André Verméglio

Legumes symbioses : Absence of Nod genes in photosynthetic bradyrhizobia

Leguminous plants (such as peas and soybeans) and rhizobial soil bacteria are symbiotic partners that communicate through molecular signaling pathways, resulting in the formation of nodules on legume roots and occasionally stems that house nitrogen-fixing bacteria. Nodule formation has been assumed to be exclusively initiated by the binding of bacterial, host-specific lipochito-oligosaccharidic Nod factors, encoded by the nodABC genes, to kinase-like receptors of the plant. Here we show by complete genome sequencing of two symbiotic, photosynthetic, Bradyrhizobium strains, BTAi1 and ORS278, that canonical nodABC genes and typical lipochito-oligosaccharidic Nod factors are not required for symbiosis in some legumes. Mutational analyses indicated that these unique rhizobia use an alternative pathway to initiate symbioses, where a purine derivative may play a key role in triggering nodule formation.

The Photosynthetic Bacterial Reaction Center II

Alexander Ogrodnik, Thomas Langenbacher, Ulrich Eberl, Martin Yolk and Maria E. Michel-Beyerle Institut fUr Physikalische und Theoretische Chemie Technische Universitiit Miinchen, Lichtenbergstr.4, D-8046 Garching (Germany) Electric field effects on reaction centers from Rb. sphaeroides have been detected in ps transient absorption at 90K. Upon application of a field of 7·IOSY /cm the bleaching of the bacteriopheophytin at the A-branch (HA, 545nm) monitored after 30ps mirrors a reduction of the Quantum yield of P+HA formation by 0:11% (P being the special pair). A similar field induced reduction of the Quantum yield is observed when probing the bleaching of P at 870nm. However, this effect evolves with a time constant of o:900ps. We conclude that the loss channel employs an intermediate state with this lifetime before repopulating the ground state. Field induced formation of P+Hii (B-branch) was excluded from measurements in the Qx transition of HB at 533nm. Measurements in the blue and red wings of the 800nm Qy band of the two bacteriochlorophylls (BA and BB) are compatible with the state P+Bii being the long-lived intermediate involved in the loss of quantum yield.

Supramolecular organization of the photosynthetic apparatus of Rhodobacter sphaeroides.

Native tubular membranes were purified from the purple non‐sulfur bacterium Rhodobacter sphaeroides. These tubular structures contain all the membrane components of the photosynthetic apparatus, in the relative ratio of one cytochrome bc1 complex to two reaction centers, and ∼24 bacteriochlorophyll molecules per reaction center. Electron micrographs of negative‐stained membranes diffract up to 25 Å and allow the calculation of a projection map at 20 Å. The unit cell (a = 198 Å, b = 120 Å and γ = 103°) contains an elongated S‐shaped supercomplex presenting a pseudo‐2‐fold symmetry. Comparison with density maps of isolated reaction center and light‐harvesting complexes allowed interpretation of the projection map. Each supercomplex is composed of light‐harvesting 1 complexes that take the form of two C‐shaped structures of ∼112 Å in external diameter, facing each other on the open side and enclosing the two reaction centers. The remaining positive density is tentatively attributed to one cytochrome bc1 complex. These features shed new light on the association of the reaction center and the light‐harvesting complexes. In particular, the organization of the light‐harvesting complexes in C‐shaped structures ensures an efficient exchange of ubihydroquinone/ubiquinone between the reaction center and the cytochrome bc1 complex.

Bacteriophytochrome controls photosystem synthesis in anoxygenic bacteria

Plants use a set of light sensors to control their growth and development in response to changes in ambient light. In particular, phytochromes exert their regulatory activity by switching between a biologically inactive red-light-absorbing form (Pr) and an active far-red-light absorbing form (Pfr). Recently, biochemical and genetic studies have demonstrated the occurrence of phytochrome-like proteins in photosynthetic and non-photosynthetic bacteria—but little is known about their functions. Here we report the discovery of a bacteriophytochrome located downstream from the photosynthesis gene cluster in a Bradyrhizobium strain symbiont of Aeschynomene. The synthesis of the complete photosynthetic apparatus is totally under the control of this bacteriophytochrome. A similar behaviour is observed for the closely related species Rhodopseudomonas palustris, but not for the more distant anoxygenic photosynthetic bacteria of the genus Rhodobacter, Rubrivivax or Rhodospirillum. Unlike other (bacterio)phytochromes, the carboxy-terminal domain of this bacteriophytochrome contains no histidine kinase features. This suggests a light signalling pathway involving direct protein–protein interaction with no phosphorelay cascade. This specific mechanism of regulation may represent an important ecological adaptation to optimize the plant–bacteria interaction.

Selenite and tellurite reduction by Shewanella oneidensis

ABSTRACT Shewanella oneidensis MR-1 reduces selenite and tellurite preferentially under anaerobic conditions. The Se(0) and Te(0) deposits are located extracellularly and intracellularly, respectively. This difference in localization and the distinct effect of some inhibitors and electron acceptors on these reduction processes are taken as evidence of two independent pathways.

Characterization of the Reduction of Selenate and Tellurite by Nitrate Reductases

ABSTRACT Preliminary studies showed that the periplasmic nitrate reductase (Nap) of Rhodobacter sphaeroides and the membrane-bound nitrate reductases of Escherichia coli are able to reduce selenate and tellurite in vitro with benzyl viologen as an electron donor. In the present study, we found that this is a general feature of denitrifiers. Both the periplasmic and membrane-bound nitrate reductases of Ralstonia eutropha, Paracoccus denitrificans, and Paracoccus pantotrophus can utilize potassium selenate and potassium tellurite as electron acceptors. In order to characterize these reactions, the periplasmic nitrate reductase of R. sphaeroides f. sp. denitrificans IL106 was histidine tagged and purified. The Vmax andKm were determined for nitrate, tellurite, and selenate. For nitrate, values of 39 μmol · min−1 · mg−1 and 0.12 mM were obtained for Vmax and Km, respectively, whereas the Vmax values for tellurite and selenate were 40- and 140-fold lower, respectively. These low activities can explain the observation that depletion of the nitrate reductase in R. sphaeroides does not modify the MIC of tellurite for this organism.

Reduction of Technetium(VII) by Desulfovibrio fructosovorans Is Mediated by the Nickel-Iron Hydrogenase

ABSTRACT Resting cells of the sulfate-reducing bacteriumDesulfovibrio fructosovorans grown in the absence of sulfate had a very high Tc(VII)-reducing activity, which led to the formation of an insoluble black precipitate. The involvement of a periplasmic hydrogenase in Tc(VII) reduction was indicated (i) by the requirement for hydrogen as an electron donor, (ii) by the tolerance of this activity to oxygen, and (iii) by the inhibition of this activity by Cu(II). Moreover, a mutant carrying a deletion in the nickel-iron hydrogenase operon showed a dramatic decrease in the rate of Tc(VII) reduction. The restoration of Tc(VII) reduction by complementation of this mutation with nickel-iron hydrogenase genes demonstrated the specific involvement of the periplasmic nickel-iron hydrogenase in the mechanism in vivo. The Tc(VII)-reducing activity was also observed with cell extracts in the presence of hydrogen. Under these conditions, Tc(VII) was reduced enzymatically to soluble Tc(V) or precipitated to an insoluble black precipitate, depending on the chemical nature of the buffer used. The purified nickel-iron hydrogenase performed Tc(VII) reduction and precipitation at high rates. These series of genetic and biochemical approaches demonstrated that the periplasmic nickel-iron hydrogenase of sulfate-reducing bacteria functions as a Tc(VII) reductase. The role of cytochromec3 in the mechanism is also discussed.

Inhibition of a respiratory activity by short saturating flashes in Chlamydomonas: Evidence for a chlororespiration

Abstract Excitation with short actinic flashes (2 μs) of oxygenated dark-adapted Chlamydomonas cells deposited on a bare O2 platinum electrode induces an increase of the amperometric signal after the first two flashes. Mass spectrometer experiments performed in the presence of 18O2 showed that this signal was not due to the photolysis of water (H216O). The insensitivity of this signal to 10 μM DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea), its stimulation by acetate or high O2 concentration as well as its inhibition by cyanide indicate that these flash-induced changes in O2 concentration were related to the inhibition of a respiratory process. Because this rather fast inhibition of respiration is insensitive to antimycin A and to salicyl hydroxamic acid, inhibitors of mitochondrial respiration, and because it occurs on a single flash illumination, we conclude that the related respiratory activity takes place inside the chloroplast (chlororespiration) and not in the mitochondria. This interpretation is confirmed by the quite high Km(O2) of this process (about 23 μM) compared to those measured for the mitochondrial reactions (0.2 μM for the cytochrome oxidase pathway and 5.5 μM for the alternative pathway). In a mutant lacking Photosystem I activity, no photoinhibition of respiration was observed. We conclude from the above results that the light-induced inhibition of chlororespiration is due to the oxidation by Photosystem I activity of electron carriers common to both photosynthetic and chlororespiratory chains.

Orientation of chromophores in reaction centers of Rhodopseudomonas sphaeroides: A photoselection study

The relative orientation of the pigments of reaction centers from Rhodopseudomonas sphaeroides has been studied by the photoselection technique. A high value (+0.45) of p=(delta AV--delta AH)/(delta AV + delta AH) is obtained when exciting and observing within the 870 nm band which is contradictory to the results of Mar and Gingras (Mar, T. and Gringras, G. (1976) Biochim. Biophys. Acta 440, 609-621) and Shuvalov et al. (Shuvalov, V.A., Asadov, A.A. and Krakhmaleva, I.N. (1977) FEBS Lett. 16, 240-245). It is shown that the low values of p obtained by both groups were erroneous due to excitation conditions. Analysis of the polarization of light-induced changes when exciting with polarized light in single transitions (spheroiden band and bacteriopheophytin Qx bands) enable us to propose a possible arrangement of the pigments within the reaction center. It is concluded that the 870 nm band corresponds to a single transition and is one of the two bands of the primary electron donor (P-870). The second band of the bacteriochlorophyll dimer is centered at 805 nm. The Qx transitions of the molecules constituting the bacteriochlorophyll dimer are nearly parallel (angle less than 25 degrees). The two bacteriopheophytin molecules present slightly different absorption spectra in the near infra-red. Both bacteriopheophytin absorption bands are subject to a small shift under illumination. The angle between the Qy bacteriopheophytin transitions is 55 degrees or 125 degrees. Both Qy transitions are nearly perpendicular to the 870 nm absorption band. Finally, the carotenoid molecules makes an angle greater than 70 degrees with the 870 nm band and the other bacteriochlorophyll molecules.

Menaquinone as pool quinone in a purple bacterium

Purple bacteria have thus far been considered to operate light-driven cyclic electron transfer chains containing ubiquinone (UQ) as liposoluble electron and proton carrier. We show that in the purple γ-proteobacterium Halorhodospira halophila, menaquinone-8 (MK-8) is the dominant quinone component and that it operates in the QB-site of the photosynthetic reaction center (RC). The redox potentials of the photooxidized pigment in the RC and of the Rieske center of the bc1 complex are significantly lower (Em = +270 mV and +110 mV, respectively) than those determined in other purple bacteria but resemble those determined for species containing MK as pool quinone. These results demonstrate that the photosynthetic cycle in H. halophila is based on MK and not on UQ. This finding together with the unusual organization of genes coding for the bc1 complex in H. halophila suggests a specific scenario for the evolutionary transition of bioenergetic chains from the low-potential menaquinones to higher-potential UQ in the proteobacterial phylum, most probably induced by rising levels of dioxygen 2.5 billion years ago. This transition appears to necessarily proceed through bioenergetic ambivalence of the respective organisms, that is, to work both on MK- and on UQ-pools. The establishment of the corresponding low- and high-potential chains was accompanied by duplication and redox optimization of the bc1 complex or at least of its crucial subunit oxidizing quinols from the pool, the Rieske protein. Evolutionary driving forces rationalizing the empirically observed redox tuning of the chain to the quinone pool are discussed.