Red Alga Porphyridium Purpureum Research Articles

Analysis of the complete plastid genome of the unicellular red alga Porphyridium purpureum

We determined the complete nucleotide sequence of the plastid genome of the unicellular marine red alga Porphyridium purpureum strain NIES 2140, belonging to the unsequenced class Porphyridiophyceae. The genome is a circular DNA composed of 217,694 bp with the GC content of 30.3%. Twenty-nine of the 224 protein-coding genes contain one or multiple intron(s). A group I intron was found in the rpl28 gene, whereas the other introns were group II introns. The P. purpureum plastid genome has one non-coding RNA (ncRNA) gene, 29 tRNA genes and two nonidentical ribosomal RNA operons. One rRNA operon has a tRNA(Ala)(UGC) gene between the rrs and the rrl genes, whereas another has a tRNA(Ile)(GAU) gene. Phylogenetic analyses suggest that the plastids of Heterokontophyta, Cryptophyta and Haptophyta originated from the subphylum Rhodophytina. The order of the genes in the ribosomal protein cluster of the P. purpureum plastid genome differs from that of other Rhodophyta and Chromalveolata. These results suggest that a large-scale rearrangement occurred in the plastid genome of P. purpureum after its separation from other Rhodophyta.

Analysis of horizontal genetic transfer in red algae in the post-genomics age

The recently published genome of the unicellular red alga Porphyridium purpureum revealed a gene-rich, intron-poor species, which is surprising for a free-living mesophile. Of the 8,355 predicted protein-coding regions, up to 773 (9.3%) were implicated in horizontal genetic transfer (HGT) events involving other prokaryote and eukaryote lineages. A much smaller number, up to 174 (2.1%) showed unambiguous evidence of vertical inheritance. Together with other red algal genomes, nearly all published in 2013, these data provide an excellent platform for studying diverse aspects of algal biology and evolution. This novel information will help investigators test existing hypotheses about the impact of endosymbiosis and HGT on algal evolution and enable comparative analysis within a more-refined, hypothesis-driven framework that extends beyond HGT. Here we explore the impacts of this infusion of red algal genome data on addressing questions regarding the complex nature of algal evolution and highlight the need for scalable phylogenomic approaches to handle the forthcoming deluge of sequence information.

Algal endosymbionts as vectors of horizontal gene transfer in photosynthetic eukaryotes

Photosynthesis in eukaryotes occurs in the plastid, an organelle that is derived from a single cyanobacterial primary endosymbiosis in the common ancestor of the supergroup Plantae (or Archaeplastida) that includes green, red, and glaucophyte algae and plants. However a variety of other phytoplankton such as the chlorophyll c-containing diatoms, dinoflagellates, and haptophytes contain a red alga-derived plastid that traces its origin to secondary or tertiary (eukaryote engulfs eukaryote) endosymbiosis. The hypothesis of Plantae monophyly has only recently been substantiated, however the extent and role of endosymbiotic and horizontal gene transfer (EGT and HGT) in algal genome evolution still remain to be fully understood. What is becoming clear from analysis of complete genome data is that algal gene complements can no longer be considered essentially eukaryotic in provenance; i.e., with the expected addition of several hundred cyanobacterial genes derived from EGT and a similar number derived from the mitochondrial ancestor. For example, we now know that foreign cells such as Chlamydiae and other prokaryotes have made significant contributions to plastid functions in Plantae. Perhaps more surprising is the recent finding of extensive bacterium-derived HGT in the nuclear genome of the unicellular red alga Porphyridium purpureum that does not relate to plastid functions. These non-endosymbiont gene transfers not only shaped the evolutionary history of Plantae but also were propagated via secondary endosymbiosis to a multitude of other phytoplankton. Here we discuss the idea that Plantae (in particular red algae) are one of the major players in eukaryote genome evolution by virtue of their ability to act as “sinks” and “sources” of foreign genes through HGT and endosymbiosis, respectively. This hypothesis recognizes the often under-appreciated Rhodophyta as major sources of genetic novelty among photosynthetic eukaryotes.

IDENTIFICATION OF COENZYME Q10 FROM PORPHYRIDIUM PURPUREUM (RHODOPHYTA) BY MATRIX-ASSISTED LASER DESORPTION IONIZATION CURVED FIELD REFLECTRON MASS SPECTROMETRY1.

Antioxidant agents from natural sources are currently the focus of scientific interest and are part of several natural product screenings. Coenzymes Q (CoQ, ubiquinones) are integral parts of the electron transport chain of the inner mitochondrial membrane. As antioxidants they protect phospholipids against peroxidation and are also involved in various processes of tissue protection. Their natural occurrence was validated for Saccharomyces cerevisiae as CoQ6 , for Escherichia coli as CoQ8 , and for humans as CoQ10 . After carrying out a preparative reversed-phase (RP)-HPLC separation of extracts isolated from unicellular red alga Porphyridium purpureum (Bory) K. M. Drew et R. Ross, it was possible to identify a 2,3-dimethoxy-5-methyl-6-decaprenyl-1,4-benzoquinone (CoQ10 ) within these extracts using a matrix-assisted laser desorption ionization (MALDI) curved field reflectron (CFR) mass spectrometer. Detected mass fragments showed a high significance and could be structurally interpreted for both commercialized standard and CoQ10 isolated from P. purpureum.

NITZSCHIA OVALIS (BACILLARIOPHYCEAE) MONO LAKE STRAIN ACCUMULATES 1,4/2,5 CYCLOHEXANETETROL IN RESPONSE TO INCREASED SALINITY(1).

The growth of microalgae in hypersaline conditions requires that cells accumulate osmoprotectants. In many instances, these are polyols. We isolated the diatom Nitzschia ovalis H. J. Arn. from the saline and alkaline water body Mono Lake (CA, USA). This isolate can grow in salinities ranging from 5 to 120 parts per thousand (ppt) of salt but normally at 90 ppt salinity. In this report, we identified the major polyol osmoprotectant as 1,4/2,5 cyclohexanetetrol by electron ionization-mass spectrometry (EI-MS), (1) H, (13) C nuclear magnetic resonance spectroscopy (NMR), and infrared (IR) and showed an increase in cellular concentration in response to rising salinity. This increase in the cyclitol concentration was evaluated by gas chromatography of the derived tetraacetylated cyclohexanetetrol obtaining an average of 0.7 fmol · cell(-1) at 5 ppt and rising to 22.5 fmol · cell(-1) at 120 ppt. The 1,4/2,5 cyclohexanetetrol was also detected in the red alga Porphyridium purpureum. Analysis of the free amino acid content in N.ovalis cultures exposed to changes in salinity showed that proline and lysine also accumulate with increased salinity, but the cellular concentration of these amino acids is about 10-fold lower than the concentration of 1,4/2,5 cyclohexanetetrol. The comparison of amino acid concentration per cell with cyclitol suggests that this polyol is important in compensating the cellular osmotic pressure due to increased salinity, but other physiological functions could also be considered.

Identification and characterization of a new carbonic anhydrase in the marine diatom Phaeodactylum tricornutum

A cDNA encoding a new isoenzyme of β-type carbonic anhydrase (CA; EC 4.2.1.1) in the marine diatom Phaeodactylum tricornutum Bohlin has been cloned. The cDNA contained an open reading frame of 819 bp, which encodes a polypeptide of 273 amino acids. This gene, which is designated as ptca2, was found to be highly homologous (83% at the nucleotide level) to the previously isolated intracellular β-CA gene from Phaeodactylum tricornutum (ptca1). Comparison of the deduced amino acid sequence of ptca2 with β-CAs from other sources demonstrated that PtCA2 possesses the completely conserved zinc coordination residues of β-CA. The N-terminus 19 amino acid sequence of PtCA2 was predicted to be an endoplasmic reticulum-targeting signal, suggesting localization of the protein in an organelle or in the periplasmic space. Quantitative analysis of mRNA accumulation of ptca2 using real-time polymerase chain reaction revealed a significant level of mRNA accumulation even under 5% CO2 and a 3.5-fold increase in accumulation upon acclimation of the diatom to air. This indicates that ptca2 belongs to a constitutive class of enzyme that responds only weakly to the ambient CO2 concentration. The sequences of both ptca1 and ptca2 were shown to be grouped into a phylogeny that is composed of mixture of sequences from the eucarya and procarya domains, including sequences from the red alga Porphyridium purpureum, the green alga Coccomyxa, the red mold Neurospora crassa, and the yeast Saccharomyces cerevisiae.Key words: carbonic anhydrase, marine diatom, inorganic carbon concentrating mechanism (CCM), Phaeodactylum tricornutum.

A gene homologous to beta-type carbonic anhydrase is essential for the growth of Corynebacterium glutamicum under atmospheric conditions.

Carbonic anhydrase catalyzes the interconversion of CO(2) and bicarbonate. We focused on this enzyme in the amino acid-producing organism Corynebacterium glutamicum in order to assess the availability of bicarbonate for carboxylation reactions essential to growth and for those required for L-lysine overproduction. A whole-genome sequence revealed two genes encoding putative beta-type and gamma-type carbonic anhydrases in C. glutamicum. These genes encode polypeptides containing zinc ligands strictly conserved in each type of carbonic anhydrase and were designated bca and gca, respectively. Internal deletion of the chromosomal bca gene resulted in a phenotype showing severely reduced growth under atmospheric conditions (0.04% CO(2)) on both complete and minimal media. The growth defect of the Delta bca strain was restored under elevated CO(2) conditions (5% CO(2)). Introduction of the red alga Porphyridium purpureum carbonic anhydrase gene ( pca) could compensate for the bca deletion, allowing normal growth under an atmospheric level of CO(2). In contrast, the Delta gca strain behaved identically to the wild-type strain with respect to growth, irrespective of the CO(2) conditions. Attempts to increase the dosage of bca, gca, and pca in the defined L-lysine-producing strain C. glutamicum AHD-2 led to no discernable effects on growth and production. Northern blot analysis indicated that the bca transcript in strain AHD-2 and another L-lysine producer, C. glutamicum B-6, was present at a much higher level than in the wild-type strain, particularly during exponential growth phases. These results indicate that: (1) the bca product is essential to achieving normal growth under ordinary atmospheric conditions, and this effect is most likely due to the bca product's ability to maintain favorable intracellular bicarbonate/CO(2) levels, and (2) the expression of bca is induced during exponential growth phases and also in the case of L-lysine overproduction, both of which are conditions of higher bicarbonate demand.

Porphyridium purpureum (formerly P. cruentum) contains β-carotene but no ⍺-carotene

Conflicting reports on the carotene content of the unicellular red alga Porphyridium purpureum (BORY) DREW et Ross induced us to reinvestigate the carotenoid pigments of this red algal species. But using high-performance liquid chromatography (HPLC) in combination with absorbance spectroscopy, we identified β-carotene as the only carotene in various strains of P. purpureum. These results together with the absence of β,e-xanthophylls indicate that the pathway forming β,e-carotenoids is absent in P. purpureum.

Crystal structure of E. coli beta-carbonic anhydrase, an enzyme with an unusual pH-dependent activity.

Carbonic anhydrases fall into three distinct evolutionary and structural classes: alpha, beta, and gamma. The beta-class carbonic anhydrases (beta-CAs) are widely distributed among higher plants, simple eukaryotes, eubacteria, and archaea. We have determined the crystal structure of ECCA, a beta-CA from Escherichia coli, to a resolution of 2.0 A. In agreement with the structure of the beta-CA from the chloroplast of the red alga Porphyridium purpureum, the active-site zinc in ECCA is tetrahedrally coordinated by the side chains of four conserved residues. These results confirm the observation of a unique pattern of zinc ligation in at least some beta-CAS: The absence of a water molecule in the inner coordination sphere is inconsistent with known mechanisms of CA activity. ECCA activity is highly pH-dependent in the physiological range, and its expression in yeast complements an oxygen-sensitive phenotype displayed by a beta-CA-deletion strain. The structural and biochemical characterizations of ECCA presented here and the comparisons with other beta-CA structures suggest that ECCA can adopt two distinct conformations displaying widely divergent catalytic rates.

Growth and product formation of Porphyridium purpureum

A mathematical model was developed to describe the influence ofcultivation conditions on the growth and formation of macromolecularproducts of the red alga Porphyridium purpureum. Severalmacroscopic regulation mechanisms were shown to influence productformation.The growth and formation of pigments and polysaccharides and the releaseof polysaccharides were studied in different photobioreactors under batchand continuous cultivation conditions. The light intensity tested rangedfrom the compensation point to conditions nearly saturating for growth.Temperature, gas and the medium were kept constant during theexperiments in the photobioreactors, but were varied in supplementaryexperiments in shaken flasks.A non-linear relationship was found between the biomass concentrationand the polysaccharides excreted to the medium. Light intensity did notinfluence polysaccharide formation directly, but was the main externalfactor influencing pigment composition. These two factors are consideredin a structural model of functional macromolecules to give a properdescription of the product regulation. Based on the experimental results amodel is proposed that describes the formation of functionalmacromolecules in this alga.

The adsorption kinetics of metal ions onto different microalgae and siliceous earth

In the present work the adsorption kinetics of the six metal ions aluminum, zinc, mercury, lead, copper, and cadmium onto living microalgae were measured. The freshwater green microalga Scenedesmus subspicatus, the brackish water diatom Cyclotella cryptica, the seawater diatom Phaeodactylum tricornutum, and the seawater red alga Porphyridium purpureum were the subject of investigation. In most cases the adsorption rate of the metals could be well described by using the equation of the Langmuir adsorption rate expression. Inverse parameter estimation allowed the determination of the rate constants of the adsorption process and the maximum metal content of the algae. The highest values for the rate constant were obtained for Porphyridium purpureum followed by Phaeodactylum tricornutum. High values for the maximum content were obtained for Cyclotella cryptica and Scenedesmus subspicatus. The maximum rate constant was 24.2 l h −1 for the adsorption of Hg to Porphyridium purpureum whereas the maximum metal content (0.243 g g −1) was obtained for Zn on Cyclotella cryptica. A comparison of these values with those obtained for the mineral siliceous earth exhibiting low maximum content and high adsorption rates reveals that the mechanism of adsorption onto the algae is a mixture of adsorption and accumulation.

Crystallization and preliminary X-ray diffraction studies of a beta-carbonic anhydrase from the red alga Porphyridium purpureum.

The beta-carbonic anhydrase from the red alga Porphyridium purpureum was heterologously expressed, purified and crystallized. The crystals belong to space group P2(1) (unit-cell parameters a = 63.8, b = 113.9, c = 73.8 A, beta = 104.1 degrees) with two subunits per asymmetric unit and diffract to 2.5 A resolution.

Kinetic model of extracellular polysaccharide production by the unicellular red alga Porphyridium purpureum

Kinetic model of extracellular polysaccharide production by the unicellular red alga Porphyridium purpureum

The effect of salinity, nitrate concentration, pH and temperature on eicosapentaenoic acid (EPA) production by the red unicellular alga Porphyridium purpureum

The effects of salinity, nitrate concentration, pH and temperature on eicosapentaenoic acid (EPA) production by Porphyridium purpureum were studied. Both systematic and stepwise optimization were used in order to determine the best conditions for production of EPA. An analysis method for fatty acids was also developed. The highest EPA production per cell was achieved at pH 7.6 with 41.8 g l −1 NaCl and 12.1 mM nitrate in the production medium and using a growth temperature of 8°C.

Porphyridium purpureum (Rhodophyta) from red and green light: characterization of photosystem I and determination of in situ fluorescence spectra of the photosystems

Recent investigations have shown the existence of a light-harvesting complex (LHC) of photosystem I (PSI) with a fluorescence maximum at 680 nm in the red alga Porphyridium purpureum (Wolfe et al., Nature, 367 (1994) 566–568). We have attempted to determine whether the composition of this red algal LHC is invariable or subject to light acclimatization and whether the 680–690 nm fluorescence band of intact algae, which is usually ascribed to photosystem II (PSII), could be partially due to the LHC of PSI. For this purpose, we characterized PSI core complexes and PSI holocomplexes consisting of core and LHC from algae grown under red or green light conditions and calculated the in situ spectra of the photosystems by normalizing the thylakoid spectra of red and green light algae to an identical PSI or PSII concentration. The composition of the LHC seems to be independent of the light quality, since no significant difference was found in the antenna size and the pigment composition of the PSI holocomplexes from red and green light algae; both holocomplex preparations contained 150–160 chlorophylls, 21–22 zeaxanthins and 23–25 β-carotenes per reaction centre, while the core complexes contained approximately 90 chlorophylls, 15 carotenes and no zeaxanthin. Both holocomplex preparations had a fluoroscence maximum (77 K) at 718 nm. A short-wavelength emission band, as reported by Wolfe et al., was not found in the spectra of the isolated PSI complexes or in the PSI spectrum calculated from thylakoid spectra. This indicates that the LHC does not contribute to the fluorescence spectrum of intact cells, but transfers energy very effectively to the PSI core complex.

Acclimation processes in the light harvesting complex of the red alga Porphyridium purpureum (Bory) Drew et Ross, according to irradiance and nutrient availability

ABSTRACTThe time‐course of acclimation (0‐5h) of the red alga Porphyridium purpureum with respect to total proteins, phycoerythrin (PE) and phycobilisomes (PBS) has been studied at different N availability and different light regimes. After a high N input, acclimation takes place in two phases. The first one, which is photoindependent is characterized by simultaneous increase of proteins and PE. At low N input, this first phase is not detected. In the second phase the PE content increases only under low light together with an increase of the PBS size, followed probably by an increase in the number of PBS. The effectiveness of the energy transfer increases under these conditions. A rapid decrease in the PBS size correlated with a decrease of the energy transfer is observed at high irradiance. Free PE plays an important role in the organization‐disorganization of the PBS at low N concentration (inverse correlation between free PE and PE attached to PBS). Free PE is not accumulated in the cell after a high N input at high irradiance. Independently of photoacclimation, two species of PBS appear with different PE content and different capacities to aggregate with other compounds. A clear correlation appears between the level of coupling of the PBS and the fluorescence ‘in vivo’ of the whole cells. The comparison between dissociated and undissociated PBS as well as between PBS obtained after the different acclimation processes allows the determination of the presence of two linker polypeptides probably associated with B‐PE (37 and 32–5 kDa) and two associated with PC and APC (27 and 25 kDa). That suggests that acclimation of PBS requires a parallel stoichiometric response of biliproteins and the linker polypeptides involved in the efficiency of the energy transfer.

Alterations in Fatty Acid Composition of Marine Red Alga Porphyridium purpureum by Environmental Factors

The effects of culture temperature, illumination time, aeration and concentrations of NH 4 Cl, phosphorus and NaCl in the medium on population growth and fatty acid composition of P. purpureum which exhibited the highest productivity of EPA and AA among 84 strains of marine microalgae tested were investigated. It was observed that the level of C 20:5 is high and the levels of C 20:4 and C 18 fatty acid groups are low under optimal growth conditions. The reverse phenomenon was observed under growth-limiting conditions. It was concluded that we should be able to regulate and maintain high levels of EPA and/or AA by modifying this algal environment

Fractionation of thylakoid membranes from Porphyridium purpureum using the detergent N-lauryl-?-iminodipropionate

Two green fractions, thought to represent the chlorophyll-antennae of photosystems I (PSI) and II (PSII), were isolated from the red alga Porphyridium purpureum by solubilisation of the thylakoid membranes using the detergent N-lauryl-ß-iminodipropionate and subsequent sucrose-density-gradient centrifugation. No release of pigments from the pigment-protein complexes was detected during isolation. The fractions were analyzed with respect to their chlorophyll-protein pattern, spectral properties and pigment composition. The supposed PSII antenna fraction contained both the major carotenoids of P. purpureum, β-carotene and zeaxanthin, and showed a long-wavelength absorption maximum at 672 nm and a low-temperature fluorescence maximum at 692-694 nm. Polypeptides of this fraction cross-reacted with antibodies raised against the PSII polypeptides D1, CP43 and CP47 from higher plants. The PSI fraction could perform P-700 photooxidation and showed a long-wavelength absorbance maximum at 679 nm and a low-temperature fluorescence maximum at 718 nm. It contained β-carotene as the only carotenoid. The fluorescence excitation spectrum of the fraction and measurements of the photochemical activity of a thylakoid preparation excited with light that is preferentially absorbed either by chlorophyll (433 nm) or by carotenoids (495 nm) indicate that β-carotene serves as a very efficient antenna-pigment in PSI. In contrast, only a small amount of energy transfer from the carotenoids to chlorophyll could be observed with the supposed PSII fraction.

Sustained production of arachidonic and eicosapentaenoic acids by the red alga Porphyridium purpureum cultured in a light/dark cycle

Abstract Sustained production of AA and EPA, which were accumulated at high levels without a change of fatty acid composition, was demonstrated by the red alga, Porphyridium purpureum . This alga produced 5.1 mg of AA and 5.7 mg of EPA for 1.0/culture in a one-week operation on a light/dark cycle.

Growth and ATP levels inPorphyridium purpureum(Rhodophyceae, Bangiales) cultured in the presence of surfactants

Growth and ATP levels were studied in the unicellular red alga Porphyridium purpureum cultured on a synthetic agar medium in the presence of seven different commonly used synthetic surfactants. The surfactants were used in concentrations of 0–50 ppm; the alga did not tolerate higher concentrations of the surfactants. Of the surfactants tested, three “mild” compounds (sodium desoxycholate, Synperonic NP9 (a nonyl phenol ethoxylate) and Triton X-100) did not change the normal ATP levels found in control algae (30–45 μg ATP gdw-1). In contrast, the surfactants Ufasan 65 (a linear alkylbenzene sulphonate, LAS) and Genapol ZRO (a polyglycol ether sulphate) increased the ATP levels markedly at concentrations of 30–40 ppm. A similar large increase was observed at 10 ppm of TEGO-Betain L7 (an amphoteric betaine derivative) and Ungerol N2-70 (an ethoxylated fatty alcohol sulphate). The effects of the three “mild” surfactants on the growth of P. purpureum were not linear: at 50 ppm all were growth-inhibitory, and a...