三种紫球藻亲缘关系RAPD分析

利用RAPD技术对铜绿紫球藻（Porphyridium aerugineum 755）,淡色紫球藻（Porphyridium purpureum 806）和紫球藻（Porphyridium cruentum）的亲缘关系进行分析。从50个随机引物中筛选出25个种间多态性较强,重复性较好的引物。检测到233个位点。在233个条带中有186个多态性位点。多态位点比率为79.73%,平均每个引物扩增9个条带,多态性条带7个。聚类分析结果表明：铜绿紫球藻、淡色紫球藻和紫球藻之间的平均遗传距离为0.455,其中铜绿紫球藻和淡色紫球藻之间的遗传距离为0.413,铜绿紫球藻和紫球藻之间的遗传距离为0.556,淡色紫球藻和紫球藻之间的遗传距离最小为0.396。所以由RAPD试验的分析结果可以得出:三种紫球藻为独立的种, 其中淡色紫球藻和紫球藻是亲缘关系较近的两个不同种。紫球藻属种间个体DNA多态性比较丰富,因此利用RAPD技术可以从DNA水平上检测紫球藻属种间差异。     

A selenite-induced decrease in the lipid content of a red alga

Two unicellular marine algae (Dunaliella primolecta and Porphyridium cruentum) have been found to contain a selenium-inducible, non-enzymatic glutathione peroxidase activity when cultured in the presence of selenite. To test the possibility that selenium functions in vivo as an antioxidant in these algae, a detailed examination of the lipid content of algae cultured in the presence or absence of selenite was conducted. If selenium augments the antioxidant defenses of algal cells, an increase in the content of oxidation-sensitive lipids would be expected. The fatty acid, chlorophyll, phospholipid and glycolipid content of the green alga D. primolecta was not affected by growth in selenite. At low light intensity there was a moderate decrease in the chlorophyll and polyunsaturated fatty acid content of the red alga P. cruentum when cultured in selenite. At higher light intensity the content of all fatty acids, phospholipid, glycolipid, chlorophyll, carotenoid and phycoerythrin decreased in P. cruentum grown in selenite. Since growth in selenite did not increase the quantity of oxidation-sensitive lipids in either alga, there is no evidence for an in vivo functioning of selenium as an antioxidant. Instead, the observed decrease in lipids of the red alga P. cruentum can best be explained as a selenite-induced oxidative effect.     

A selenium-induced peroxidation of glutathione in algae

Two unicellular marine algae cultured in media containing sodium selenite were examined for glutathione peroxidase activity. The 400 g supernatant from disrupted cells of both the green alga Dunaliella primolecta and the red alga Porphyridium cruentum were able to enhance both the H₂O₂ and the tert-butyl hydroperoxide dependent oxidation of glutathione. The glutathione peroxidation activity of D. primolecta was reduced only slightly by heating the 400 g supernatant, a 30% decrease in the rate with H₂O₂ and 10% decrease in the rate with t-BuOOH being observed. Heating caused the H₂O₂ dependent activity in P. cruentum to be reduced by only 30%, but the activity with t-BuOOH was reduced by 90%. Freezing decreased the t-BuOOH dependent activity of P. cruentum by 90%, but did not lower the t-BuOOH dependent activity of D. primolecta or the H₂O₂ dependent activity of either alga. It was concluded that the heat and cold stable, glutathione peroxidation was non-enzymatic in nature. A variety of small molecules (ascorbate, Cu(NO₃)₂, selenocystine, dimethyldiselenide and selenomethionine) were shown to be able to enhance the hydroperoxide dependent oxidation of glutathione in the assay system employed in this study. Such compounds could be responsible for the activity observed in algae. The heat and cold labile t-BuOOH reductase activity of P. cruentumwas possibly enzymatic, but was not attributable to the presence of glutathione-S-transferase. Both algae, when cultured in the presence of added selenite, displayed an approximate doubling of the non-enzymatic H₂O₂ and t-BuOOH dependent glutathione oxidase activities. The heat and cold labile t-BuOOH reductase activity of P. cruentum was unaltered when the alga was grown in the presence of added selenite. These observations are consistent with the hypothesis that selenium compounds present in the algae are responsible for the selenium induced glutathione peroxidation.     

Mixotrophic production of phycoerythrin and exopolysaccharide by the microalga Porphyridium cruentum

The ability of the unicellular rhodophyte Porphyridium cruentum to grow mixotrophically on the soluble fraction of Solarium tuberosum meal was tested. At the beginning of stationary phase Porphyridium cruentum produced 7 μg ml⁻¹ of phycoerythrin and 129 μg ml⁻¹ of total soluble exopolysaccharide when cultured autotrophically. When cultured mixotrophically with the soluble fraction of Solanum tuberosum meal, the productivity increased to 10 μg ml⁻¹ of phycoerythrin and 330 μg ml⁻¹ of total soluble exopolysaccharide. When the soluble fraction of S. tuberosum meal was supplied together with nitrate and phosphate, the productivity of phycoerythrin increased to 21 μg ml⁻¹ while the production of total soluble exopolysaccharide decreased to 195 μg ml⁻¹. Results demonstrate that the soluble fraction of S. tuberosum meal can be used as substrate for the production of phycoerythrin and exopolysaccharide by P. cruentum improving the results obtained with the autotrophic culture medium.     

The extracellular polysaccharides of porphyridium cruentum and porphyridium aerugineum

The extracellular mucilages from Porphyridium cruentum and P. aerugineum contain D-xylose, D-glucose, D-and L-galactose, 3-O-methylxylose, 3- and 4-O-methyl-galactose, and D-glucoronic acid in the approximate molar proportions of 3:1:2.5:0.13:0.13:0.8 and 1.7:1:1.1:0.3:0.6:0.5, respectively. In addition, P cruentum mucilage contains 2-O-methylhexose (0.13) and 2-O-methylglucuronic acid (0.2), whereas P. aeruginum mucilage is devoid of these two sugars but contains 2,4-di-O-methylgalactose (0.5). Both polysaccharides contain ～10% of half-ester sulphate and appear to be linked to ～5% of protein. Attempted fractionation into homopolysaccharides was unsuccessful. Methylation, periodate oxidation, and partial hydrolysis studies revealed that the glucuronic acid is 1,3-linked and is attached solely to O–3 of D-galactose in both mucilages. The 2-O-methylglucuronic acid in P. cruentum is linked to O–4 of L-galactose. Xylose, glucose, and galactose are present in both mucilages as end groups, and 1,3- and 1,4-linked residues, with galactose and glucose also present as, 1,3,4-linked or sulphated residues. Molecular weight determinations on Sepharose 4B indicate a $\text{M}$_w of 4 x 10(su6) for P. cruentum mucilage and 5 x 10⁶ for that from P. aerugineum.     

Overproduction of gamma-Linolenic and Eicosapentaenoic Acids by Algae

The pharmaceutical interest and limited availability of γ-linolenic acid (GLA) and eicosapentaenoic acid (EPA) prompted the search for genetic means for increasing the production of these fatty acids from algal sources. Cell lines of Spirulina platensis and Porphyridium cruentum resistant to the growth inhibition of the herbicide Sandoz 9785 were selected by serial transfers of the culture in the presence of increasing concentrations of the herbicide. The resistant cell lines of S. platensis overproduced GLA and those of P. cruentum overproduced EPA and were stable for at least 50 generations in the absence of the inhibitor.     

Isolation and Function of Allophycocyanin B of Porphyridium cruentum

Allophycocyanin B was purified to homogeneity from the eukaryotic red alga Porphyridium cruentum. This biliprotein is distinct from the allophycocyanin of P. cruentum with respect to subunit molecular weights, and spectroscopic and immunological properties. The purified allophycocyanin B has a long wavelength absorption maximum at 669 nm at room temperature and at 675 nm at −196 C while the fluorescence emission maximum is at 673 nm at room temperature and 679 nm at −196 C. The emission spectrum of allophycocyanin shifted only 1 nm, from 659 to 660 nm, on cooling to −196 C, and was the same with allophycocyanin crystals as it was with pure solutions of the pigment. Phycobilisomes from P. cruentum have a major fluorescence emission band at 680 nm at −196 C which emanates from the small amount of allophycocyanin B present in the phycobilisomes. Light energy absorbed by the bulk of the biliprotein pigments is transferred to allophycocyanin B with high efficiency.     

Preparation of different molecular weight polysaccharides from Porphyridium cruentum and their antioxidant activities

Hermetical microwave was used to degrade Porphyridium cruentum polysaccharides from 2918 to 256.2, 60.66 and 6.55 kDa. The antioxidant properties of different molecular weight polysaccharides were evaluated by determining the scavenging ability of free radicals, inhibitory effects on lipid peroxidation in liver homogenates and hemolysis of mouse erythrocytes. Analysis of physicochemical properties confirmed that microwave degradation might not markedly change the chemical components of the polysaccharides. High-molecular-weight polysaccharides from P. cruentum had no obvious antioxidant activity, but low-molecular-weight fragments after degradation exerted an inhibitory effect on oxidative damage. The 6.55-kDa fragment had stronger antioxidant activity than the 60.66 and 256-kDa fragments.     

Photosynthetic Light Responses May Explain Vertical Distribution of Hymenophyllaceae Species in a Temperate Rainforest of Southern Chile

Some epiphytic Hymenophyllaceae are restricted to lower parts of the host (<60 cm; 10–100 μmol photons m^-2 s^-1) in a secondary forest of Southern Chile; other species occupy the whole host height (≥10 m; max PPFD >1000 μmol photons m^-2 s^-1). Our aim was to study the photosynthetic light responses of two Hymenophyllaceae species in relation to their contrasting distribution. We determined light tolerance of Hymenoglossum cruentum and Hymenophyllum dentatum by measuring gas exchange, PSI and PSII light energy partitioning, NPQ components, and pigment contents. H. dentatum showed lower maximum photosynthesis rates (A_max) than H. cruentum, but the former species kept its net rates (A_n) near A_max across a wide light range. In contrast, in the latter one, A_n declined at PPFDs >60 μmol photons m^-2 s^-1. H. cruentum, the shadiest plant, showed higher chlorophyll contents than H. dentatum. Differences in energy partitioning at PSI and PSII were consistent with gas exchange results. H. dentatum exhibited a higher light compensation point of the partitioning of absorbed energy between photochemical Y(PSII) and non-photochemical Y(NPQ) processes. Hence, both species allocated energy mainly toward photochemistry instead of heat dissipation at their light saturation points. Above saturation, H. cruentum had higher heat dissipation than H. dentatum. PSI yield (YPSI) remained higher in H. dentatum than H. cruentum in a wider light range. In both species, the main cause of heat dissipation at PSI was a donor side limitation. An early dynamic photo-inhibition of PSII may have caused an over reduction of the Qa⁺ pool decreasing the efficiency of electron donation to PSI. In H. dentatum, a slight increase in heat dissipation due to acceptor side limitation of PSI was observed above 300 μmol photons m^-2s^-1. Differences in photosynthetic responses to light suggest that light tolerance and species plasticity could explain their contrasting vertical distribution.     

Energy transfer from photosystem II to photosystem I in Porphyridium cruentum

Rates of photooxidation of P-700 by green (560 nm) or blue (438 nm) light were measured in whole cells of Porphyridium cruentum which had been frozen to ?196 °C under conditions in which the Photosystem II reaction centers were either all open (dark adapted cells) or all closed (preilluminated cells). The rate of photooxidation of P-700 at ?196 °C by green actinic light was approx. 80% faster in the preilluminated cells than in the dark-adapted cells. With blue actinic light, the rates of P-700 photooxidation in the dark-adapted and preilluminated cells were not significantly different. These results are in excellent agreement with predictions based on our previous estimates of energy distribution in the photosynthetic apparatus of Porphyridium cruentum including the yield of energy transfer from Photosystem II to Photosystem I determined from low temperature fluorescence measurements.     

Action Spectra for Photosystems I and II in Formaldehyde Fixed Algae

Action spectra were obtained for photosystems I and II in chemically fixed algal cells and for photosystem I in unfixed lysozyme treated cells. Untreated algal cells yielded neither of the 2 light reactions with the reaction mixtures used. The action spectra for photosystem I in the blue-green alga Anacystis nidulans and red alga Porphyridium cruentum follow the absorption spectrum of chlorophyll a with a small peak in the region of the accessory pigments. In the green alga Chlorella pyrenoidosa the photosystem I action spectrum follows the absorption spectrum of chlorophyll a. Photosystem II action spectra in A. nidulans and P. cruentum follow the absorption spectra of the accessory pigments while that in C. pyrenoidosa is shifted slightly toward the blue spectral region. These results provide additional evidence that formaldehyde fixed cells are valid models for studying the light reactions of photosynthesis.     

Comparative properties of ferredoxins from a marine and freshwater species of Porphyridium

Ferredoxins were isolated from the freshwater red alga Porphyridium aerugineut, and from Porphyridium cruentum, a related marine species. A sin     

Effective Absorption Cross-Sections in Porphyridium cruentum: Implications for Energy Transfer between Phycobilisomes and Photosystem II Reaction Centers

Effective absorption cross-sections for O₂ production by Porphyridium cruentum were measured at 546 and 596 nanometers. Although all photosystem II reaction centers are energetically coupled to phycobilisomes, any single phycobilisome acts as antenna for several photosystem II reaction centers. The cross-section measured in state I was 50% larger than that measured in state II.     

Influence of microalgae cell wall characteristics on protein extractability and determination of nitrogen-to-protein conversion factors

Additional evidence about the influence of the cell wall physical and chemical characteristics on protein extractability was determined by calculating the conversion factors of five different microalgae known to have different cell wall composition, and their protein extracts. The conversion factors obtained for crude rigid cell walled Chlorella vulgaris, Nannochloropsis oculata and Haematococcus pluvialis were 6.35, 6.28 and 6.25, respectively, but for their protein extracts the values were lower with 5.96, 5.86 and 5.63. On the other hand, conversion factor obtained for fragile cell walled microalgae Porphyridium cruentum and Athrospira platensis was 6.35 for the former and 6.27 for the latter, with no significant difference for their protein extract with 6.34 for the former and 6.21 for the latter. In addition, the highest hydro-soluble protein percentage recovered from total protein was for P. cruentum 80.3 % and A. platensis 69.5 % but lower for C. vulgaris with 43.3 %, N. oculata with 33.3 % and H. pluvialis with 27.5 %. The study spotted the light on the influence of the cell wall on evaluating the conversion factor and protein extractability. In addition, it showed the necessity of finding the conversion factor every time accurate protein quantification is required, and proved that there is not a universal conversion factor that can be recommended.     

Mechanism of the light state transition in photosynthesis. IV. Picosecond fluorescence spectroscopy of Anacystis nidulans and Porphyridium cruentum in state 1 and state 2 at 77 K

The sequential energy-transfer pathway through the phycobilin pigments to chlorophyll a was investigated as a function of the state transition in the cyanobacterium Anacystis nidulans and the red alga Porphyridium cruentum. The fluorescence decay kinetics of the phycobilin pigments and chlorophyll a were determined for cells frozen at 77 K in state 1 and state 2 using a single-photon timing fluorescence spectroscopy apparatus with picosecond resolution. Time-resolved 77 K fluorescence emission spectra were also obtained for both species in state 1 and state 2. In both A. nidulans and P. cruentum the transition to state 1 was accompanied by a large increase in the apparent fluorescent lifetime of chlorophyll a associated with PS II (emission peak at 695 nm). There were smaller increases in the lifetime of the terminal phycobilin emitter (685 nm) in both species and no change in phycocyanin (645 nm) or allophycocyanin (660 nm). Time-resolved spectra showed sequential emission from phycocyanin, allophycocyanin, the terminal phycobilin emitter and chlorophyll a. Spectral red shifts were observed with time for all emission peaks with the exception of the terminal phycobilin emitter. In A. nidulans this peak showed a small blue shift with time. The results are interpreted as evidence for an effective uncoupling of PS II chlorophyll a from subsequent energy transfer to PS I chlorophyll a upon transition to state 1. Our recently proposed model for the mechanism of the state transition in phycobilisome-containing organisms is discussed in terms of a decrease in the energy transfer overlap between PS II chlorophyll a and PS I chlorophyll a in state 1.     

Mechanism of the light state transition in photosynthesis. II. Analysis of phosphorylated polypeptides in the red alga,Porphyridium cruentum

The possible involvement of a reversible protein phosphorylation event in the regulation of excitation energy distribution was studied in the red alga Porphyridium cruentum. Whole cells were incubated in phosphate-depleted growth medium containing carrier-free [³²P]orthophosphate for several hours to label the intracellular phosphate pools, and they were then converted to State 1 or State 2 by illumination using blue or green light, respectively. The successful transition to State 1 or State 2 was verified by 77 K fluorescence spectroscopy of the chlorophyll emission and the cells were then denatured using either acetone, trichloroacetic acid or boiling detergent. The whole cell lysates were solubilized, treated with RNAase, and analyzed for phosphoproteins by SDS-polyacrylamide gel electrophoresis. At least twelve polypeptides were found to be phosphorylated but no changes in specific radioactivity of the polypeptides were detected when samples from cells in State 1 and State 2 were compared. We conclude that a reversible protein phosphorylation event is not implicated in the state transition in P. cruentum. A model is presented for the mechanism of the light state transition in organisms that contain phycobilisomes which is different from the mechanism of energy distribution proposed for higher plants.     

Stoichiometry of Photosystem I, Photosystem II, and Phycobilisomes in the Red Alga Porphyridium cruentum as a Function of Growth Irradiance

Cells of the red alga Porphyridium cruentum (ATCC 50161) exposed to increasing growth irradiance exhibited up to a three-fold reduction in photosystems I and II (PSI and PSII) and phycobilisomes but little change in the relative numbers of these components. Batch cultures of P. cruentum were grown under four photon flux densities of continuous white light; 6 (low light, LL), 35 (medium light, ML), 180 (high light, HL), and 280 (very high light, VHL) microeinsteins per square meter per second and sampled in the exponential phase of growth. Ratios of PSII to PSI ranged between 0.43 and 0.54. About three PSII centers per phycobilisome were found, regardless of growth irradiance. The phycoerythrin content of phycobilisomes decreased by about 25% for HL and VHL compared to LL and ML cultures. The unit sizes of PSI (chlorophyll/P₇₀₀) and PSII (chlorophyll/Q_A) decreased by about 20% with increase in photon flux density from 6 to 280 microeinsteins per square meter per second. A threefold reduction in cell content of chlorophyll at the higher photon flux densities was accompanied by a twofold reduction in β-carotene, and a drastic reduction in thylakoid membrane area. Cell content of zeaxanthin, the major carotenoid in P. cruentum, did not vary with growth irradiance, suggesting a role other than light-harvesting. HL cultures had a growth rate twice that of ML, eight times that of LL, and slightly greater than that of VHL cultures. Cell volume increased threefold from LL to VHL, but volume of the single chloroplast did not change. From this study it is evident that a relatively fixed stoichiometry of PSI, PSII, and phycobilisomes is maintained in the photosynthetic apparatus of this red alga over a wide range of growth irradiance.     

Effects of light intensity on the growth rate of the red alga Porphyridium cruentum

The red alga, Porphyridium cruentum, which is one of the potential sources of arachidonic acid, was cultured in batch and continuous vessels. The growth rates in batch cultures were correlated to the mean light intensity in the vessels, and the cell concentrations in continuous cultures were estimated by those results. The yield of arachidonic acid was about 1.2 g per 10¹² cell at cell concentrations ranging from 0.5 to 1.5 × 10¹⁰ cell/l and independent of the mean light intensity.     

Characterization of a Purified Photosystem II-Phycobilisome Particle Preparation from Porphyridium cruentum

Detergent preparations isolated from thylakoids of the red alga Porphyridium cruentum, in a sucrose, phosphate, citrate, magnesium chloride medium consist of phycobilisomes and possess high rates of photosystem II activity. Characterization of these particles shows that the O₂-evolving activity is stable for several hours and the pH optimum is about 6.5 to 7.2. Response of the system to light, electron donors and acceptors, and inhibitors verify that the observed activity, measured both as O₂ evolution and 2,6-dichlorophenol-indophenol reduction, is due to photosystem II. Furthermore, photosystem II is functionally coupled to the phycobilisome in this preparation since green light, absorbed by phycobilisomes of P. cruentum, is effective in promoting both O₂ evolution and 2,6-dichlorophenol-indophenol reduction. Photosystem II activity declines when light with wavelengths shorter than 665 nm is removed. Both 3-(3,4-dichlorophenyl)-1,1-dimethylurea and atrazine inhibit photosystem II activity in this preparation, indicating that the herbicide binding site is a component of the photosystem II-phycobilisome particle.     

Metabolism of delta-Aminolevulinic Acid in Red and Blue-Green Algae

δ-Aminolevulinic acid was incorporated in vivo into C-phycocyanin and B-phycoerythrin in two species of the Rhodophyta (Cyanidium caldarium, Porphyridium cruentum) and three species of the Cyanophyta (Anacystis nidulans, Plectonema boryanum, Phormidium luridum). Amino acid analysis of phycocyanin-¹⁴C from C. caldarium cells which had been incubated with δ-aminolevulinate-4-¹⁴C showed that 84% of the radioactivity incorporated was present in the phycocyanobilin chromophore and less than 16% of the radioactivity cochromatographed with amino acids. These results indicate that δ-aminolevulinate is utilized predominantly via the porphyrin pathway in C. caldarium. Conversely, analysis of phycocyanin-¹⁴C prepared from cells of A. nidulans, P. boryanum, and P. luridum which had been incubated with radiolabeled δ-aminolevulinate demonstrated that 85%, 81%, and 93%, respectively, of the radioactivity incorporated cochromatographed with amino acids. The ratio of incorporated radioactivity in amino acids and phycoerythrobilin was 40:60 in P. cruentum phycoerythrin obtained from cells which had been incubated with δ-aminolevulinate-4-¹⁴C. Succinate-2-3-¹⁴C appeared to be as good a carbon source of amino acids as did C₄ and C₅ of δ-aminolevulinate. These data demonstrate a major alternate route (other than the porphyrin pathway) of δ-aminolevulinate metabolism in red and blue-green algae. The factors responsible for the extent to which δ-aminolevulinate is utilized for synthesis of porphyrins and their derivatives and routes of δ-aminolevulinate catabolism in the organisms employed are discussed.