质体醌库和细胞色素b6f参与调控蓝细菌 Synechocystis sp. PCC 6803的状态转换

用调制叶绿素荧光研究了对苯醌(1,4-benzoquinone,BQ)和二溴百里香醌(2,5-dibromo-3-methyl-6-isopropyl-1,4-benzoquinone,DBMIB)对蓝细菌Synechocystissp.PCC6803状态转换的作用。BQ和DBMIB是质体醌(PQ)的类似物,两者均可充当PQ的电子受体。其中,DBMIB能够和细胞色素b6f的Qo位点特异结合。在没有作用光的情况下,BQ诱导暗适应的蓝细菌进入状态Ⅰ;相反,DBMIB诱导Syne鄄chocystis6803向状态Ⅱ转换。据此提出,在生理状态下蓝细菌根据PQ库的氧化还原状态调节状态转换;细胞色素b6f参与此调控过程。     

Interaction of electron acceptors with thylakoids from halophytic and non-halophytic species

Thylakoid membranes isolated from halophytic species showed differences in their interactions with ionic and lipophilic electron acceptors when compared to thylakoids from non-halophytes. FeCN was considerably less efficient as electron acceptor with halophyte thylakoids, supporting much lower rates of O₂ evolution and having a lower affinity. FeCN accepted electrons at a different, DMMIB insensitive, site with these thylakoids. 1,4-Benzo-quinones with less positive midpoint potentials were less effective in accepting electrons from halophyte thylakoids compared to nonhalophyte thylakoids, also reflected in lower rates of O₂ evolution and lower affinity. Considering the lipolphilic nature and the fact that there was no apparent change in the site donating electrons to the quinones, an alteration in the midpoint potential of this site by about +100mV is postulated for the halophyte thylakoids.Abbreviations AMPD 2-amino-2-methyl-1,3-propanediol - Cyt b₆/f cytochrome b₆/f complex - DBMIB 2,5-dibromo-6-isopropyl-3-methyl-1,4-benzoquinone - DCBQ 2,6-dichloro-1,4-benzoquinone - DCIP 2,6-dichlorophenol-indolphenol - DMBQ 2,5-dimethyl-1,4-benzoquinone - E_m7 midpoint redox potential at pH 7.0, FeCN-K₃Fe(CN)₆ - HNQ 5-hydroxy-1,4-naphthoquinone - MV methylviologen - NQ 1,4-naphthoquinone - PBQ phenyl-1,4-benzoquinone - PC plastocyanin - PQ plastoquinone     

The relationship between state II to state I transitions and cyclic electron flow around photosystem I

The effects of electron acceptors, inhibitors of electron flow and uncouplers and inhibitors of photophosphorylation on a state II to I transition were studied. 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) did not inhibit the state II to I transition. By contrast, 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), methyl viologen and antimycin A inhibited the transition indicating that the cyclic electron flow around photosystem I, but not the oxidation of electron carriers (such as plastoquinone), induced the state II to I transition. Uncouplers, but not inhibitors of photophosphorylation, inhibited the state transition suggesting that the proton transport through the cyclic electron flow was related to the transition.     

The relationship between state II to state I transitions and cyclic electron flow around photosystem I

The effects of electron acceptors, inhibitors of electron flow and uncouplers and inhibitors of photophosphorylation on a state II to I transition were studied. 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) did not inhibit the state II to I transition. By contrast, 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), methyl viologen and antimycin A inhibited the transition indicating that the cyclic electron flow around photosystem I, but not the oxidation of electron carriers (such as plastoquinone), induced the state II to I transition. Uncouplers, but not inhibitors of photophosphorylation, inhibited the state transition suggesting that the proton transport through the cyclic electron flow was related to the transition.     

Flash oxygen yield patterns of autotrophically and photoheterotrophically grown Chlamydobotrys stellata in the presence and absence of lipophilic acceptors

The obligate phototrophic green alga Chlamydobotrys stellata does not evolve oxygen when grown in CO₂-free atmosphere on acetate. With the application of the lipophilic acceptor 2,6-dichloro-p-benzoquinone it was investigated whether this phenomenon is caused by the inactivation of the water-splitting system or by an inhibition of the electron transport chain. It was found that in the presence of DCQ, the photoheterotrophic alga exhibited a normal period-4 flash oxygen pattern, but the steady state yield was only 25% of that measured in the autotrophic cells. After DCQ addition, the initial distribution of S-states and the values of the transition probabilities proved to be the same in the autotrophic and photoheterotrophic algae. These results indicate that photoheterotrophic growth conditions inhibit the electron transport of Chl. stellata behind the acceptor site of DCQ, but the water-splitting system remains active with a reduced oxygen evolving capacity.Abbreviations Chl chlorophyll - DCQ 2,6-dichloro-p-benzoquinone - DCMU 3-(3,4)-dichlorophenyl)-1,1-dimethylurea - DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - pBQ 1,4-benzoquinone - PS I photosystem I - PS II photosystem II     

Three classes of ubiquinone analogs regulate the mitochondrial permeability transition pore through a common site

To identify the structural features required for regulation of the mitochondrial permeability transition pore (PTP) by ubiquinone analogs (Fontaine, E., Ichas, F., and Bernardi, P. (1998) J. Biol. Chem. 40, 25734-25740), we have carried out an analysis with quinone structural variants. We show that three functional classes can be defined: (i) PTP inhibitors (ubiquinone 0, decylubiquinone, ubiquinone 10, 2,3-dimethyl-6-decyl-1,4-benzoquinone, and 2,3,5-trimethyl-6-geranyl-1,4-benzoquinone); (ii) PTP inducers (2,3-dimethoxy-5-methyl-6-(10-hydroxydecyl)-1,4-benzoquinone and 2,5-dihydroxy-6-undecyl-1,4-benzoquinone); and (iii) PTP-inactive quinones that counteract the effects of both inhibitors and inducers (ubiquinone 5 and 2,3,5-trimethyl-6-(3-hydroxyisoamyl)-1,4-benzoquinone) . The structure-function correlation indicates that minor modifications in the isoprenoid side chain can turn an inhibitor into an activator, and that the methoxy groups are not essential for the effects of quinones on the PTP. Since the ubiquinone analogs used in this study have a similar midpoint potential and decrease mitochondrial production of reactive oxygen species to the same extent, these results support the hypothesis that quinones modulate the PTP through a common binding site rather than through oxidation-reduction reactions. Occupancy of this site can modulate the PTP open-closed transitions, possibly through secondary changes of the PTP Ca(2+) binding affinity.     

The origin of the temperature-induced fluorescence fluctuation in <Emphasis Type="Italic">Spirulina platensis</Emphasis>: temperature-sensitive mobility of PQ molecules

Temperature effects on state transitions have been studied in the cyanobacterium Spirulina platensis. At lower temperatures the time to reach completion took longer and the extent of the state transitions was larger. Effects were limited to the temperature range below the phase transition temperature of the membrane lipids. In the presence of the artificial electron acceptor phenyl-1,4-benzoquinone (PBQ) state transitions became completely temperature-independent. State transitions induced by a change in the light climate or in darkness by a switch from aerobic to anaerobic conditions responded similar to temperature; the occurrence of state transitions solely by a change of the temperature has been excluded. Our conclusion is that the temperature-dependent mobility of plastoquinone molecules in the thylakoid membranes is the intrinsic cause of temperature effects on state transitions.     

Kinetic analyses of state transitions of the cyanobacterium Synechococcus sp. PCC 7002 and its mutant strains impaired in electron transport

The state transitions of the cyanobacterium Synechococcus sp. PCC 7002 and of three mutant strains, which were impaired in PsaE-dependent cyclic electron transport (psaE(-)), respiratory electron transport (ndhF(-)) and both activities (psaE(-)ndhF(-)), were analyzed. Dark incubation of the wild type and psaE(-) cells led to a transition to state 2, while the ndhF(-) strains remained in state 1 after dark incubation. The ndhF(-) cells adapted to state 2 when the cells were incubated under anaerobic conditions or in the presence of potassium cyanide; these results suggest that the ndhF(-) cells were inefficient in performing state 1 to state 2 transitions in the dark unless cytochrome oxidase activity was inhibited. In the state 2 to state 1 transition of wild-type cells induced by light in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), there was still a significant reduction of the interphotosystem electron carriers by both respiration and cyclic electron flow around PSI. Kinetic analysis of the state 2 to state 1 transition shows that, in the absence of PSII activity, the relative contribution to the reduced state of the interphotosystem electron carriers by respiratory and cyclic electron transfer is about 72% and 28%, respectively. The state 2 to state 1 transition was prevented by the cytochrome b(6)f inhibitor 2,5-dibromo-3-methyl-6-isopropylbenzoquinone (DBMIB). On the other hand, the state 1 to state 2 transition was induced by DBMIB with half times of approximately 8 s in all strains. The externally added electron acceptor 2,5-dimethyl-benzoquinone (DMBQ) induced a state 2 to state 1 transition in the dark and this transition could be prevented by DBMIB. The light-induced oxidation of P700 showed that approximately 50% of PSI could be excited by 630-nm light absorbed by phycobilisomes (PBS) under state 2 conditions. P700 oxidation measurements with light absorbed by PBS also showed that the dark-induced state 1 to state 2 transition occurred in wild-type cells but not in the ndhF(-) cells. The possible mechanism for sensing an imbalanced light regime in cyanobacterial state transitions is discussed.     

Synthesis and biological activity of derivatives of ubiquinone: photoaffinity analogues containing the 4-azido-2-nitroanilino group

The photoaffinity analogues of ubiquinone 2,3-dimethoxy-5-methyl-6-[2-[1-oxo-3-(4-azido-2-nitroanilino) propoxy]-3-methylbutyl]-1,4-benzoquinone (2'-ANAP-Q-1) and 2,3-dimethoxy-5-methyl-6-[3-[1-oxo-3-(4-azido-2-nitroanilino) propoxy]-3-methylbutyl]-1,4-benzoquinone (3'-ANAP-Q-1) have been synthesized. The required intermediate alcohols 2,3-dimethoxy-5-methyl-6-(2-hydroxy-3-methylbutyl)-1,4-benzoquinone and 2,3-dimethoxy-5-methyl-6-(3-hydroxy-3-methylbutyl)-1,4-benzoquinone were prepared in good yield from ubiquinone 1 by hydration of the side-chain double bond via hydroboration or acid catalysis, respectively. These alcohols were then coupled with 3-(4-azido-2-nitroanilino)propanoic acid, with p-toluenesulfonyl chloride in dry pyridine, to give 2'- and 3'-ANAP-Q-1. The synthetic methods presented should be of general utility in the preparation of derivatives of ubiquinone in which a reactive or reporter group is relatively close to the ubiquinone ring. By use of membrane vesicles prepared from a ubi-men-strain of Escherichia coli described previously [Wallace, B., & Young, I. G. (1977) Biochim. Biophys. Acta 461, 84-100], it has been shown that 2'- and 3'-ANAP-Q-1 substitute for ubiquinone 8 in the NADH, succinate, and D-lactate oxidase systems. Thus, these compounds may be of value in labeling respiratory chain proteins that interact with ubiquinone.     

Light-Induced Phosphate Binding in Relation to Photophosphorylation and Levels of ATP, ADP and AMP in the Green Alga Scenedesmus

Synchronous cultures of Scenedesmus obtusiusculus Chod. were starved for phosphorus for 48 h. Such cells develop an efficient mechanism for phosphate binding which is very sensitive to metabolic inhibitions. Phosphate binding, fluctuations in the ATP pool during dark-light-dark transitions, and steady state levels of ATP, ADP and AMP were studied. The experiments were carried out in a CO₂-free N₂ atmosphere. DCMU, phloridzin and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB) were used as inhibitors of photophosphorylation. Light-induced phosphate uptake was inhibited to various extents by all the inhibitions. The dark-light-dark transition experiments show that neither the light-induced increment in ATP nor the decrease at darkening are affected by DCMU, but DBMIB and phloridzin inhibit both processes. DCMU seems to affect the regulation of the ATP pool size. The steady state levels of the adenylate pools were almost the same in the light as in the dark, and they were also little sensitive to the inhibitors. In unpoisoned cells in the light the steady state ATP/ADP ratio was 1.7 and the energy charge was 0.66. The rates of phosphate binding are not correlated to any of the adenylate parameters studied. This is probably due to the diverse effects of the inhibitors on light-stimulated production of reducing equivalents, photophosphorylation and transfer of energy from the chloroplast to the cytoplasm.     

The effects of 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone on isolated pea chloroplasts

The effects of a quinone analogue, 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone(DBMIB), on the photochemical activities of isolated pea chloroplastshave been investigated. DBMIB completely inhibits pseudo-cyclicflow with methyl viologen (MV) and partially inhibits non-cyclicflow with ferricyanide (FeCN) in both coupled and uncoupledsystems. It is also shown that, under some circumstances, DBMIBacts like an uncoupler in that it stimulates electron flow andinhibits cyclic photophosphorylation. ¹ Present address: 32(1), 6 1/4 Mile, Prome Road, Kamayut P.O., Rangoon, Burma. (Received May 21, 1973; )     

Studies on the mechanism of destabilization of the positive charges trapped in the photosynthetic water-splitting enzyme system Y by a deactivation-accelerating agent

The mechanism of the 2-(3,4,5-trichloro)anilino-3,5 dinitrothiophene (ANT 2S)-induced cyclic electron flow leading to the discharge of the higher-trapped-hole accumulation states S₂ and S₃ in the photosynthetic water-splitting enzyme system Y of chloroplasts has been investigated. It was found:

1. 1. Under normal conditions the ANT 2s-catalyzed cycle includes both light reactions.

2. 2. By selective kinetical inhibition of the electron flow through P700—either by histone treatment or by 2,5-dibromo-3-methyl-6-isopropyl-1,4-benzoquinone blockage—the ANT 2s-induced deactivation of S₂ and S₃ is not significantly changed. Hence, System I activity is not a functional prerequisite for the ANT 2s-catalyzed discharge of S₂ and S₃.

3. 3. The reciprocal half time of the ANT 2s-induced decay of the relative average oxygen yield per flash, as a function of the time t_d between the flashes representing the degree of the Acceleration of the Deactivation Reactions of the water-splitting enzyme system (ADRY) effect, is nearly linearly related to the ANT 2s concentration within the range of 10⁻⁷–10⁻⁶ M.

4. 4. In respect to the mode of action of ANT 2s two different types of mechanism have been discussed: fixed-place mechanism and mobile-catalyst mechanism.

5. 5. Based on the experimental data the conclusion has been drawn that the ADRY agent ANT 2s probably acts as a mobile catalyst.

Isolation and identification of secondary metabolites of a strain ofAspergillus terreus,a common food contaminant

2-Hydroxy 3-methyl 1,4-benzoquinone 5,6 epoxide was identified as secondary metabolite of a strain ofAspergillus terreus, a common contaminant of animal feeds. In addition, the following compounds were also tentatively identified to be produced by this organism: 2-hYdroxy 3-methyl 1,4-benzoquinone; 2-methyl 1,4-benzoquinone 5,6-epoxide; naphthazarin epoxide; and 2-hydroxy 3-methyl 1,4-benzoquinone 5, 6-epoxide.     

State 1-State 2 transitions in the cyanobacterium Synechococcus 6301 are controlled by the redox state of electron carriers between Photosystems I and II

The mechanism by which state 1-state 2 transitions in the cyanobacterium Synechococcus 6301 are controlled was investigated by examining the effects of a variety of chemical and illumination treatments which modify the redox state of the plastoquinone pool. The extent to which these treatments modify excitation energy distribution was determined by 77K fluorescence emission spectroscopy. It was found that treatment which lead to the oxidation of the plastoquinone pool induce a shift towards state 1 whereas treatments which lead to the reduction of the plastoquinone pool induce a shift towards state 2. We therefore propose that state transitions in cyanobacteria are triggered by changes in the redox state of plastoquinone or a closely associated electron carrier. Alternative proposals have included control by the extent of cyclic electron transport around PS I and control by localised electrochemical gradients around PS I and PS II. Neither of these proposals is consistent with the results reported here.Abbreviations DBMIB 2,5-dibromo-3methyl-6-isopropyl-p-benzoquinone - Chl chlorophyll - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DQH₂ duroquinol (tetramethyl-p-hydroquinone) - LHC II light-harvesting chlorophyll a/b-binding protein of PS II - Light 1 light predominantly exciting PS I - Light 2 light predominantly exciting PS II - M.V. methyl viologen - PS photosystem     

Characterization and Genetic Analysis of Mutant Strains of Escherichia coli K-12 Accumulating the Ubiquinone Precursors 2-Octaprenyl-6-Methoxy-1,4-Benzoquinone and 2-Octaprenyl-3-Methyl-6-Methoxy-1,4-Benzoquinone

The ubiquinone precursors, 2-octaprenyl-6-methoxy-1,4-benzoquinone and 2-octaprenyl-3-methyl-6-methoxy-1,4-benzoquinone, were isolated from ubiquinone-deficient mutants of Escherichia coli and identified by nuclear magnetic resonance and mass spectrometry. Mutants accumulating 2-octaprenyl-6-methoxy-1,4-benzoquinone and 2-octaprenyl-3-methyl-6-methoxy-1,4-benzoquinone were shown to carry mutations in genes designated ubiE and ubiF, respectively. The ubiE gene was shown to be cotransducible with metE (minute 75) and close to two other genes concerned with ubiquinone biosynthesis. The ubiF gene was located close to minute 16 by cotransduction with the lip, gltA, and entA genes.     

Evidence for alpha-Tocopherol Function in the Electron Transport Chain of Chloroplasts

The effect of three different stable radicals-2,2-diphenyl-1-picrylhydrazyl, 1,3,5-triphenyl-verdazyl, and galvinoxyl-was studied in photosystem II of spinach (Spinacia oleracea) chloroplasts. Inhibition by the three was noted on dimethylbenzoquinone reduction in presence of 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB) and on silicomolybdate reduction in presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) in photosystem II and on the H₂O → methylviologen reaction encompassing both photosystems. Inhibition of all photosystem II reactions except silicomolybdate reduction could be partially restored by α-tocopherol or by 9-ethoxy-α-tocopherone but not by other quinones or radical chasers. On this basis, a functional role for α-tocopherol in the electron transport chain of spinach chloroplasts between the DCMU and DBMIB inhibition sites is postulated.     

A comparison study on ribonuclease A modifications induced by substituted p-benzoquinones

In this paper, we present our investigation on ribonuclease A (RNase) modifications induced by 1,4-benzoquinone (PBQ), 2-methyl-1,4-benzoquinone (MBQ), and 2-chloro-1,4-benzoquinone (CBQ). The goal of the study was to evaluate quinone-induced protein modifications as well as substituent effects, utilizing several techniques such as SDS–PAGE, fluorescence spectroscopy, microscopy, and LC-ESI⁺-QTOF-MS. SDS–PAGE experiments revealed that all quinones modify RNase through oligomerization as well as polymeric aggregation; with CBQ functioning as the most efficient quinone while MBQ was least efficient. The fluorescence emission was found to be less intense and the anisotropy values were found to be slightly higher for the modified RNase compared to the unmodified RNase. UV–Vis spectroscopy indicated that all three quinones formed adducts in which they were covalently linked to RNase. Confocal imaging analysis showed that the presence of CBQ resulted in massive RNase aggregation, while PBQ-treated RNase formed much smaller aggregates. MBQ-treated RNase exhibited micrographic features that closely resembled those of the unmodified RNase. LC-ESI⁺-QTOF-MS studies indicated the nature of PBQ- and CBQ-induced RNase modifications are complex mainly due to simultaneously occurrence of both adduct formation and oligomerization. Kinetic studies on quinone reactivity toward lysine revealed the rank order of CBQ > PBQ ≫ MBQ, based on the second-order rate constants. We also utilized scanning electron microscopy in order to investigate the effect of modified RNase on the biomineralization of salts.     

Development of a new E. coli strain to detect oxidative mutation and its application to the fungicide o-phenylphenol and its metabolites

Oxidative mutation is mainly induced by reactive oxygen species (ROS), such as the superoxide anion radical (O(2)(-)) and hydrogen peroxide (H(2)O(2)). However, in Escherichia coli (E. coli), ROS are eliminated by enzymes such as superoxide dismutase and catalase, which are coded by sodAB and katEG genes. In this study, to detect mutagens that induce oxidative mutation, a mutant (WP2katEGsodAB) with katEG and sodAB deleted was constructed by gene manipulation of E. coli WP2. H(2)O(2) and menadione sodium bisulfite generated mutation in WP2katEGsodAB but not in WP2. o-Phenylphenol (OPP) and its metabolites (phenylhydroquinone (PHQ) and phenyl-1,4-benzoquinone (PBQ)), which had been shown to be negative in the Ames test but reported to be carcinogenic, induced mutation in WP2katEGsodAB but not in WP2. These results suggest that the new assay may be useful for the detection of oxidative mutagens.     

Benzoquinones from Costus speciosus seeds

In addition to α-tocopherolquinone and 5α-stigmast-9(11)-en-3β-ol, two new quinones have been isolated from the seeds of Costus speciosus and characterized as 6-methyl dihydrophytylplastoquinone (2,5,6-trimethyl-3-(3,7,11,15-tetramethylhexadecyl)-1,4-benzoquinone) and dihydrophytylplastoquinone (5,6-dimethyl-3-(3,7,11,15-tetramethylhexadecyl)-1,4-benzoquinone) respectively by physical data and chemical studies.     

La photooxydation du cytochrome b-559, en presence de carbonylcyanure-p-trifluorométhoxyphenylhydrazone et de 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone,ou de p-benzoquinone,chez trois mutants non-photosynthétiques de Chlamydomonas reinhardti

Cytochrome b-559 photooxidation in the presence of carbonyl cyanide p-trifluorometh-oxyphenylhydrazone and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone or p-benzoquinone in three non-photosynthetic mutants of Chlamydomonas reinhardtiStudies of absorbance changes related to the cytochrome b-559 photooxidation induced by FCCP, with and without addition of 3-p-chlorophenyl-1, 1-dimethylurea (CMU), DBMIB or p-benzoquinone, in whole cells and in chloroplast fragments of Chlamydomonas reinhardti, were carried out. In addition to the wild type, three strains of non-photosynthetic mutants were used: Fl 5, which lacks P 700; Fl 9 and Fl 15, which are deficient in bound cytochrome c-553 and in cytochrome b-563.In the presence of FCCP, whole cells and chloroplast fragments of the four strains showed a System II-dependent photooxidation of cytochrome b-559. This photooxidation was inhibited by CMU but it occurred again in presence of FCCP, CMU and DBMIB. In chloroplast fragments, cytochrome b-559 photooxidation was also inhibited by an excess of FCCP; it was recovered, likewise, by addition of DBMIB. In whole cells, the highest measured redox changes were: 1 μmol oxidized cytochrome b-559 per 1 mmol chlorophyll, corresponding approximately to about one seventh (wild type, Fl 5) or one fifth (Fl 9, Fl 15) of the total amount of this cytochrome.Another kind of cytochrome b-559 photooxidation, CMU-insensitive, also occurred in the mutants Fl 9 and Fl 15 and in the wild type, but not in the mutant Fl 5. This latter kind of photooxidation was observed with chloroplast fragments in the presence of FCCP and CMU and also with whole cells in the presence of FCCP, CMU and p-benzoquinone. These reactions can be attributed to the Photosystem I; they do not require the intervention of the cytochrome c-553.A high-potential form of cytochrome b-559, hydroquinone-reducible, was involved in these two kinds of photooxidation. In addition, a lower potential form, reducible only by ascorbate, appeared to be able to interfere also.An interpretation is attempted, taking into consideration the various effects of FCCP and DBMIB, at different concentrations, on photosynthetic electron transport.