Phylogeny of anoxygenic filamentous phototrophic bacteria of the family Oscillochloridaceae as inferred from comparative analyses of the rrs, cbbL, and nifH genes

Phylogeny of anoxygenic filamentous phototrophic bacteria (AFPB) of the family Oscillochloridaceae (Oscillochloris trichoides DG6^T and the recently isolated strains Oscillochloris sp. R and C6) was studied based on comparative analyses of the genes coding for 16S rRNA (rrs), ribulose-1,5-bisphosphate carboxylase/oxygenase (cbbL), and nitrogenase (nifH). The sequences of the genes studied proved to be identical in the three strains, which is in agreement with data obtained earlier that showed a lack of differentiating phenotypic distinctions between these strains; therefore, it is proposed that the new strains should be identified as representatives of the species O. trichoides. Using an earlier designed system of oligonucleotide primers and a specially designed additional primer, fragments of the cbbL genes of the “red-like” form I RuBisCO were amplified and sequenced for all of the O. trichoides strains. Analysis of the cbbL genes suggested a separate position of the bacteria studied in the phylogenetic tree, where O. trichoides strains formed an independent branch, which, apart from this species, also included the only studied species of gram-positive facultatively chemoautotrophic bacteria, Sulfobacillus acidophilus. In the phylogenetic tree inferred from the analysis of nifH genes, the bacteria under study also formed a new separate branch, deviating near the root, which indicated a lack of relatedness between them and other phototrophic bacteria. The data obtained support the conclusion that AFPB has an ancient origin and their allocation as one of the main evolutionary lineages of eubacteria, which was made based on the analysis of ribosomal genes.     

Phylogeny of Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase Genes in Haloalkaliphilic Obligately Autotrophic Sulfur-Oxidizing Bacteria of the Genus <Emphasis Type="Italic">Thioalkalivibrio</Emphasis>

Fragments of genes of the “green-like” form I ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) of eight species of haloalkaliphilic obligately autotrophic sulfur-oxidizing bacteria of the genus Thioalkalivibrio have been revealed and sequenced using previously developed oligonucleotide primers. The data obtained are used for the construction of phylogenetic trees on the basis of nucleotide sequences of RuBisCO genes and their conceptual translations into amino acid sequences. Comparative analysis of the 16S rRNA and RuBisCO gene trees reveals discrepancies between their topologies. According to a RuBisCO gene analysis, the genus Thioalkalivibrio is not monophyletic, and its inner divergence conforms to the significant morphological differences observed between the species. Presumably, horizontal (interspecies) gene transfer was involved in the evolution of the genus Thioalkalivibrio.__________Translated from Mikrobiologiya, Vol. 74, No. 3, 2005, pp. 378–386.Original Russian Text Copyright © 2005 by Tourova, Spiridonova, Berg, Kuznetsov, Sorokin.     

基于pufM基因的乌梁素海富营养化湖区好氧不产氧光合细菌系统发育多样性分析

好氧不产氧光合细菌(AAPB)的多样性在海洋中已经广泛研究,但在富营养化湖泊中却研究甚少。通过构建和分析AAPB光合中心合成中的关键基因pufM克隆文库,以期揭示乌梁素海富营养化湖区AAPB分布及其系统发育多样性,探讨其在富营养化湖泊中的功能和作用。对乌梁素海红圪卜湖区水体文库中的52个克隆子进行分析,产生了28个OTU,文库覆盖度达到71.4%,反映出文库有较好的代表性。同源性和系统发育分析结果表明,乌梁素海红圪卜湖区AAPB有较高的多样性,与我们之前所发现的同一湖区总细菌多样性较低形成鲜明对比。所获得的序列分属7个亚群,即γ-Proteobacteria(44.2%,含Group-1,-2和-3共3个亚群)、β-Proteobacteria(21.2%)、Rhodobacter-like(7.69%)及2个未知亚群unknown Group-1(21.2%)和Group-2(5.77%)。其中γ-Proteobacteria占到总克隆的44.2%,在低盐的乌梁素海环境中出现高比例的γ-Proteobacteria之前并未见类似报道,并且乌梁素海中存在一些可能是富营养化湖泊特有的AAPB。这表明AAPB可能在富营养化湖泊生态系统的维持和稳定中有重要作用。     

Regulation of Rubisco by inhibitors in the light

2-carboxy-D-arabinitol-1-phosphate (CA1P) bound to Rubisco either in leaf extracts or after purification can be displaced by SO₄^2? ions. Thus, treatment of leaf extracts with a buffer containing 200 mol m^?3 SO₄^2? displaces any bound CA1P and enables measurement of maximum car-boxylation potential. In tobacco leaves, the activity following treatment with SO₄^?2 ions (‘maximal activity’) is greater than the total Rubisco activity. The ratio of the two activities altered in a dynamic way with fluctuations in irradiance. Even in species which do not produce significant amounts of CA1P, the maximal activity greatly exceeded the total activity. Anion exchange separation of components in acid extracts confirmed the absence of CA1P in tobacco leaves harvested above an irradiance of 300 μmol quanta m^?2 s^?1, but the presence of another inhibitor of Rubisco. These results are consistent with the regulation of Rubisco activity by inhibitors other than CA1P which, like CA1P, can be displaced by SO₄^2? ions.     

A point mutation in the N-terminus of ribulose-1,5-bisphosphate carboxylase affects ribulose-1,5-bisphosphate binding

Mutagenesis in vitro of the gene encoding the large subunit of ribulose-1,5-bisphosphate carboxylase/ oxygenase (EC 4.1.1.39) from Anacystis nidulans was used to generate novel enzymes. Two conserved residues, threonine 4 and lysine 11 in the N-terminus were changed. The substitution of threonine 4 with serine or valine had little effect on the kinetic parameters. The substitution of lysine 11 with leucine, which is non-polar, increased the K _m for ribulose-1,5-bisphosphate from 82 to 190 M but its replacement with glutamine, which has polar properties, had no appreciable effect.Abbreviations Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase - RuBP ribulose-1,5-bisphosphate - LSU large sub-unit of Rubisco - SSU small subunit of Rubisco We thank Dr. S. Gutteridge (DuPont, Wilmington, USA) for structural information and for his comments on the results described. The technical assistance of Mr. A. Cowland and Mr. I. Major was invaluable.     

Chloroplastic Carbon Dioxide Concentration in Norway Spruce (Picea Abies [L.] Karst.) needles relates to the position within the crown

Differences between sun (E) and shaded (S) foliage were studied in a Norway spruce (Picea abies [L.] Karst.) stand. Response curves describing the dependence of the CO2 assimilation rate (PN) on the CO2 concentration at the catalytic site of ribulose-1,5-bisphosphate carboxylase/oxygenase, RuBPCO (PN-Cc) were estimated using the simultaneous measurements of chlorophyll fluorescence and leaf gas exchange. Higher PN, higher electron transport (Ja), higher carboxylation capacity (Vc), and higher RuBPCO activity () for sun acclimated needles was found. The S-needles had higher portion of internal limitation and higher CO2 compensation concentration () than the E-needles. Because higher degree of limitation of photosynthesis by carboxylation was ascertained, it can be assumed that photosynthesis in shade foliage is limited mainly by lower carboxylation capacity and by low chloroplastic CO2 concentration     

Growth and Photosynthetic Characteristics of Ceriops Roxburghiana under NaCl Stress

The plant growth, net photosynthetic rate (PN), intercellular CO2 concentration (ci), and dry matter production of Ceriops roxburghiana Arn. were significantly increased with increasing salinity from 0 to 400 mM NaCl. At 600 mM NaCl, shoot and root lengths, and dry mass were significantly depressed with respect to control. Absence of diurnal fluctuation of concentrations of organic acids, and the low activity of phosphoenolpyruvate carboxylase and high activity of ribulose-1,5-bisphosphate carboxylase confirmed the operation of C3 pathway in Ceriops even at increasing salinity.     

Km (CO2) values of ribulose-1,5-bisphosphate carboxylase in grasses of different C4 type

Statistical analysis of K_m (CO₂) values of ribulose-1,5-bisphosphate (RuBP) carboxylase from 35 C₄ grass species shows that the mean value for PEP-carboxykinase (PCK) type C₄ species (41.4±s.e. 2.2 μM CO₂) is significantly different from that of NAD-malic enzyme (NAD-ME) type species (55.3±3.1 μM CO₂) or NADP-malic enzyme (NADP-ME type species (52.5±s.e. 2.0μM CO₂). These C₄ type differences remain detectable within both the eu-panicoid and chloridoid grass subfamilies. By contrast, no between-subfamily differences were found within C₄ types. Variation in K_m (CO₂) values of RuBP carboxylase may be related to in vivo differences in CO₂ concentration at the enzyme site, mediated perhaps by differences in CO₂-leakiness of C₄ leaf ‘photosynthetic carbon reduction’ (PCR or ‘Kranz’) tissue.     

Seasonal Changes in the Structure of the Anoxygenic Phototrophic Bacterial Community in Lake Mogilnoe,a Relict Lake on Kil'din Island in the Barents Sea

An anaerobic phototrophic bacterial community in Lake Mogilnoe, a relict lake on Kil'din Island in the Barents Sea, was studied in June 1999 and September 2001. Irrespective of the season, the upper layer of the anaerobic zone of this lake had a specific species composition of sulfur phototrophic bacteria, which were dominated by the brown-colored green sulfur bacterium Chlorobium phaeovibrioides. The maximum number of sulfur phototrophic bacteria was observed in June 1999 at a depth of 9 m, which corresponded to a concentration of bacteriochlorophyll (Bchl) e equal to 4.6 mg/l. In September 2001, the maximum concentration of this pigment (3.4 mg/l) was found at a depth of 10 m. In both seasons, the concentration of Bchl a did not exceed 3 μg/l. Purple sulfur bacteria were low in number, which can be explained by their poor adaptation to the hydrochemical and optical conditions of the Lake Mogilnoe water. In June 1999, the water contained a considerable number of Pelodictyon phaeum microcolonies and Prosthecochloris phaeoasteroides cell chains, which was not the case in September 2001. A 16S rDNA-based phylogenetic analysis of pure cultures of phototrophic bacteria isolated from the lake water confirmed that the bacterial community is dominated by Chl. phaeovibrioides and showed the presence of three minor species, Thiocystis gelatinosa, Thiocapsa sp., and Thiorhodococcus sp., the last of which is specific to Lake Mogilnoe.     

Responses of Penultimate and Flag Leaves of Wheat to Different Nitrogen Supply

Uniculm wheat (Triticum aestivum L.) was grown to maturity at four concentrations of nitrogen corresponding to 3 (N1), 6 (N2), 9 (N3) and 12 (N4) g m^–2. Penultimate and flag leaves were examined throughout the ontogeny. Sub-optimal concentrations of N resulted in sharp decline in both area and dry mass of the leaves. Decline in leaf area was due to fewer mesophyll cells. Net photosynthetic rate (P_N) increased up to full expansion, remained constant for about a week and then declined. P_N, nitrogen, ribulose-1,5-bis-phosphate carboxylase/oxygenase (RuBPCO) amount and activity, chlorophyll and soluble protein contents were similar at all the N concentrations. Both amount and activity of RuBPCO in the flag leaf were about two fold higher as compared to penultimate leaf, but P_N was similar. This indicates the presence of an excess amount of RuBPCO in the flag leaf.     

Effects of Phytoplasma Infection on Pigments, Chlorophyll-Protein Complex and Photosynthetic Activities in Field Grown Apple Leaves

Changes in contents of pigments, chlorophyll-protein complex, and photosynthetic activities were investigated in field grown apple (Malus pumila Mill.) leaves infected by Apple Proliferation phytoplasma. The contents of chlorophyll a+b (Chl) and carotenoids (Car) markedly decreased in infected leaves. Similar results were also observed for content of total soluble proteins and ribulose-1,5-bisphosphate carboxylase activity. When various photosynthetic activities were followed in isolated thylakoids, phytoplasma infection caused a marked inhibition of whole chain and photosystem 2 (PS2) activity. Smaller inhibition of photosystem 1 (PS1) activity was observed even in severely infected leaves. The artificial exogenous electron donors, MnCl₂ diphenyl carbazide, and NH₂OH, did not restore the loss of PS2 activity in both mildly and severely infected leaves. Similar results were obtained by Chl fluorescence measurements. The marked loss of PS2 activity in infected leaves was due to the reduction of contents of chlorophyll and light-harvesting chlorophyll-protein 2 complexes. This revised version was published online in July 2006 with corrections to the Cover Date.     

Small-angle X-ray study on the structure of ribulose-1,5-bisphosphate carboxylase-oxygenase from Rhodospirillum rubrum

The quaternary structure of ribulose-1,5-bisphosphate carboxylase-oxygenase (rubisco) from Rhodospirillum rubrum, an enzyme consisting of two large subunits, L₂, was investigated by small-angle X-ray scattering. In the presence of HCO ₃ ^- and Mg²⁺, rubisco is in the active state and displays a radius of gyration of 2.96 nm, a maximum diameter of 9.5 nm and a volume of 170 nm³. A model is presented where the subunits are arranged back-to-back, rotated relative to each other by 90°, and shifted by 1.3 nm. Upon inactivation by removal of HCO ₃ ^- and Mg²⁺, the model swells slightly without any distinct changes in configuration. This contrasts with our previous observations with rubisco from Alcaligenes eutrophus, an enzyme composed of small (S) and large (L) subunits, L₈S₈, where inactivation gives rise to substantial changes in configuration.Abbreviations RuBP Ribulose-1,5-bisphosphate - 3-PGA 3-phosphoglyceric acid     

Photosynthetic carbon fixation in the corollas of Petunia hybrida

Corollas of Petunia hybrida (cv. Hit Parade Rosa) flowers fixed ¹⁴CO₂ under both light and dark conditions. Rates of light fixation were much higher in mature pink corollas than in young, green corollas [57 and 9 nmol (ngchl)¹ min^-1], paralleling the development of chloroplasts in these tissues. Stomatal conductance in corollas was only 12% of that in green leaves, mainly due to the presence of few, and non-functioning stomata in the corolla. The activity and concentration of ribulose bisphosphate carboxylase (EC 4.1.1.39) in corolla extracts were only about 30% (per unit Chi) of those in extracts from green leaves. These results, together with previous results, might indicate a coordinated reduction in activity of systems participating in photosynthesis in corollas. The fixation products following a 6 s pulse with ¹⁴CO₂, were typical of C, plants in both corollas and green leaves, but a higher level of β-carboxylation products was found in the corollas. The activity of phosphoenol-pyruvate carboxylase (EC 4.1.1.31) (per unit protein) was similar in both tissues. Although the total carbon fixed by the corolla constituted only a small part of the metabolites required for flower development, certain photosynthetic metabolites might have a regulatory role in flower development.     

Elevated CO<Subscript>2</Subscript> and temperature differentially affect photosynthesis and resource allocation in flag and penultimate leaves of wheat

Differences in acclimation to elevated growth CO₂ (700 μmol mol⁻¹, EC) and elevated temperature (ambient +4 °C, ET) in successive leaves of wheat were investigated in field chambers. At a common measurement CO₂, EC increased photosynthesis and the quantum yield of electron transport (Φ) early on in the growth of penultimate leaves, and later decreased them. In contrast, EC did not change photosynthesis, and increased Φ at later growth stages in the flag leaf. Contents of chlorophyll (Chl), ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCO), and total soluble protein were initially higher and subsequently lower in penultimate than flag leaves. EC decreased RuBPCO protein content relative to soluble protein and Chl contents throughout the development of penultimate leaves. On the other hand, EC initially increased the RuBPCO:Chl and Chl a/b ratios, but later decreased them in flag leaves. In the flag leaves but not in the penultimate leaves, ET initially decreased initial and specific RuBPCO activities at ambient CO₂ (AC) and increased them at EC. Late in leaf growth, ET decreased Chl contents under AC in both kinds of leaves, and had no effect or a positive one under EC. Thus the differences between the two kinds of leaves were due to resource availability, and to EC-increased allocation of resources to photon harvesting in the penultimate leaves, but to increased allocation to carboxylation early on in growth, and to light harvesting subsequently, in the flag leaves.     

Nitrogen accumulation and mobilization in Citrus leaves throughout the annual cycle

The changes of protein and amino acid contents of young and old leaves from field citrus trees [ Citrus sinensis (L.) Osbeck cv. Washington Navel] were studied throughout the year. the total protein content of old leaves decreased during the spring and summer growths, whereas in young developing leaves it increased rapidly. The end of the spring and summer flushes was followed by a quick recovery of the initial protein content due to a process of reversible senescence. The evolution of SDS-PAGE proteinograms indicated that most of the foliar proteins contributed to the nitrogen mobilization during the spring and summer growth periods. A protein band of molecular weight ca 55,000, probably containing the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase, decreased relatively more than the others. The amino acid content of the leaves increased during winter time due to the accumulation of proline, which was mobilized during the spring flush. The possible nitrogen storage function of proline is discussed.     

Potential Consequences of Virus Infection for Shade-Sun Acclimation in Leaves

Attention is drawn to literature evidence that leaf virus infection may interfere with synthesis of ribulose-1,5-bisphosphate carboxylase oxygenase, and with the activity of photosystem II, two main requirements for shade-sun acclimation. Preliminary experiments which show that virus infections resulting in these interactions can lead to photosynthetic physiology diagnostic of nitrogen limited plants and/or shade ecotypes are presented. Other observations on the diversity of virus infections in sun populations of wild species commonly found in the shade are presented. Together, this evidence leads to the hypothesis that virus infection may well confine individuals of some species to shaded habitats because infection prevents acclimation to bright light, and predisposes them to photoinhibition.     

The Model of Calvin Cycle Enzyme Organization on Thylakoid Membranes with the Involvement of the Photosystem I Lectin

Based on our own and other researchers' experimental data, a model of the light-induced assembly of Calvin cycle enzymes on the membranes of stromal thylakoids is suggested. According to this model, the components involved are: (1) the photosystem I chlorophyll–protein complex containing the P700 reaction center, which possesses a stroma-exposed and pH-dependent carbohydrate-binding site performing both photo- and chemoreceptor functions, and (2) ribulose-1,5-bisphosphate carboxylase (Rubisco), a glycoenzyme, which is a specific ligand of the lectin in the stromal thylakoids. After stroma alkalization in light, the carbohydrate-binding site of the photosystem I pigment–lectin complex is transformed into an active state, and this transformation increases the enzyme carboxylase activity.