Hiroyuki Arata

Intramolecular electron transport in quinoprotein alcohol dehydrogenase of Acetobacter methanolicus: a redox-titration study

Quinohemoprotein-cytochrome c complex alcohol dehydrogenase (ADH) of acetic acid bacteria consists of three subunits, of which subunit I contains pyrroloquinoline quinone (PQQ) and heme c, and subunit II contains three heme c components. The PQQ and heme c components are believed to be involved in the intramolecular electron transfer from ethanol to ubiquinone. To study the intramolecular electron transfer in ADH of Acetobacter methanolicus, the redox potentials of heme c components were determined with ADH complex and the isolated subunits I and II of A. methanolicus, as well as hybrid ADH consisting of the subunit I/III complex of Gluconobacter suboxydans ADH and subunit II of A. methanolicus ADH. The redox potentials of hemes c in ADH complex were -130, 49, 188, and 188 mV at pH 7.0 and 24, 187, 190, and 255 mV at pH 4.5. In hybrid ADH, one of these heme c components was largely changed in the redox potential. Reduced ADH was fully oxidized with potassium ferricyanide, while ubiquinone oxidized the enzyme partially. The results indicate that electrons extracted from ethanol at PQQ site are transferred to ubiquinone via heme c in subunit I and two of the three hemes c in subunit II. Copyright 1998 Elsevier Science B.V.

Restoration of the optimal redox state for the photosynthetic electron transfer system by auxiliary oxidants in an aerobic photosynthetic bacterium, Erythrobacter sp. OCh 114

Abstract In an aerobic photosynthetic bacterium, Erythrobacter sp. OCh 114, photosynthetic electron transfer (photooxidation of cytochromes) and light-driven proton release did not occur under anaerobic conditions. ‘Auxiliary oxidants’, such as nitrate, nitrite, trimethylamine N-oxide (TMAO) and chlorate, restored the optimal redox state for the photosynthetic electron transfer system, resulting in the photooxidation of c-type cytochrome and the release of protons from intact cells on illumination. These auxiliary oxidants oxidized c-type cytochrome(s) under anaerobic conditions in the dark, suggesting the presence of reductases for these auxiliary oxidants. The redox level of the electron transfer chain shifted towards oxidation in the presence of the auxiliary oxidants and was poised to allow photosynthetic electron transfer. KCN inhibited the photooxidation of cytochromes and the light-driven proton release in the presence of these auxiliary oxidants, except for TMAO. KCN also inhibited the auxiliary oxidant-induced oxidation of cytochromes. This suggests that KCN inhibits the reductases for these oxidants. Antimycin and myxothiazol enhanced the auxiliary oxidant-induced oxidation of cytochrome(s), suggesting the involvement of the cytochrome b-c1 complex in nitrate, chlorate and nitrite reduction.

Isolation and characterization of cytochrome b-562 from cytochrome b-c1 comlex of the photosynthetic bacterium Rhodopseudomonas sphaeroides R-26

Cytochrome b‐562 Cytochrome b‐C1 complex Rhodopseudomonas sphaeroides

Free energy change accompanying electron transfer from P870 to quinone in Rhodopseudomonas sphaeroides chromatophores

Abstract Delayed fluorescence of chromatophores of Rhodopseudomonas sphaeroides was measured to estimate the standard free energy change accompanying the electron transfer from the bacteriochlorophyll dimer (P) to the primary acceptor quinone (Q A ). The chromatophores emitted delayed fluorescence with a lifetime of about 60 ms in the presence of o -phenanthroline. By comparing the intensity of the delayed fluorescence with that of the prompt fluorescence, the standard free energy of the P + Q A − radical pair was evaluated. It was about 0.87 eV below the level of excited singlet state, P∗Q A , or 0.51 eV above the ground state, PQ A , independent of pH.

New type of delayed light emission from photosynthetic bacteria

A new type of short-wavelength delayed light emission from the photosynthetic bacteria Rhodospirillum rubrum and Rhodopseudomonas spheroides was found. The emission and action spectra suggest that magnesium protoporphyrin IX or a similar compound is the emitter. Anaerobic conditions were necessary to detect this long-lived emission. Some inhibitors of the primary processes of photosynthesis affected the intensity and the decay rate of the delayed light emission.

Free-energy change accompanying the reduction of the reaction center secondary quinone in Rhodopseudomonas sphaeroides chromatophores

Abstract The standard free-energy change accompanying the electron transfer from QA to QB was estimated from the intensity of the delayed fluorescence in chromatophores of Rhodopseudomonas sphaeroides. The value of 120 meV (at pH 8) suggests that QB− is more stable in the chromatophore membrane than in the isolated reaction center.

Some properties and occurrence of cytochrome c-552 in the aerobic photosynthetic bacterium Roseobacter denitrificans.

Characteristics and occurrence of cytochrome c-552 from an aerobic photosynthetic bacterium, Roseobacter denitrificans, were described.Relative molecular mass of the cytrochrome was 13.5 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and 15,000 by gel filtration. This cytochrome was a acidic protein having a pI of 5.6 and Em was +215 mV at pH 7.0. Absorption peaks were at 278, 408 and 524 nm in the oxidized form and 416, 523 and 552 nm in the reduced form.Amino acid composition and N-terminal amino acid sequence of cytochrome c-552 determined for 24 residues had low similarities to those of cytochrome c-551 of this bacterium, which is homologous to cytochrome c2, although the physico-chemical properties of these two cytochromes were similar to each other.Cytochrome c-552 was maximally synthesized in the light under aerobic conditions but not in the dark. The synthesis also occurred in the presence of alternative acceptors such as trimethylamine N-oxide (TMAO) and nitrate under anaerobic conditions. Our results suggest that cytochrome c-552 is involved in TMAO respiration and denitrification in R. denitrificans, although the effect of light remains to be solved.

Structure of the Membrane-intrinsic Nitric Oxide Reductase from Roseobacter denitrificans.

Membrane-intrinsic nitric oxide reductases (NORs) are key components of bacterial denitrification pathways with a close evolutionary relationship to the cytochrome oxidase (COX) complex found in aerobic respiratory chains. A key distinction between COX and NOR is the identity of the metal directly opposite heme b3 within the active site. In NOR, this metal is iron (FeB), whereas in COX, it is copper (CuB). The purified NOR of Roseobacter denitrificans contains copper and has modest oxidase activity, raising the possibility that a COX-like active site might have independently arisen within the context of a NOR-like protein scaffold. Here we present the crystal structure of the Roseobacter denitrificans NorBC complex and anomalous scattering experiments probing the identity of each metal center. Our results refute the hypothesis that copper occupies the active site and instead reveal a new metal center in the small subunit not seen in any other NOR or COX.

Trimethylamine N Oxide Respiration of Aerobic Photosynthetic Bacterium Erythrobacter sp. OCh 114

Erythrobacter sp. OCh 114 is a photosynthetic bacterium which adapts to aerobic environments (1–4). The amino acid sequence of its cytochrome c 2 (cytochrome c 551) indicated close relation of this species to the photosynthetic and non-photosynthetic species of the α-3 subdivision (5) of the purple bacteria (6).

Flash-Induced Proton Release from Spheroplasts of Rhodopseudomonas Sphaeroides

Vectorial proton translocation coupled with the electron transfer is one of the important subject in the field of the bioenergetics. The advantage of the cyclic electron transfer system of the purple photosyn-thetic bacteria in the study of the coupling mechanism is that we can make rapid stoichiometric excitation using short flashes.