Do chromium-resistant bacterial symbionts of hyperaccumulator Callitriche cophocarpa support their host in phytobial remediation of water?

Callitriche cophocarpa Sendtn. is a macrophyte widely distributed in aquatic systems of the temperate climate zone and a known hyperaccumulator of chromium. Ten pure symbiotic bacterial isolates of C. cophocarpa were obtained and identified. Three of the isolates showed the highest resistance to Cr(VI): Microbacterium sp. (Ct1), Aeromonas sp. (Ct3) and Acinetobacter sp. (Ct6). Acinetobacter sp. (Ct6) was able to survive up to a concentration of 104 mg/L (2 mM). The isolates were also able to effectively detoxify Cr(VI) by reducing it to Cr(III). We tested whether inoculation of plants with a consortium consisting of Ct1, Ct3 and Ct6 affects: (1) the phytoextraction of chromium from leachates, (2) the physiological state of plants after Cr(VI) treatment. The solutions were landfill leachates and contained 10.7 mg/L of Cr(VI) - an amount 530 times exceeding the legal limits. We influenced the plants with Cr in two steps, each lasting for 10 days, first using mature shoots and then apical ones. The highest Cr content concomitant with the highest bioconcentration factor (BCF) were found in the inoculated plants: 1274 and 119 mg/kg dry mass (d.m.), respectively. The physiological status of the plants was assessed by biometric tests and advanced chlorophyll fluorescence analyses. The photosynthetic activity of mature shoots was influenced by Cr(VI) more negatively than that of young apical shoots. The inoculation with the bacterial consortium significantly reduced the negative effect of Cr(VI) on mature organs. In some cases the inoculated mature plants exhibited photosynthetic activity that was even higher than in the control plants. The results unequivocally show a beneficial effect of C. cophocarpa inoculation with the tested isolates resulting in a significant improvement of the phytoremediation properties of this aquatic chromium hyperaccumulator.

Structural differences of cell walls in earlywood and latewood of Pinus sylvestris and their contribution to biomass recalcitrance.

Scots pine (Pinus sylvestris L.) is an evergreen coniferous tree with wide distribution and good growth performance in a range of habitats. Therefore, wood from P. sylvestris is produced in many managed forests and is frequently used in industry. Despite the importance of pine wood, we still do not fully understand its molecular structure what limits improvements in its processing. One of the basic features leading to variation in wood properties is the presence of earlywood and latewood which form annual growth rings. Here, we characterise biochemical traits that differentiate cell walls of earlywood and latewood in Scots pine. We discover that latewood is less recalcitrant to enzymatic digestion, with galactoglucomannan showing particularly pronounced difference in accessibility. Interestingly, characterisation of lignin reveals a higher proportion of coniferaldehydes in pine latewood and suggests the presence of a different linkage landscape in this wood type. With complementary analysis of wood polysaccharides this enabled us to propose the first detailed molecular model of earlywood and latewood and to conclude that the variation in lignin structure is likely the main determinant of differences in recalcitrance observed between the two wood types in pine. Our discoveries lay the foundation for improvements in industrial processes that use pine wood since we show clear pathways for increasing the efficiency of enzymatic processing of this renewable material. Our work will help guide future breeding of pine trees with desired timber properties and can help link molecular structure of softwood cell walls to function of the different types of xylem in conifers.

Secrets of Flavonoid Synthesis in Mushroom Cells.

Flavonoids are chemical compounds that occur widely across the plant kingdom. They are considered valuable food additives with pro-health properties, and their sources have also been identified in other kingdoms. Especially interesting is the ability of edible mushrooms to synthesize flavonoids. Mushrooms are usually defined as a group of fungal species capable of producing macroscopic fruiting bodies, and there are many articles considering the content of flavonoids in this group of fungi. Whereas the synthesis of flavonoids was revealed in mycelial cells, the ability of mushroom fruiting bodies to produce flavonoids does not seem to be clearly resolved. This article, as an overview of the latest key scientific findings on flavonoids in mushrooms, outlines and organizes the current state of knowledge on the ability of mushroom fruiting bodies to synthesize this important group of compounds for vital processes. Putting the puzzle of the current state of knowledge on flavonoid biosynthesis in mushroom cells together, we propose a universal scheme of studies to unambiguously decide whether the fruiting bodies of individual mushrooms are capable of synthesizing flavonoids.

Is the merA gene sufficient as a molecular marker of mercury bacterial resistance?

Gene encoding mercuric ion reductase, merA is a crucial component of the mer operon for reduction of nonorganic mercury ions into less toxic form. The merA gene or its fragments are commonly used as a molecular marker of bacterial resistance to mercury. In this study, it was tested whether the merA gene can be considered as a molecular marker of mercury bacterial resistance. For this purpose, the presence of the mer operon in bacteria isolated from the microbiota of Tussilago farfara L. growing in post-industrial mercury-contaminated and non-contaminated areas was verified by merA gene identification. Mercury resistance was determined by analyzing the bacterial growth parameters in standard Luria-Bertani (LB) medium with mercury concentration of 0.01% (w/v) and in medium without mercury addition. The results obtained showed that the merA gene was present in all T. farfara L. bacterial isolates growing in both mercury-contaminated and noncontaminated soils, however, only the isolates from mercury-contaminated areas were able to grow under mercury conditions. Although merA is commonly regarded as a molecular marker of bacterial mercury resistance, results of our research indicate the need for a verification of that statement/thesis and further investigation of bacterial mercury resistance to indicate other its key markers, structures, or mechanisms.

Brassinosteroid-lipid membrane interaction under low and high temperature stress in model systems.

BACKGROUND: In earlier studies [1], we indicated that applying brassinosteroids (BRs) to lipids that had been isolated from plants altered the physicochemical properties of the monolayers. A continuation of these dependencies using the defined model lipid systems is presented in this paper. The influence of homocastasterone (HCS) and castasterone (CS) (BRs for which the increase in concentration were characteristic of plants grown at low temperatures) on the membrane properties of their polar and the hydrophobic parts were studied. RESULTS: Changes in the electrokinetic potential indicate that both BRs decreased the negative charge of the surface, which is an important factor in modifying the contacts with the polar substances. This property of BRs has not yet been described. The studies of the interactions that occur in the hydrophobic part of the membrane were investigated using the EPR methods and Langmuir techniques. The physicochemical parameters of the lipid structure were determined, and the excess of Gibbs free energy was calculated. CONCLUSION: We conclude that examined BRs modify both the hydrophilic and hydrophobic properties of the membranes, but to a greater extent HCS. The consequence of these changes may be the attempt to maintain the stability of the membranes in stressful temperature conditions and / or to the possibility of adsorption of other substances on membranes surfaces. The change of plant metabolism towards increasing the amount of BR, mainly HCS (under cooling) may by an important factor for maintaining optimal structural properties of membranes and their functionality despite temperature changes.

ROS-Scavengers, Osmoprotectants and Violaxanthin De-Epoxidation in Salt-Stressed Arabidopsis thaliana with Different Tocopherol Composition.

To determine the role of α- and γ-tocopherol (TC), this study compared the response to salt stress (200 mM NaCl) in wild type (WT) Arabidopsis thaliana (L.) Heynh. And its two mutants: (1) totally TC-deficient vte1; (2) vte4 accumulating γ-TC instead of α-TC; and (3) tmt transgenic line overaccumulating α-TC. Raman spectra revealed that salt-exposed α-TC accumulating plants were more flexible in regulating chlorophyll, carotenoid and polysaccharide levels than TC deficient mutants, while the plants overaccumulating γ-TC had the lowest levels of these biocompounds. Tocopherol composition and NaCl concentration affected xanthophyll cycle by changing the rate of violaxanthin de-epoxidation and zeaxanthin formation. NaCl treated plants with altered TC composition accumulated less oligosaccharides than WT plants. α-TC deficient plants increased their oligosaccharide levels and reduced maltose amount, while excessive accumulation of α-TC corresponded with enhanced amounts of maltose. Salt-stressed TC-deficient mutants and tmt transgenic line exhibited greater proline levels than WT plants, lower chlorogenic acid levels, and lower activity of catalase and peroxidases. α-TC accumulating plants produced more methylated proline- and glycine- betaines, and showed greater activity of superoxide dismutase than γ-TC deficient plants. Under salt stress, α-TC demonstrated a stronger regulatory effect on carbon- and nitrogen-related metabolites reorganization and modulation of antioxidant patterns than γ-TC. This suggested different links of α- and γ-TCs with various metabolic pathways via various functions and metabolic loops.

Antioxidant and Signaling Role of Plastid-Derived Isoprenoid Quinones and Chromanols.

Plant prenyllipids, especially isoprenoid chromanols and quinols, are very efficient low-molecular-weight lipophilic antioxidants, protecting membranes and storage lipids from reactive oxygen species (ROS). ROS are byproducts of aerobic metabolism that can damage cell components, they are also known to play a role in signaling. Plants are particularly prone to oxidative damage because oxygenic photosynthesis results in O2 formation in their green tissues. In addition, the photosynthetic electron transfer chain is an important source of ROS. Therefore, chloroplasts are the main site of ROS generation in plant cells during the light reactions of photosynthesis, and plastidic antioxidants are crucial to prevent oxidative stress, which occurs when plants are exposed to various types of stress factors, both biotic and abiotic. The increase in antioxidant content during stress acclimation is a common phenomenon. In the present review, we describe the mechanisms of ROS (singlet oxygen, superoxide, hydrogen peroxide and hydroxyl radical) production in chloroplasts in general and during exposure to abiotic stress factors, such as high light, low temperature, drought and salinity. We highlight the dual role of their presence: negative (i.e., lipid peroxidation, pigment and protein oxidation) and positive (i.e., contribution in redox-based physiological processes). Then we provide a summary of current knowledge concerning plastidic prenyllipid antioxidants belonging to isoprenoid chromanols and quinols, as well as their structure, occurrence, biosynthesis and function both in ROS detoxification and signaling.

Unique biocenosis as a foundation to develop a phytobial consortium for effective bioremediation of Cr(VI)-polluted waters and sediments.

This paper analyzes a unique, aquatic phytobial biocenosis that has been forming naturally for over 20 years and operating as a filter for Cr(VI)-polluted groundwater. Our study presents a thorough taxonomic analysis of the biocenosis, including filamentous algae, vascular plants, and microbiome, together with the analysis of Cr accumulation levels, bioconcentration factors and other environmentally-significant parameters: siderophore production by bacteria, biomass growth of the plants or winter hardiness. Among 67 species identified in the investigated reservoir, 13 species were indicated as particularly useful in the bioremediation of Cr(VI)-polluted water and sediment. Moreover, three species of filamentous algae, Tribonema sp., and three easily culturable bacterial species were for the first time shown as resistant to Cr concentration up to 123 mg/dm3, i.e. 6150 times over the permissible level. The work presents a modern holistic phytobial consortium indispensable for the remediation of Cr(VI)-contaminated aquatic environment in temperate zones worldwide.

Detection of a pheomelanin-like pigment by EPR spectroscopy in the mycelium of Plenodomus biglobosus.

Melanin occurrence in Plenodomus biglobosus was investigated using electron paramagnetic (spin) resonance (EPR, ESR) spectroscopy. The fungus was isolated from living and dead leaves of European ash (Fraxinus excelsior L.). Dark pigmentation of P. biglobosus mycelium in vitro, especially on the reverse, was observed. The black coloration intensified with the age of the culture and inspired us to check if the analyzed fungus species synthesizes melanin. Melanin contains unpaired electrons, thus, EPR spectroscopy was applied, as a specific technique, to verify its presence in P. biglobosus. The EPR spectrum of the mycelium showed a very strong melanin signal, revealing pheomelanin-like features. Thus, the black pigment of P. biglobosus was clearly identified as melanin. However, no melanin was detected in the apparently dark culture medium even when zinc (II) acetate was added to increase the sensitivity of detection. Pheomelanin has many unusual biological functions but it is not commonly found in fungi. Detection of this type of melanin in P. biglobosus, which can be both endophytic or pathogenic, suggests a closer examination of the potential role of this melanin in host-parasite interaction.

Lipid Dependence of Xanthophyll Cycling in Higher Plants and Algae.

The xanthophyll cycles of higher plants and algae represent an important photoprotection mechanism. Two main xanthophyll cycles are known, the violaxanthin cycle of higher plants, green and brown algae and the diadinoxanthin cycle of Bacillariophyceae, Xanthophyceae, Haptophyceae, and Dinophyceae. The forward reaction of the xanthophyll cycles consists of the enzymatic de-epoxidation of violaxanthin to antheraxanthin and zeaxanthin or diadinoxanthin to diatoxanthin during periods of high light illumination. It is catalyzed by the enzymes violaxanthin or diadinoxanthin de-epoxidase. During low light or darkness the back reaction of the cycle, which is catalyzed by the enzymes zeaxanthin or diatoxanthin epoxidase, restores the epoxidized xanthophylls by a re-introduction of the epoxy groups. The de-epoxidation reaction takes place in the lipid phase of the thylakoid membrane and thus, depends on the nature, three dimensional structure and function of the thylakoid lipids. As the xanthophyll cycle pigments are usually associated with the photosynthetic light-harvesting proteins, structural re-arrangements of the proteins and changes in the protein-lipid interactions play an additional role for the operation of the xanthophyll cycles. In the present review we give an introduction to the lipid and fatty acid composition of thylakoid membranes of higher plants and algae. We introduce the readers to the reaction sequences, enzymes and function of the different xanthophyll cycles. The main focus of the review lies on the lipid dependence of xanthophyll cycling. We summarize the current knowledge about the role of lipids in the solubilization of xanthophyll cycle pigments. We address the importance of the three-dimensional lipid structures for the enzymatic xanthophyll conversion, with a special focus on non-bilayer lipid phases which are formed by the main thylakoid membrane lipid monogalactosyldiacylglycerol. We additionally describe how lipids and light-harvesting complexes interact in the thylakoid membrane and how these interactions can affect the structure of the thylakoids. In a dedicated chapter we offer a short overview of current membrane models, including the concept of membrane domains. We then use these concepts to present a model of the operative xanthophyll cycle as a transient thylakoid membrane domain which is formed during high light illumination of plants or algal cells.

Molecular Dynamics of Chloroplast Membranes Isolated from Wild-Type Barley and a Brassinosteroid-Deficient Mutant Acclimated to Low and High Temperatures.

Plants have developed various acclimation strategies in order to counteract the negative effects of abiotic stresses (including temperature stress), and biological membranes are important elements in these strategies. Brassinosteroids (BR) are plant steroid hormones that regulate plant growth and development and modulate their reaction against many environmental stresses including temperature stress, but their role in modifying the properties of the biological membrane is poorly known. In this paper, we characterise the molecular dynamics of chloroplast membranes that had been isolated from wild-type and a BR-deficient barley mutant that had been acclimated to low and high temperatures in order to enrich the knowledge about the role of BR as regulators of the dynamics of the photosynthetic membranes. The molecular dynamics of the membranes was investigated using electron paramagnetic resonance (EPR) spectroscopy in both a hydrophilic and hydrophobic area of the membranes. The content of BR was determined, and other important membrane components that affect their molecular dynamics such as chlorophylls, carotenoids and fatty acids in these membranes were also determined. The chloroplast membranes of the BR-mutant had a higher degree of rigidification than the membranes of the wild type. In the hydrophilic area, the most visible differences were observed in plants that had been grown at 20 °C, whereas in the hydrophobic core, they were visible at both 20 and 5 °C. There were no differences in the molecular dynamics of the studied membranes in the chloroplast membranes that had been isolated from plants that had been grown at 27 °C. The role of BR in regulating the molecular dynamics of the photosynthetic membranes will be discussed against the background of an analysis of the photosynthetic pigments and fatty acid composition in the chloroplasts.

Selected metal content and antioxidant capacity of Sambucus nigra flowers from the urban areas versus soil parameters and traffic intensity.

The metal accumulation and antioxidant contents in flowers of wild specimens of European elder (Sambucus nigra L.), a famous medicinal plant and valuable component in the urban landscape, were determined. The total reflection X-ray fluorescence revealed the presence of K, Rb, Ca, Sr, Cr, Mn, Fe, Cu, and Zn associated with flowers. A typical, large, non-industrial city with considerable traffic and atmospheric pollution resulting from smog was chosen as a place of sampling. Obtained results were correlated with selected parameters of soil and the intensity of surrounding traffic. The flowers were relatively rich in elements K, Ca, Cu, Rb, and antioxidants, while it did not accumulate heavy metals potentially bioavailable in the soil. The correlation between street traffic and the content of Fe, Cr, and Zn in elderflowers was revealed; the metal quantities were below levels harmful to humans. Flowers from the city center exhibited higher antioxidant and radical scavenging capacities comparing to plants from the areas of little traffic. The antioxidant parameters were negatively correlated with the silty fraction content and positively with the potentially bioavailable levels of Ti and Mn in soils and increased with the amount of Rb in the flowers. It was proven for the first time that the urban specimens of wild S. nigra can perform as a local source of beneficial flowers providing cost-effective support in disease prevention and treatment.

Violaxanthin conversion by recombinant diatom and plant de-epoxidases, expressed in Escherichia coli - comparative analysis.

The purpose of this research was to obtain recombinant violaxanthin de-epoxidases (VDEs) from two species. The first one was VDE of Arabidopsis thaliana (L.) Heynh. (WT Columbia strain) (AtVDE) which in vivo catalyzes conversion of violaxanthin (Vx) to zeaxanthin (Zx) via anteraxanthin (Ax). The second one was VDE of Phaeodactylum tricornutum Bohlin, 1897 (CCAP 1055/1 strain) (PtVDE) which is responsible for de-epoxidation of diadinoxanthin (Ddx) to diatoxanthin (Dtx). As the first step of our experiments, open reading frames coding for studied enzymes were amplified and subsequently cloned into pET-15b plasmid. For recombinant proteins production Escherichia coli Origami b strain was used. The molecular weight of the produced enzymes were estimated approximately at 45kDa and 50kDa for AtVDE and PtVDE, respectively. Both enzymes, purified under native conditions by immobilized metal affinity chromatography, displayed comparable activity in assay mixture and converted up to 90% Vx in 10 min in two steps enzymatic de-epoxidation, irrespective of enzyme origin. No statistically significant differences were observed when kinetics of the reactions catalyzed by these enzymes were compared. Putative role of selected amino-acid residues of AtVDE and PtVDE was also considered. The significance of the first time obtained recombinant PtVDE as a useful tool in various comparative investigations of de-epoxidation reactions in main types of xanthophyll cycles existing in nature are also indicated.

Diadinoxanthin de-epoxidation as important factor in the short-term stabilization of diatom photosynthetic membranes exposed to different temperatures.

The importance of diadinoxanthin (Ddx) de-epoxidation in the short-term modulation of the temperature effect on photosynthetic membranes of the diatom Phaeodactylum tricornutum was demonstrated by electron paramagnetic resonance (EPR), Laurdan fluorescence spectroscopy, and high-performance liquid chromatography. The 5-SASL spin probe employed for the EPR measurements and Laurdan provided information about the membrane area close to the polar head groups of the membrane lipids, whereas with the 16-SASL spin probe, the hydrophobic core, where the fatty acid residues are located, was probed. The obtained results indicate that Ddx de-epoxidation induces a two component mechanism in the short-term regulation of the membrane fluidity of diatom thylakoids during changing temperatures. One component has been termed the "dynamic effect" and the second the "stable effect" of Ddx de-epoxidation. The "dynamic effect" includes changes of the membrane during the time course of de-epoxidation whereas the "stable effect" is based on the rigidifying properties of Dtx. The combination of both effects results in a temporary increase of the rigidity of both peripheral and internal parts of the membrane whereas the persistent increase of the rigidity of the hydrophobic core of the membrane is solely based on the "stable effect."

Influence of thylakoid membrane lipids on the structure of aggregated light-harvesting complexes of the diatom Thalassiosira pseudonana and the green alga Mantoniella squamata.

The study investigated the effect of the thylakoid membrane lipids monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), sulphoquinovosyldiacylglycerol (SQDG) and phosphatidylglycerol (PG) on the structure of two algal light-harvesting complexes (LHCs). In contrast to higher plants whose thylakoid membranes are characterized by an enrichment of the neutral galactolipids MGDG and DGDG, both the green alga Mantoniella squamata and the centric diatom Thalassiosira pseudonana contain membranes with a high content of the negatively charged lipids SQDG and PG. The algal thylakoids do not show the typical grana-stroma differentiation of higher plants but a regular arrangement. To analyze the effect of the membrane lipids, the fucoxanthin chlorophyll protein (FCP) complex of T. pseudonana and the LHC of M. squamata (MLHC) were prepared by successive cation precipitation using Triton X-100 as detergent. With this method, it is possible to isolate LHCs with a reduced amount of associated lipids in an aggregated state. The results from 77 K fluorescence and photon correlation spectroscopy show that neither the neutral galactolipids nor the negatively charged lipids are able to significantly alter the aggregation state of the FCP or the MLHC. This is in contrast to higher plants where SQDG and PG lead to a strong disaggregation of the LHCII whereas MGDG and DGDG induce the formation of large macroaggregates. The results indicate that LHCs which are integrated into thylakoid membranes with a high amount of negatively charged lipids and a regular arrangement are less sensitive to lipid-induced structural alterations than their counterparts in membranes enriched in neutral lipids with a grana-stroma differentiation.

Column chromatography as a useful step in purification of diatom pigments.

Fucoxanthin, diadinoxanthin and diatoxanthin are carotenoids found in brown algae and most other heterokonts. These pigments are involved in photosynthetic and photoprotective reactions, and they have many potential health benefits. They can be extracted from diatom Phaeodactylum tricornutum by sonication, extraction with chloroform : methanol and preparative thin layer chromatography. We assessed the utility of an additional column chromatography step in purification of these pigments. This novel addition to the isolation protocol increased the purity of fucoxanthin and allowed for concentration of diadinoxanthin and diatoxanthin before HPLC separation. The enhanced protocol is useful for obtaining high purity pigments for biochemical studies.

Achromobacter xylosoxidans as a new microorganism strain colonizing high-density polyethylene as a key step to its biodegradation.

This study presents results of research on isolation new bacteria strain Achromobacter xylosoxidans able to effect on the structure of high-density polyethylene (HDPE), polymer resistant to degradation in environment. New strain of A. xylosoxidans PE-1 was isolated from the soil and identified by analysis of the 16S ribosome subunit coding sequences. The substance to be degraded was HDPE in the form of thin foil films. The foil samples were analyzed with Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy (ATR-FTIR) as well as scanning electron microscope (SEM), and the results revealed degradation of chemical structure of HDPE. About 9 % loss of weight was also detected as a result of A. xylosoxidans PE-1 effect on HDPE foil. On the basis of comparative spectral analysis of the raw material before the bacteria treatment and the spectrum from a spectra database, it was assumed that the HDPE was the only source of carbon and energy for the microorganisms. No fillers or other additives used in the plastic processing were observed in HDPE before experiments. This is the first communication showing that A. xylosoxidans is able to modify chemical structure of HDPE, what was observed both on FTIR, in mass reduction of HDPE and SEM analysis. We also observed quite good growth of the bacteria also when the HDPE was the sole carbon source in the medium. These results prove that A. xylosoxidans is an organism worth applying in future HDPE biodegradation studies.

Pb remobilization by bacterially mediated dissolution of pyromorphite Pb5(PO4)3Cl in presence of phosphate-solubilizing Pseudomonas putida.

Remediation of lead (Pb)-contaminated sites with phosphate amendments is one of the best studied and cost-effective methods for in situ immobilization. In this treatment, a very stable mineral, pyromorphite Pb5(PO4)3Cl, is formed. Several studies propose to improve this treatment method with the addition of phosphate-solubilizing bacteria (PSB). The effect of bacteria on solubilization of pyromorphite is unknown. In this study, the effect of the soil microorganisms on the stability of pyromorphite Pb5(PO4)3Cl has been investigated in a set of batch solution experiments. The mineral was reacted with Pseudomonas putida, a common soil microorganism. Dissolution of pyromorphite was enhanced by the presence of P. putida, resulting in an elevated Pb concentration in the solution. This occurred even when the bacteria were provided with an additional source of phosphate in the solution. Pyromorphite has been shown to be a potential source of nutrient phosphorus for common soil bacteria. Thus, the use of PSB in remediation treatments of Pb contaminated sites may have adverse long-term impacts on Pb immobilization. Conscious phosphate management is suggested for long-term sustainability of the in situ Pb immobilization by pyromorphite formation.

Expression of three diadinoxanthin de-epoxidase genes of Phaeodacylum tricornutum in Escherichia coli Origami b and BL21 strain.

In the diadinoxanthin cycle the epoxy group is removed from diadinoxanthin and diatoxanthin is created. This conversion takes place e.g. in diatoms with the involvement of the enzyme diadinoxanthin de-epoxidase. In one of the diatom species, Phaeodactylum tricornutum (CCAP 1055/1 strain with genome sequenced) three de-epoxidase genes (PtVDE, PtVDL1, PtVDL2) have been identified, but only one of them (PtVDE) corresponds to violaxanthin de-epoxidase, an enzyme which is commonly found in higher plants. In these studies, the expression of two de-epoxidase genes of another Phaeodactylum tricornutum strain (UTEX 646), which is commonly used in diatom studies, were obtained in Origami b and BL21 E. coli strains. The molecular masses of the mature proteins are about 49 kDa and 60 kDa, respectively, for VDE and VDL2. Both enzymes are active with violaxanthin as a substrate.