Influence of Hydrocarbon Tail Structure on Quinone Binding and Electron-Transfer Performance at the QA and QB Sites of the Photosynthetic Reaction Center Protein

Warncke, Kurt; Gunner, M. R.; Braun, B.; Gu, Lian‐Quan; Yu, Chao; Bruce, J. Malcolm; Dutton, P. Leslie

doi:10.1021/bi00191a010

Cited by 68 publications

(85 citation statements)

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“…These values are in very good agreement with those previously reported [23], and are shown in Table 2. Their similarity at different detergent concentrations indicates that the thermodynamics of the process are not altered substantially by the detergent.…”

Section: Discussionsupporting

confidence: 93%

Investigation on the detergent role in the function of secondary quinone in bacterial reaction centers

Agostiano¹,

Milano²,

Trotta³

1999

European Journal of Biochemistry

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In this paper are reported studies on the detergent role in isolated reaction centers (RC) from Rhodobacter sphaeroides, over a large range of lauryldimethylamino-N-oxide (LDAO) concentrations, in influencing the thermodynamics of the quinone exchange reaction as well as the protein aggregation. The occurrence of the quinone exchange reaction between the Q B -binding site (where Q B is the second quinone molecule of two in the RC) and the ubiquinone 0 dissolved in the different environments (water, LDAO micelles and detergent phase of the protein± detergent complex) has also been analyzed. Measurements carried out in Q B -depleted RC to which exogenous quinone has been added show that the relative amplitudes of the slow and fast phase of the recombination reaction depend on this parameter. The overall amount of the restored Q B -functionality is affected by the concentration of the LDAO in solution. Interpolation of the titration curves with a quadratic function obtained by simple considerations allowed the binding constant of UQ 0 to the Q B -binding site to be calculated. From the fitting procedure, the distribution of the quinone in the different environments present in solution was evaluated, indicating that the exchange reaction can take place only between the Q B -site and the detergent phase. The dependence of the quinone pool size upon the volume of the phase in which the interacting quinone is solubilized is also discussed. The increasing difficulty in saturating the Q B -pocket above the LDAO critical micellar concentration is finally related to the association of protein±detergent complexes to form large protein clusters.Keywords: bacterial photosynthesis; reaction center; detergents; quinone, electron transfer.The photosynthetic reaction center (RC), an integral transmembrane pigment±protein complex, represents an extremely efficient molecular machine able to perform charge separation across the membrane. The charge separation is impinged by absorption of a photon with a quantum yield close to unity. In purple nonsulfur bacteria, this process is achieved through a series of electron transfer reactions, from a bacteriochlorophyll special pair (D) to a primary ubiquinone acceptor (Q A ). The charge separated state D + Q 2 A is stabilized by replacement of the electron on D + by reduced cytochrome c 2 followed by a forward electron transfer to a secondary ubiquinone molecule (Q B ). The latter quinone is loosely bound to a relatively polar protein domain from which it can freely exchange with a membrane quinone pool.Because of its hydrophobicity, the RC is solubilized and purified by using detergents to disrupt the membrane and keep it in solution forming a protein±detergent complex [ 1]. RCs from Rhodopseudomonas viridis and Rhodobacter sphaeroides have been extensively studied and the three-dimensional structure of the protein±detergent complex is known for both the ordered protein portion [2±4] and for the disordered detergent part [4,6]. The complex has a core formed by the protein surrounded by a r...

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Section: Discussionsupporting

confidence: 93%

Investigation on the detergent role in the function of secondary quinone in bacterial reaction centers

Agostiano¹,

Milano²,

Trotta³

1999

European Journal of Biochemistry

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“…Unlike some branes with wild-type or mutant nitrate reductase A overexpressed, together with menadiol and duroquinol, analogues of more selective reductases and dehydrogenases, therefore, nitrate reductase accepts both menaquinol and ubiquinol as substrates, the physiological quinols menaquinol and ubiquinol. We set out to determine the kinetic behaviour of menadiol and duroquinol though apparently not demethylmenaquinol (Wissenbach et al, 1990(Wissenbach et al, , 1992.…”

Section: Fig 1 Arrangement Of Fe-s Centres In the β Subunit Of Nitrmentioning

confidence: 99%

“…The apparent alignment of the haem groups in the γ subunit none head groups at one of the sites of this centre showed that the complete absence of a tail structure did not decrease the rate (Berks et al, 1995) may allow the electrons to pass from one haem to the other; the distances from edge to edge and from of forward electron transfer by more than 5Ϫ10-fold (Gunner and Dutton, 1989;Warncke et al, 1994). iron to iron obtained by modelling are compatible with electron transfer between the haems.…”

Section: Mutant Forms Of Nitrate Reductase With β Subunits That Lackmentioning

confidence: 99%

Kinetics of Membrane‐Bound Nitrate Reductase A from Escherichia Coli with Analogues of Physiological Electron Donors

Giordani

Buc

Cornish‐Bowden

et al. 1997

European Journal of Biochemistry

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We have compared the steady-state kinetics of wild-type nitrate reductase A and two mutant forms with altered β subunits. To mimic conditions in vivo as closely as possible, we used analogues of the physiological quinols as electron donors and membranes with overexpressed nitrate reductase A in preference to a purified Aβγ complex. With the wild-type enzyme both menadiol and duroquinol supply their electrons for the reduction of nitrate at rates that depend on the square of the quinol concentration, menadiol having the higher catalytic constant. The results as a whole are consistent with a substitutedenzyme mechanism for the reduction of nitrate by the quinols.Kinetic experiments suggest that duroquinol and menadiol deliver their electrons at different sites on nitrate reductase, with cross-inhibition. Menadiol inhibits the duroquinol reaction strongly, suggesting that menaquinol may be the preferred substrate in vivo. To examine whether electron transfer from menadiol and duroquinol for nitrate reduction requires the presence of all of the Fe-S centres, we have studied the steady-state kinetics of mutants with β subunits that lack an Fe-S centre. The loss of the highest-potential Fe-S centre results in an enzyme without menadiol activity, but retaining duroquinol activity; the kinetic parameters are within a factor of two of those of the wild-type enzyme, indicating that this centre is not required for the duroquinol activity. The loss of a low-potential Fe-S centre affects the activity with both quinols: the enzyme is still active but the catalytic constants for both quinols are decreased by about 75%, indicating that this centre is important but not essential for the activity.The existence of a specific site of reaction on nitrate reductase for each quinol, together with the differences in the effects on the two quinols produced by the loss of the Fe-S centre of ϩ80 mV, suggests that the pathways for transfer of electrons from duroquinol and menadiol are not identical.Keywords : nitrate reductase; quinol ; menadiol; duroquinol.A respiratory system constituted by formate dehydrogenase et al., 1989). The A and β subunits are located on the cytoplasmic side of the membrane and form an Aβ complex that binds to the and nitrate reductase is induced in Escherichia coli under anaerobic conditions in the presence of nitrate (Pichinoty, 1969; Ruiz-membrane by interacting with the γ subunit, a very hydrophobic protein embedded in the membrane (Sodergren et al., 1988). Herrera and DeMoss, 1969). It allows the flow of electrons to nitrate from the formate that is produced when pyruvate is conEach subunit carries a different set of redox centres (Blasco verted to acetyl-CoA by pyruvate-formate lyase. The electron et al., 1989 ; Guigliarelli et al., 1992). The 139-kDa A subunit transfer is strongly dependent on the presence of a quinone, and contains the active site for the reduction of nitrate to nitrite and either ubiquinone (a benzoquinone) or menaquinone (a naph-carries a Mo cofactor (molybdopterin guanine dinucleotide)...

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“…11 It has been also reported that two isoprene units of ubiquinone could be modeled in the native structure of complex II by computational analysis using a proteinligand docking program. 12 Napyradiomycin A1 is structurally similar to ubiquinone ( Figure 1a); therefore, it is likely that napyradiomycin A1 binds with the ubiquinone-binding site of complexes I and II through its terpenoid residue.…”

mentioning

confidence: 99%

Napyradiomycin A1, an inhibitor of mitochondrial complexes I and II

et al. 2012

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We have previously proposed a cell-based screening method 'EGFinduced (epidermal growth factor) filopodium protrusion assay' to identify mitochondrial electron transport inhibitors or glycolysis inhibitors. Filopodia are spike-like cell membrane projections that contribute to tumor metastasis. Previously, we have reported that mitochondrial electron transport inhibition resulted in the inhibition of EGF-induced filopodium protrusion in human adenocarcinoma A431 cells only when their glycolytic pathways were restricted. 1 By using the inhibition of filopodium protrusion as an indicator, we identified napyradiomycin A1 (Figure 1a; isolated from Streptomyces antimycoticus NT17), 2 which was previously identified as an antibacterial antibiotic, 3 as a candidate of mitochondrial electron transport inhibitor. A431 cells were treated with napyradiomycin A1 with or without 10 mM 2-deoxy-D-glucose (Sigma-Aldrich, St Louis, MO, USA) for 30 min, followed by 30 ng ml À1 EGF (Sigma-Aldrich) stimulation and observation under a microscope. As shown in Figure 1b, 20 mM napyradiomycin A1 inhibited EGF-induced filopodium protrusion in A431 cells only in the presence of the glycolytic enzyme hexokinase inhibitor 2-deoxy-D-glucose. Mitochondrial electron transport inhibitor rotenone (Sigma-Aldrich) also inhibited filopodium protrusion only in the presence of 2-deoxy-D-glucose. Furthermore, it was reported that co-treatment with a mitochondrial electron transport inhibitor and a glycolytic inhibitor markedly decreased intracellular ATP levels. 1 We then tested whether napyradiomycin A1 decreased ATP levels in A431 cells in which glycolytic pathways were restricted. As intracellular ATP levels were not affected by EGF stimulation (data not shown), we measured intracellular ATP levels under the condition where A431 cells were treated with napyradiomycin A1 with or without 10 mM 2-deoxy-D-glucose for 30 min in the absence of EGF. After incubation, intracellular ATP levels were measured using a Cell Titer-Glo Luminescent Cell Viability Assay Kit (Promega, Madison, WI, USA) with a luminometer (Wallac; Perkin-Elmer, Waltham, MA, USA). As shown in Figure 1c, napyradiomycin A1 treatment did not decrease cellular ATP levels; however, 20 mM napyradiomycin A1 markedly decreased cellular ATP levels in the presence of 2-deoxy-D-glucose in A431 cells. Furthermore, ATP levels in HeLa cells also decreased under co-treatment with napyradiomycin A1 and 2-deoxy-D-glucose (data not shown). These results suggested that napyradiomycin A1 inhibited mitochondrial electron transport in cancer cells.Next, we examined whether napyradiomycin A1 actually inhibited mitochondrial electron transport in vitro by using submitochondrial particles (SMP) obtained from the bovine heart. In order to prepare SMP, bovine hearts were homogenized in MSH buffer (210 mM mannitol, 70 mM sucrose, 1 mM DTT, 1 mM EGTA, 0.1% BSA and 10 mM HEPES pH 7.4) with a Potter-Elvehjem homogenizer (Nippon genetics, Tokyo, Japan). Homogenates were centrifuged at 1000 g for 10 min, and the res...

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Influence of Hydrocarbon Tail Structure on Quinone Binding and Electron-Transfer Performance at the QA and QB Sites of the Photosynthetic Reaction Center Protein

Cited by 68 publications

References 50 publications

Investigation on the detergent role in the function of secondary quinone in bacterial reaction centers

Investigation on the detergent role in the function of secondary quinone in bacterial reaction centers

Kinetics of Membrane‐Bound Nitrate Reductase A from Escherichia Coli with Analogues of Physiological Electron Donors

Napyradiomycin A1, an inhibitor of mitochondrial complexes I and II

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