Unicellular freshwater cyanobacterium Synechococcus elongatus PCC 7942 is the model organism for studying the circadian clock in cyanobacteria. Despite tremendous work over the last decade in identification of clock-related loci and elucidation of molecular mechanisms of the central oscillator, many details of the basic steps in generating circadian rhythms of biological processes remain unsolved and many components are still missing. A transposon-mediated mutagenesis and sequencing strategy has been adopted to disrupt essentially every locus in the genome so as to identify all of the loci that are involved in clock function. The complete genome sequence has been determined by a combination of shotgun sequences and transposon-mediated sequences. The S. elongatus PCC 7942 genome is 2,695,903 bp in length, and has a 55.5% GC content. Automated annotation identified 2,856 protein-coding genes and 51 RNA coding loci. A system for community refinement of the annotation was established. Organization and characteristic features of the genome are discussed in this dissertation. More than 95% of the PCC 7942 genome has been mutagenized and mutants affected in approximately 30% of loci have been screened for defects in circadian function. Approximately 70 new clock loci that belong to different functional categories have been discovered through a team effort. Additionally, functional analysis of insertion mutants revealed that the Type-IV pilus assembly protein PilN and the RNA chaperon Hfq are involved in transformation competence of S. elongatus cells. Functional analysis of an atypical short period kaiA insertional mutant showed that the short period phenotype is caused mainly by the truncation of KaiA by three amino acid residues. The interaction between KaiC and the truncated KaiA is weakened as shown by fluorescence anisotropy analysis. Deletion analysis of pANL, the large endogenous plasmid, implies that two toxin-antitoxin cassettes were responsible for inability to cure cells of this plasmid. In summary, the results indicate that this functional genomics project is very promising toward fulfilling our goal to assemble a comprehensive view of the cyanobacterial circadian clock. The mutagenesis reagents and dataset generated in this project will also benefit the greater scientific community.

Abstract This research investigated the spatial and temporal distributions of Trichodesmium spp. and Richelia intracellularis in the South China Sea. The surveys covered the period from July 2000 to July 2002. A total of eight cruises, including spring, summer and fall were conducted. The sampling stations located between 18~22o N and 115~122 o E , over the continental shelf, the slope, and the basin of the northern South China Sea. Trichodesmium biomass was higher in summer and fall than spring. There was no significant difference in biomass among shelf , slope and basin. The averaged biomass was 69

This research investigated the factors influencing the structure of the phytoplanktori community in Chaffey Dam, which is located in sub-tropical Australia. In particular, the research aimed to determine the influence of light at time scales ranging from seconds to seasons, on the growth and photophysiology of the cyanobacterium Anabaena circinalis. On a large scale, field monitoring programs between 1987 and 1997 indicated that the phytoplankton community of Chaffey Dam was dominated by colonial or relatively large phytoplankton that move either with the aid of flagella or can be positively buoyant. Diatoms contributed only a minor component, which may be the result of the reservoir being stratified for much of the year. Several of the dominant taxa bloomed in each of the seasons during the eleven year period, with some blooms lasting >9 months, indicating that environmental variability between seasons can be low. In contrast to other studies, A. circinalis was more likely to grow and bloom during the cooler months (March-October). A two-year intensive monitoring program (1995-1997) identified a seasonal progression that was similar in both years. Chlorophytes occurred in spring, Ceratium in mid summer, a relatively clear period in February, A. circinalis in March and cryptomonads in winter. On a smaller scale, short-term (2-3 day) in-situ and laboratory enclosure experiments found that the light and nutrient requirements of the dominant taxa varied. In comparison to most other phytoplankton, A. circinalis cells disappeared at very rapid rates when supplied irradiances <10 (umol photons m-2 s-1. Over several days of darkness, the filaments broke apart and the cell numbers declined. The experiments also showed that at certain times, field populations of A. circinalis were subject to high losses at all irradiances. Laboratory studies investigating the influence of inter- and intra-daily changes in light availability showed that the growth rate of A. circinalis was not affected by the frequency of daytime light:dark cycles, indicating that the rate of water mixing will not have major influence on its growth if the total daily light dose is maintained. It was also found that A. circinalis cultures did not accumulate large reserves of energy in the form of carbohydrate, other than that required for one night. This strategy may enable the colonies to have a high level of buoyancy each morning so that they float quickly to the surface waters and obtain sufficient light each day to minimise losses. However, this strategy limits the ability of A. circinalis to grow and maintain vital cell processes during extended periods of low irradiances and may be a factor causing them to be susceptible to cell breakdown. Weekly measurements of algal growth rates in Chaffey Dam identified two factors that may have acted singly or simultaneously to influence the development of A. circinalis blooms during 1996 and 1997. The blooms developed during a 4-6 week period when the mean irradiance in the surface mixed layer (SML) was sufficient to prevent high losses. Secondly, the blooms developed when soluble phosphorus in the epilimnion was relatively high but soluble nitrogen was low. This may have favoured A. circinalis, which has the potential to fix atmospheric nitrogen. The decline of A. circinalis blooms was correlated with a deepening of the SML and a reduction of the mean daytime irradiance within the SML. Their decline did not appear to be related to nutrient limitation or to changes in zooplankton concentrations. This research also developed a physiological technique for tracking daily changes in the mean daytime irradiance of A. circinalis and for estimating cell growth rate. This method is based on chlorophyll-a fluorescence quenching analysis of the state transition mechanism, which regulates light availability between the photosystems. The mean daytime irradiance of A. circinalis showed a strong relationship with the degree of non-photochemical quenching (qn), whereas the relative change to the maximum fluorescence showed a strong relationship with cell growth. It is anticipated that this method will provide a useful research tool for determining the relative importance of light and other factors on the net growth of A. circinalis and other cyanobacteria.

Many multicellular cyanobacteria produce specialized nitrogenfixing heterocysts. During diazotrophic growth of Anabaena (Nostoc) sp. strain PCC 7120, a regulated developmental pattern of single heterocysts separated by about 10 to 20 photosynthetic vegetative cells is maintained along filaments. Heterocyst structure and metabolic activity function together to accommodate oxygensensitive nitrogen fixation, catalyzed by nitrogenase. In this work, we show that the promoter of the nifHDK genes that encode nitrogenase, lies upstream from the intergenic region between nifH and nifU. Excision of the fdxN element is required for transcription of the nifHDK genes. Fluorescence microscopy of reporter strain PnifHDgfp, in the chromosomal nif locus indicated that expression of nifHDK is blocked in mutants that are unable to excise the fdxN element after nitrogen deprivation. We proposed that a promoter upstream of the element, likely PnifB, is required for transcription of the nifHDK genes. Indeed, the PnifHDgfp reporter at an ectopic site did not show GFP fluorescence. A PnifBgfp reporter was expressed specifically in heterocysts indicating that a promoter for the nifB gene lies in the intergenic region upstream of nifB. A stem loop structure located in the intergenic region between nifH and nifU may act as a processing site for production of nifHDK transcripts. We also provide evidence that DevH, a transcriptional regulator, is involved in regulating the nifBfdxNnifSUHDK genes. DevH is a protein belonging to the cAMP receptor protein (CRP) family of proteins that are widespread in bacteria and regulate genes in response to a gamut of physiological conditions. We show that DevH binds specifically to the nifB upstream region but not to the immediate upstream region of nifH. We predict that DevH binds to an NtcAlike binding site upstream of nifB and functions as an activator of the nifBfdxNnifSUHDK genes. Finally, we show that sigE, which is expressed at 16 hours after nitrogen deprivation, is required for normal expression of some heterocyst specific genes, including nifHDK. A sigE mutant shows delayed and reduced expression of nifHDK and some middle and late genes. We hypothesize that DevH in concert with SigE upregulates the expression of nifHDK in heterocysts after nitrogen deprivation.

No description available.

La communication intercellulaire se produit non seulement chez les eucaryotes, mais aussi chez certaines bactéries. Un tel exemple est la cyanobactérie filamenteuse Anabaena sp. PCC 7120, capable de former des hétérocystes suite à une carence en azote combiné. Un filament d'Anabaena est coordonné comme une unité multicellulaire; comment les cellules communiquent-elles le long de chaque filament et comment échangent-elles des ressources nutritionnelles demeurent des mécanismes encore mal élucidés. Des études récentes ont démontré que des molécules de petites tailles peuvent être échangées entre les cytoplasmes à travers des jonctions intercellulaires. De plus, le périplasme semble être continu le long de chaque filament, avec une membrane extérieure commune pour toutes les cellules. Toutefois, il n’est pas déterminé si le "périplasme continu" peut servir comme une route alternative pour les échanges moléculaires le long des filaments.Dans cette étude, la propriété du périplasme chez Anabaena a été évaluée par le suivi du mouvement de protéines fluorescentes (GFP ou iLOV) en utilisant des techniques microscopiques. Les protéines fluorescentes ont été exportées vers l'espace périplasmique, soit d'un hétérocyste soit d’une cellule végétative. Nous avons pu montrer que ces protéines fluorescentes restent dans le périplasme de la cellule d’origine, et que la GFP peut diffuser librement, mais seulement dans le périplasme d'un hétérocyste ou d’une cellule végétative. Ainsi, bien que le périplasme semble être continu le long du filament, une barrière intercellulaire semble exister pour empêcher la libre diffusion des protéines à la taille de ~27 kDa (GFP) ou ~13 kDa (iLOV). La couche de peptidoglycane pourrait constituer cette barrière et nous estimons que la limite pour la diffusion à travers cette barrière se situe entre 0.53 et 13 kDa.En parallèle, les voies métaboliques des cellules végétatives et des hétérocystes ont été comparées en utilisant une approche transcriptomique. L'expression différentielle des gènes impliqués dans le métabolisme nous permet d’appréhender la nature des métabolites pouvant être échangées entres ces deux types cellulaires. / Cell-cell communication occurs not only in eukaryotes but also in bacteria. One such example is the filamentous cyanobacterium Anabaena sp. PCC 7120, which is able to differentiate a specialized cell type named heterocyst upon nitrogen deprivation. A filament of Anabaena is coordinated as a multicellular unity; how the cells along each filament communicate and exchange resources are not yet fully understood. Recent studies demonstrated that small molecules can be rapidly exchanged from cytoplasm to cytoplasm through intercellular junctions. In addition, the periplasm appears to be continuous along each filament, with a shared outer membrane for all cells. However, whether the ‘continuous periplasm’ serves as an alternative route for molecular exchanges along the filament remains unknown. In this study, the property of periplasm in Anabaena was assessed by monitoring the movement of fluorescent proteins (GFP or iLOV) using microscopic techniques. Fluorescent proteins were exported to the periplasmic space of either a heterocyst or a vegetative cell and their diffusion was tested. We found that both GFP and iLOV remains in the producing cells, and at least GFP could diffuse freely in the periplasm of a heterocyst or a vegetative cell but failed to cross cell borders. Thus although periplasm appears to be continuous along the filament, barriers exist to prevent free diffusion of proteins up to the size of ~27 kDa (GFP) or ~13 kDa (iLOV). One candidate as diffusion barrier in the periplasm may be the peptidoglycan and we estimate the limit for diffusion of the barrier in the range between 0.53 to 13 kDa. In parallel, the biosynthetic pathways operating in vegetative cells and heterocysts were compared using oligonucleotide microarray. Differential expression of the genes involved in amino acids metabolism give clues as to which nitrogen-containing compounds might serve as the transfer vehicle in cell-cell exchanges.

The linear peptide bisebromoamide was isolated by the Suenaga group in 2009 from the marine cyanobacterium Lyngbya sp. It exhibits antiproliferative activity at nanomolar levels against a wide range of cell lines. Current SAR data indicates that there is some flexibility in the structure with respect to stereochemistry, but the range of modifications that have been biologically tested is limited, as reviewed in Chapter 1. Bisebromoamide contains a number of non-commercial amino acids and an oxopropyl pyrrolidine moiety which had not been found in a natural product previously. Several new synthetic routes towards the non-commercial amino acid fragments have been developed, as described in Chapter 2, including two ring-closure-based approaches to the substituted proline derivative 4-methyl proline (4-MePro). While the presence of six amide bonds makes solid phase peptide synthesis (SPPS) an appealing approach to the synthesis of bisebromoamide, the 4-MePro moiety is attached to a thiazoline and it is well documented that the α-position of an amino acid will racemise, under both acidic and basic conditions, when attached to a thiazoline or oxazoline. Previous reports indicated that the methyl group of the thiazoline was not essential for biological activity and so to increase stability it was replaced with a thiazole. The total synthesis of a series of novel bisebromoamide analogues, via an SPPS approach which enables facile modification of the final structure, is described in Chapter 3. The simple and adaptable SPPS route developed lends itself to SAR studies and allows modifications such as an alanine scan, truncations and incorporation of modified proline derivatives to be achieved rapidly. The promising anticancer activity of bisebromoamide makes the biological activity of these analogues of particular interest and the results of current biological testing are reported in Chapter 4.

No ambiente aquático a maior fonte de nitrogênio encontra-se na forma de nitrato que necessita sofrer uma redução para formação de compostos biologicamente aproveitáveis, tais como aminoácidos, bases nitrogenadas e compostos nitrogenados. A assimilação de nitrogênio é um processo que ocorre em duas etapas catalisadas seqüencialmente pelas enzimas nitrato redutase (NR) e nitrito redutase (NiR). A NR catalisa a redução do nitrato a nitrito (etapa considerada limitante na assimilação de nitrogênio), sendo este posteriormente reduzido a amônio pela NiR. NR está amplamente distribuída e encontrada em diferentes organismos, incluíndo bactérias, fungos, cianobactérias, plantas terrestres e algas. Neste trabalho estudamos a NR de Microcystis aeruginosa que é uma cianobactéria tóxica de água doce encontrada principalmente em reservatórios de água. A toxina microcistina quando liberada por esta microalga está associada com problemas de saúde em humanos e animais. Foi mostrado que a NR de M. aeruginosa pertence a classe das NRs biespecíficas para NADH e NADPH. Apresenta constante de Michaelis-Menten aparente (Km) de 1,5 e 1,6 mM para NADPH e NADH, respectivamente. Ainda, Km aparente para nitrato foi estimado em 0,6 mM. As condições ótimas de ensaio encontradas foram em pH 10,0 e temperatura em 40ºC. A exposição da M. aeruginosa ao herbicida oxifluorfeno (10 &#181;g/L) promoveu a inibição de NiR, possibilitando a quantificação de &#8226;NO formado via NR, enzima que teve sua atividade 6 vezes maior durante a exposição a este agente. O estudo da enzima NR é de fundamental importância para a compreensão da regulação da expressão de enzimas assimiladoras de nitrogênio bem como dos mecanismos de nutrição e crescimento desta microalga. / Nitrate is the major source of nitrogen in the aquatic environment, which must be reduced before incorporation into biological compounds, such as amino acids, nitrogen bases and nitrogen compounds. The nitrogen assimilation process occurs in a two-step reaction catalyzed by 2 enzymes working sequentially, nitrate reductase (NR) and nitrite reductase (NiR). The NR catalyzes the reduction of nitrate to nitrite (is considered the limiting step in the nitrogen assimilation) being this later reduced to ammonium by NiR. NR is widely distributed and found in different organisms, including bacterium, fungus, cyanobacterium, plants and algae. In this work we study the NR of Microcystis aeruginosa, a toxic microalga, mainly found in water reservoirs. The microcystin toxin released by M. aeruginosa is associated with problems of health in humans and animals. We report that NR of M. aeruginosa belongs to a biespecific group of NRs for NADH and NADPH. It presents Michaelis-Menten\'s constant (Km) as 1.5 and 1.6 mM for NADPH and NADH, respectively. The apparent Km for nitrate was estimated as 0.6 mM. The optimum conditions of assay found were at pH 10.0 and temperature of 40ºC. The exposition of M. aeruginosa to the herbicide oxyfluorfen (10 &#181;g/L) promotes the inhibition of NiR, and it makes possible to quantify the &#8226;NO produced by NR, whose has it activity 6 hold higher during the agent exposition. In order to understand the regulation of nitrogen assimilation enzymes, as well as, the mechanisms of nutrition and growth of this algae, the study of the NR enzyme is of crucial importance.

Orientadores: Telma Teixeira Franco, Eduardo Jacob-Lopes / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia Química / Made available in DSpace on 2018-08-24T13:35:37Z (GMT). No. of bitstreams: 1 Francisco_ErikaCristina_D.pdf: 1584353 bytes, checksum: 7fa11e2a9de2d170e12cddabf84ac647 (MD5) Previous issue date: 2014 / Resumo: O objetivo do estudo foi avaliar a produção de biomassa e o acúmulo lipídico da cianobactéria Phormidium sp. a partir do cultivo heterotrófico com diferentes fontes de carbono orgânico, estudar diferentes estratégias de cultivo empregando a manipueira (água residual do processo para obtenção da farinha de mandioca) e avaliar o acúmulo lipídico a partir de diferentes fontes de nitrogênio. Primeiramente avaliou-se a capacidade da cianobactéria Phormidium sp. em se desenvolver a partir de 15 diferentes fontes de carbono orgânico exógeno. Os resultados indicaram o amido de mandioca e a maltodextrina como as fontes com maior potencial de exploração para a produção de biomassa e lipídeos. Em um segundo momento, foi realizada a intensificação do processo de produção de biomassa a partir do amido de mandioca empregando-se um planejamento experimental. Os resultados indicaram que relação C/N de 68 e temperatura de 30ºC são as condições operacionais ideais. A extrapolação do cultivo em Erlenmeyers para as operações em batelada e batelada com alimentação descontínua em biorreatores resultou em produtividades de biomassa de 50,72 mg/L.h e 42,13 mg/L.h, respectivamente. A terceira etapa do projeto contemplou o estudo do cultivo heterotrófico da cianobactéria empregando diferentes modos de cultivo utilizando manipueira como substrato, inicialmente através de diferentes concentrações (v/v) de manipueira em água (5, 10, 20, 40, 60, 80 e 100%). Os resultados demonstraram que a melhor concentração da manipueira foi a de 60% (v/v). Em seguida, iniciou-se cultivos em biorreator, nos modos de batelada (Sistema A) e batelada com alimentação descontínua, empregando-se manipueira como meio de cultivo e diferentes soluções de alimentação. No primeiro sistema alimentado (Sistema B), empregou-se uma solução de amido de mandioca concentrado, e no segundo (Sistema C), o biorreator foi alimentado com manipueira não diluída. As alimentações foram efetuadas assim que a concentração de DQO no biorreator atingisse uma concentração de 6000mg/L. A extrapolação para cultivos em biorreator resultou em maiores produtividades em biomassa (53,12mg/L.h) e lipídica (7,45mg/L.h) sob batelada (sistema A). A última etapa do projeto contemplou o estudo de diferentes fontes de nitrogênio (extrato de levedura, nitrato de sódio, nitrito de sódio, molibidato de amônio e ureia) no cultivo heterotrófico da Phormidium sp. empregando amido de mandioca como fonte de carbono orgânico. A partir dos resultados obtidos selecionou-se o nitrato de sódio sob razão C/N de 60 como a fonte com maior potencial na produção de lipídeos, resultando na quantidade de lipídeos, produtividades lipídica e de biomassa de 20,58%, 7,62mg/L.h e 37,02mg/L.h, respectivamente. O processo de depleção do nitrato de sódio resultou em um aumento na quantidade de lipídeos (25,07%) e na produtividade lipídica (10,47mg/L.h) / Abstract: The aim of the study was to evaluate the production of biomass and lipid accumulation of cyanobacteria Phormidium sp. from heterotrophic cultivation with different sources of organic carbon, study diferente strategies of cultures employing cassava wastewater and analyse the lipid accumulation from different nitrogen sources. First we assessed the ability of the cyanobacterium Phormidium sp. to growth in 15 different exogenous sources of organic carbon. Results indicated that the cassava starch and maltodextrin as those with the highest potential for exploitation for the production of biomass and lipids. In a second step, the increase in biomass production process was made from cassava starch employing an experimental design. The results indicated that the C/N of 68 and temperature of 30ºC are the optimal operating conditions. Extrapolation of cultivation in flasks for batch and batch with fed discontinuous operations in bioreactors resulted in biomass productivity of 50.72 mg/L.h and 42.13mg/L.h, respectively. The third stage of the project involved the study of the heterotrophic system of cyanobacteria using different modes of cultivation using cassava wastewater as substrate, initially through different concentrations (v/v) of wastewater in water (5, 10, 20, 40, 60, 80 and 100%). The results showed that the best concentration of cassava was 60%. Then, in a bioreactor cultivation was started in batch mode (System A), and batch with descontinuous feeds, using the culture medium as wastewater and different feed solutions. In the first feed system (System B) used a concentrated solution of cassava starch, and in second (System C), the bioreactor was fed with undiluted wastewater. The feeds were made so that the concentration of COD in the bioreactor to achieve a concentration of 6000mg/L. The extrapolation to cultivations in bioreactor resulted in productivity of biomass (53.12 mg/L.h) and lipid (7.45 mg/L.h) under simple batch (System A). The last stage of the project involved the study of different nitrogen sources (ammonium molybdate, sodium nitrate, sodium nitrite, urea and yeast extract) in heterotrophic cultures of Phormidium sp. using cassava starch as a source of organic carbon. From the results obtained, we selected sodium nitrate under C/N ratio of 60 as the source with the highest potential in the production of lipids, resulting in the amount of lipids, productivity of lipid and biomass of 20.58%, 7.62 mg/L.h and 37.02 mg/L.h, respectively. The process of depletion of sodium nitrate resulted in an increase in the amount of lipids (25.07%) and the lipid productively (10.47mg/L.h) / Doutorado / Engenharia de Processos / Doutora em Engenharia Quimica

No ambiente aquático a maior fonte de nitrogênio encontra-se na forma de nitrato que necessita sofrer uma redução para formação de compostos biologicamente aproveitáveis, tais como aminoácidos, bases nitrogenadas e compostos nitrogenados. A assimilação de nitrogênio é um processo que ocorre em duas etapas catalisadas seqüencialmente pelas enzimas nitrato redutase (NR) e nitrito redutase (NiR). A NR catalisa a redução do nitrato a nitrito (etapa considerada limitante na assimilação de nitrogênio), sendo este posteriormente reduzido a amônio pela NiR. NR está amplamente distribuída e encontrada em diferentes organismos, incluíndo bactérias, fungos, cianobactérias, plantas terrestres e algas. Neste trabalho estudamos a NR de Microcystis aeruginosa que é uma cianobactéria tóxica de água doce encontrada principalmente em reservatórios de água. A toxina microcistina quando liberada por esta microalga está associada com problemas de saúde em humanos e animais. Foi mostrado que a NR de M. aeruginosa pertence a classe das NRs biespecíficas para NADH e NADPH. Apresenta constante de Michaelis-Menten aparente (Km) de 1,5 e 1,6 mM para NADPH e NADH, respectivamente. Ainda, Km aparente para nitrato foi estimado em 0,6 mM. As condições ótimas de ensaio encontradas foram em pH 10,0 e temperatura em 40ºC. A exposição da M. aeruginosa ao herbicida oxifluorfeno (10 &#181;g/L) promoveu a inibição de NiR, possibilitando a quantificação de &#8226;NO formado via NR, enzima que teve sua atividade 6 vezes maior durante a exposição a este agente. O estudo da enzima NR é de fundamental importância para a compreensão da regulação da expressão de enzimas assimiladoras de nitrogênio bem como dos mecanismos de nutrição e crescimento desta microalga. / Nitrate is the major source of nitrogen in the aquatic environment, which must be reduced before incorporation into biological compounds, such as amino acids, nitrogen bases and nitrogen compounds. The nitrogen assimilation process occurs in a two-step reaction catalyzed by 2 enzymes working sequentially, nitrate reductase (NR) and nitrite reductase (NiR). The NR catalyzes the reduction of nitrate to nitrite (is considered the limiting step in the nitrogen assimilation) being this later reduced to ammonium by NiR. NR is widely distributed and found in different organisms, including bacterium, fungus, cyanobacterium, plants and algae. In this work we study the NR of Microcystis aeruginosa, a toxic microalga, mainly found in water reservoirs. The microcystin toxin released by M. aeruginosa is associated with problems of health in humans and animals. We report that NR of M. aeruginosa belongs to a biespecific group of NRs for NADH and NADPH. It presents Michaelis-Menten\'s constant (Km) as 1.5 and 1.6 mM for NADPH and NADH, respectively. The apparent Km for nitrate was estimated as 0.6 mM. The optimum conditions of assay found were at pH 10.0 and temperature of 40ºC. The exposition of M. aeruginosa to the herbicide oxyfluorfen (10 &#181;g/L) promotes the inhibition of NiR, and it makes possible to quantify the &#8226;NO produced by NR, whose has it activity 6 hold higher during the agent exposition. In order to understand the regulation of nitrogen assimilation enzymes, as well as, the mechanisms of nutrition and growth of this algae, the study of the NR enzyme is of crucial importance.