Aryl-homoserine lactone quorum sensing in stem-nodulating photosynthetic bradyrhizobia.

Many Proteobacteria possess LuxI-LuxR-type quorum-sensing systems that produce and detect fatty acyl-homoserine lactone (HSL) signals. The photoheterotroph Rhodopseudomonas palustris is unusual in that it produces and detects an aryl-HSL, p-coumaroyl-HSL, and signal production requires an exogenous source of p-coumarate. A photosynthetic stem-nodulating member of the genus Bradyrhizobium produces a small molecule signal that elicits an R. palustris quorum-sensing response. Here, we show that this signal is cinnamoyl-HSL and that cinnamoyl-HSL is produced by the LuxI homolog BraI and detected by BraR. Cinnamoyl-HSL reaches concentrations on the order of 50 nM in cultures of stem-nodulating bradyrhizobia grown in the presence or absence of cinnamate. Acyl-HSLs often reach concentrations of 0.1-30 µM in bacterial cultures, and generally, LuxR-type receptors respond to signals in a concentration range from 5 to a few hundred nanomolar. Our stem-nodulating Bradyrhizobium strain responds to picomolar concentrations of cinnamoyl-HSL and thus, produces cinnamoyl-HSL in excess of the levels required for a signal response without an exogenous source of cinnamate. The ability of Bradyrhizobium to produce and respond to cinnamoyl-HSL shows that aryl-HSL production is not unique to R. palustris, that the aromatic acid substrate for aryl-HSL synthesis does not have to be supplied exogenously, and that some acyl-HSL quorum-sensing systems may function at very low signal production and response levels.

Time-Resolved Study of Interprotein Signaling Process of a Blue Light Sensor PapB-PapA Complex.

PapB is a short BLUF protein from the purple bacterium Rhodopseudomonas palustris, and it has the function of regulating the phosphodiesterase activity of its partner protein PapA. Because PapA is an enzymatic protein containing an EAL domain, which hydrolyzes a cyclic-dimeric-GMP, the PapB-PapA complex regulates, for example, biofilm formation in a light-dependent manner. The reaction dynamics of PapB, as well as dynamics of signal transduction in the PapB-PapA complex, were studied mostly by time-resolved transient grating and CD methods. It was found that although PapB exists as a monomer in both dark and light states, the diffusion coefficient slightly increases and partial molar volume changes upon light irradiation with time constants of 80 µs, 3.2 ms, and 24 ms. The last conformational change leads to a signaling state that is attributed to the α-helix unfolding in the C-terminal region of the BLUF domain. Interestingly, when PapA is added to a PapB solution to form a PapB-PapA complex, the diffusion coefficient decreases significantly, with a time constant of 26 ms, which is similar to the rate of the last conformational change of PapB. It was found that this change is due to the conformational change of PapA, and the light signal from PapB is transmitted through the conformational change of PapB. Compared with another BLUF protein, BlrP1, which possesses the EAL effector domain and requires photoexcitation of two monomer units of the dimer for a major conformational change, the conformational change of the PapB-PapA complex is induced by excitation of one monomer PapB among the heterotetramer.

Phototrophic growth of Rubrivivax gelatinosus in poultry slaughterhouse wastewater.

Rubrivivax gelatinosus was grown in Pfennig's synthetic medium (PM) and in treated wastewater from poultry slaughterhouse (TW) to assess growth profiles for biomass production. Cultures inoculated at 1% (v/v) were grown under anaerobiosis at 30+/-2 degrees C and 1400+/-200 lux for 12 days. Regular absorbance curves for R. gelatinosus were found both on PM and TW. On PM, the highest dry weight of biomass, 0.39 gL(-1), was achieved in the 216-h culture and the highest specific growth rate of 0.2960 h(-1) occurred in the 24-h culture. On TW, the highest biomass of 0.57 gL(-1) was also obtained in the 216-h culture and the highest specific growth rate, 0.1970 h(-1), was achieved in the 48-h culture. For productivity and chemical oxygen demand investigations, the cultivation was accomplished in the TW under anaerobiosis at 32+/-2 degrees C and 4000+/-500 lux, for 10 days. Productivity was 0.085 g biomass (d.w.) L(-1) day(-1), with a COD decrease of 91%.

Spatially-resolved fluorescence-detected two-dimensional electronic spectroscopy probes varying excitonic structure in photosynthetic bacteria.

Conventional implementations of two-dimensional electronic spectroscopy typically spatially average over ~1010 chromophores spread over ~104 micron square area, limiting their ability to characterize spatially heterogeneous samples. Here we present a variation of two-dimensional electronic spectroscopy that is capable of mapping spatially varying differences in excitonic structure, with sensitivity orders of magnitude better than conventional spatially-averaged electronic spectroscopies. The approach performs fluorescence-detection-based fully collinear two-dimensional electronic spectroscopy in a microscope, combining femtosecond time-resolution, sub-micron spatial resolution, and the sensitivity of fluorescence detection. We demonstrate the approach on a mixture of photosynthetic bacteria that are known to exhibit variations in electronic structure with growth conditions. Spatial variations in the constitution of mixed bacterial colonies manifests as spatially varying peak intensities in the measured two-dimensional contour maps, which exhibit distinct diagonal and cross-peaks that reflect differences in the excitonic structure of the bacterial proteins.

Structural determination of dihydro- and tetrahydrogeranylgeranyl groups at the 17-propionate of bacteriochlorophylls-a.

In the final stage of bacteriochlorophyll (BChl) biosynthesis, the presence of BChl-a molecules possessing dihydrogeranylgeranyl and tetrahydrogeranylgeranyl groups at the 17-propionate has been reported. However, the molecular structures of such BChls-a have not yet been determined in terms of the positions of CC double bonds in the 17(2)-ester. In this study, we isolated significant amounts of such pure BChls-a from Rhodopseudomonas palustris and determined their structures by both mass spectrometry and (1)H and (13)C NMR spectroscopy. The determined structures enable us to discuss a stepwise reduction from a geranylgeranyl to phytyl substituent.

Characterization of a separate small domain derived from the 5' end of 23S rRNA of an alpha-proteobacterium.

We demonstrate the presence of a separate processed domain derived from the 5' end of 23S rRNA in ribosomes of Rhodopseudomonas palustris, a member of the alpha-++proteobacteria. Previous sequencing studies predicted intervening sequences (IVS) at homologous positions within the 23S rRNA genes of several alpha-proteobacteria, including R.palustris, and we find a processed 23S rRNA 5' domain in unfractionated RNA from several species. 5.8S rRNA from eukaryotic cytoplasmic large subunit ribosomes and the bacterial processed 23S rRNA 5' domain share homology, possess similar structures and are both derived by processing of large precursors. However, the internal transcribed spacer regions or IVSs separating them from the main large subunit rRNAs are evolutionarily unrelated. Consistent with the difference in sequence, we find that the site and mechanism of IVS processing also differs. Rhodopseudomonas palustris IVS-containing RNA precursors are cleaved in vitro by Escherichia coli RNase III or a similar activity present in R.palustris extracts at a processing site distinct from that found in eukaryotic systems and this results in only partial processing of the IVS. Surprisingly, in a reaction unlike characterized cases of eubacterial IVS processing, an RNA segment larger than the corresponding DNA insertion is removed which contains conserved sequences. These sequences, by analogy, serve to link the 23S rRNA 5' rRNA domains or 5.8S rRNAs to the main portion of other prokaryotic 23S rRNAs or to eukaryotic 28S rRNAs, respectively.

Direct measurement of pure absorbance spectra of living phototrophic microorganisms.

The pure absorbance of turbid cell suspensions of various phototrophic microorganisms were determined by collecting the scattered light. A conventional spectrophotometer was used, equipped with an intergrating sphere as receiver unit, which allowed precise measurements of the absorbance in the range from zero to 0.1. In the wavelength range 300--1100 nm, where photosynthesis occurs, light scattered only once by a bacterial cell retains predominantly the forward direction. This allows measurements of pure absorption, when the concentration of cells which the light has to pass through is small. For example, by comparison of measurements of pigmented and nonpigmented cell suspensions of Rhodopseudomonas acidophila, it was shown that the total sum of scattered light can be collected. The best results were obtained using cuvettes with a light path of 0.1 cm or 0.2 cm to measure cell suspensions of about 0.2 mg dry weight per ml. For R. acidophila this corresponds to 1--3 cell layers. Extinction-, absorbance- and scattering spectra for R. acidophila are presented, in addition to the absorbance spectra for Rhodospirillum rubrum, Aphanocapsa and Scenedesmus.

Detergent structure in crystals of the integral membrane light-harvesting complex LH2 from Rhodopseudomonas acidophila strain 10050.

Integral membrane proteins are solubilized by their incorporation into a detergent micelle. The detergent micelle has a critical influence on the formation of a three-dimensional crystal lattice. The bulk detergent phase is not seen in X-ray crystal structures of integral membrane proteins, due to its disordered character. Here, we describe the detergent structure present in crystals of the peripheral light-harvesting complex of the purple bacteria Rhodopseudomonas acidophila strain 10050 at a maximal resolution of 12A as determined by neutron crystallography. The LH2 molecule has a toroidal shape and spans the membrane completely in vivo. A volume of 16% of the unit cell could be ascribed to detergent tails, localized on both the inner and outer hydrophobic surfaces of the molecule. The detergent tail volumes were found to be associated with individual LH2 molecules and had no direct role in the formation of the crystalline lattice.

[Isolation and identification of a photosynthetic bacteria producing coenzyme Q10].

13 isolates producing Coenzyme Q10 (CoQ10) of purple non sulfur photosynthetic bacteria were enriched out of pond sludge, one isolate named 2c was selected based on its high CoQ10 content and identified systematically. The gram-negative and short-rod shaped strain 2c is 0.6microm - 0.9microm x 1.2microm - 2.0 microm, has one long flagellum on one end of the cell and contains lamellar intracytoplasmic membrane(ICM) system parallel to cytoplasmic membrane. Cultures produce red pigments in the light. Live cells under phototrophic conditions contain bacteriochlorophyll a and carotenoids. 2c grows anaerobically in the light and aerobically in the dark. Optimal growth occurs at 30degreesC - 35degreesC and at pH7.0 -pH8.0. Various organic compounds are used as photosynthetic electron donors and carbon sources. Peptone and (NH4 )2SO4 are its better nitride source,yeast extracts stimulates its growth. A phylogenetic analysis based on 16S rDNA gene sequences reveales that strain 2c gathers a cluster with 3 strains of Rhodopseudomonas palustris whose accession number in GenBank are AY751758, DQ001155, DQ001158, respectively. 2c subcultures 15 generations stably at least. The results presented here demonstrated strain 2c is Rhodopseudomonas palustris.

[Isolation and identification of Rhodopseudomonas acidophila from Fujian province, China].

A strain P301 of Gram negative purple nonsulfur bacterium was isolated from the ditch's mud collected in Fuzhou, Fujian. The cells are rod-shaped. Multiplication occur by budding without stalk formation. Growth optimum pH is 5.0-5.7. No growth factors required. The cells contain bacteriochlorophyll a. The photosynthetic membrane system consists of parallel lamellae. According to Bergery's Manual of Determinative Bacteriology, 8th ed. (1974) and 9th ed. (1994), the strain was identified to be Rhodopseudomonas acidophila. But on the side of enzyme activity, utilization of organic substrates and others the strain some differ from the type strain described by N Pfennig (1969). Therefore the strain P301 was identified to be Rhodopseudomonas acidophila var. fujianensis n. var.

Acyclic carotenoid and cyclic apocarotenoid cleavage by an orthologue of lignostilbene-α,ß-dioxygenase in Rhodopseudomonas palustris.

Carotenoid cleavage oxygenases catalyse formation of apocarotenoids and the precursors of phytohormones, abscisic acid and strigolactones through oxidative cleavage at specific double bonds of carotenoids. A gene encoding a presumed bacterial oxygenase homologous to lignostilbene-α,ß-dioxygenases has been found in the genome of Rhodopseudomonas palustris. By analysing apocarotenoids in recombinant Escherichia coli strains, it was found that the presumed oxygenase catalyses the 15,15' double bond cleavage of lycopene and neurosporene. Cell lysate containing the recombinant protein cleaved all-trans-ß-apo-8'-carotenal at the 15,15' double bond into retinal and apo-8',15'-apocarotene-dial. These data demonstrate for the first time that the orthologue of lignostilbene-α,ß-dioxygenase found in the carotenogenic phototrophic bacterium has the 15,15' double bond cleavage activity towards both the acyclic carotenoids and cyclic apocarotenoid.

Bioconversion characteristics of Rhodopseudomonas palustris CQK 01 entrapped in a photobioreactor for hydrogen production.

The performance of the entrapped-cell photobioreactor during H2 production was assessed by using glucose as substrate in a continuous operation mode. The maximal hydrogen production rate and light conversion efficiency, 2.61 mmol/L/h and 82.3%, were obtained at a HRT of 11.4 h, an substrate loading rate of 4.2 mmol/h and an illumination of 590 nm and 6000 lux, the corresponding hydrogen yield and total energy efficiency were 0.62 mmol H2/(mmol glucose) and 4.8%, respectively. The results indicate the H2 production system illuminated at 590 nm wavelength engaged in energy storage for H2 production due to more ATP synthesized in primary reaction center, and was of higher energy recovery capacity. Furthermore, the total energy efficiency was far lower than the corresponding light conversion efficiency due to intermediates production.

Enhanced hydrogen production by Rhodopseudomonas palustris CQK 01 with ultra-sonication pretreatment in batch culture.

In the present study, the photo-hydrogen production performances by Rhodopseudomonas palustris CQK 01 growing from the inoculated cells with ultra-sonication pretreatment (R. palustris CQK 01-USP) were experimentally investigated in batch culture and compared with that without pretreatment (R. palustris CQK 01-NP). It was found that the ultra-sonication pretreatment modified membrane morphology and broke up part of the cells, resulting in improvement of membrane permeability and bacterial activities and hence, helping the improvement of hydrogen production. The hydrogen production rate, hydrogen yield and energy conversion efficiency with R. palustris CQK 01-USP were increased to be nearly 2 times higher than that with R. palustris CQK 01-NP. The parametric study showed that under the conditions of initial glucose concentration 50 mmol/l, inoculum size 12%, illumination wavelength 590 nm, the photobioreactor with R. palustris CQK 01-USP obtained the optimal hydrogen production rate 0.54 mmol/l/h, hydrogen yield 1.2 mol-H2/mol-glucose and energy conversion efficiency 9.03%.