy on the red Chls of PSI of A. platensis ► Fluorescence spectra are composed of sharp ZPLs and broad intensity distributions ► ZPLs distributed around 714 nm can be assigned to red states belonging to C708. ► Strong electron–phonon coupling and rapid spectral diffusion give rise to broad bands ► Red Chls of A. platensis/T. elongatus rely on Chl aggregates with similar properties
Keywords: Photosystem I; Single-molecule spectroscopy; Long-wavelength antenna chlorophyll; Energy transfer; Arthrospira platensis;
Enhanced charge-independent mitochondrial free Ca2 + and attenuated ADP-induced NADH oxidation by isoflurane: Implications for cardioprotection by Bhawana Agarwal; Amadou K.S. Camara; David F. Stowe; Zeljko J. Bosnjak; Ranjan K. Dash (453-465).
Modulation of mitochondrial free Ca2 + ([Ca2 +]m) is implicated as one of the possible upstream factors that initiates anesthetic-mediated cardioprotection against ischemia–reperfusion (IR) injury. To unravel possible mechanisms by which volatile anesthetics modulate [Ca2 +]m and mitochondrial bioenergetics, with implications for cardioprotection, experiments were conducted to spectrofluorometrically measure concentration-dependent effects of isoflurane (0.5, 1, 1.5, 2 mM) on the magnitudes and time-courses of [Ca2 +]m and mitochondrial redox state (NADH), membrane potential (ΔΨm), respiration, and matrix volume. Isolated mitochondria from rat hearts were energized with 10 mM Na+- or K+-pyruvate/malate (NaPM or KPM) or Na+-succinate (NaSuc) followed by additions of isoflurane, 0.5 mM CaCl2 (≈ 200 nM free Ca2 + with 1 mM EGTA buffer), and 250 μM ADP. Isoflurane stepwise: (a) increased [Ca2 +]m in state 2 with NaPM, but not with KPM substrate, despite an isoflurane-induced slight fall in ΔΨm and a mild matrix expansion, and (b) decreased NADH oxidation, respiration, ΔΨm, and matrix volume in state 3, while prolonging the duration of state 3 NADH oxidation, respiration, ΔΨm, and matrix contraction with PM substrates. These findings suggest that isoflurane's effects are mediated in part at the mitochondrial level: (1) to enhance the net rate of state 2 Ca2 + uptake by inhibiting the Na+/Ca2 + exchanger (NCE), independent of changes in ΔΨm and matrix volume, and (2) to decrease the rates of state 3 electron transfer and ADP phosphorylation by inhibiting complex I. These direct effects of isoflurane to increase [Ca2 +]m, while depressing NCE activity and oxidative phosphorylation, could underlie the mechanisms by which isoflurane provides cardioprotection against IR injury at the mitochondrial level.► We examined mechanisms of isoflurane on modulating mitochondrial bioenergetics and [Ca2 +]m. ► Isoflurane increased state 2 [Ca2 +]m independent of changes in ΔΨm and matrix volume. ► Increased [Ca2 +]m is due to inhibition of mitochondrial Na+/Ca2 + exchanger (NCE). ► Isoflurane decreased NADH oxidation, respiration, ΔΨm, and matrix volume in state 3. ► Isoflurane delayed state 3 duration of bioenergetic variables by complex I inhibition.
Keywords: Cardiac IR injury; Anesthetic; Cardioprotection; Mitochondrion; Ca2 + uptake/efflux; Bioenergetics;