Anabaena Strains Research Articles

Comparative proteomics of oxidative stress response in three cyanobacterial strains native to Indian paddy fields

Three strains of photoautotrophic, heterocystous, nitrogen-fixing cyanobacterium Anabaena, native to Indian paddy fields, were examined for their tolerance and proteomic response to the frequently used weedicide paraquat (methyl viologen). Anabaena 7120 (LD50 dose: 2μM for 6h) and Anabaena L-31 (LD50 dose: 2μM for 5h) showed distinctly better tolerance than Anabaena doliolum (LD50 dose: 2μM for 3h), to methyl viologen induced oxidative stress. The proteomic response, at respective LD50 dose, was mapped by 2D gel protein electrophoresis followed by protein identification by MALDI-ToF mass spectrometry. About 92 and 41 oxidative stress-responsive proteins were identified from Anabaena L-31 and A. doliolum, respectively, and compared with methyl viologen responsive proteins reported from Anabaena 7120 earlier. Upregulation of proteins involved in oxidative stress alleviation and protein homeostasis and downregulation of photosynthesis and carbon metabolism related enzymes appeared to underlie the oxidative stress response in all three Anabaena strains. Reduced photosynthesis and cellular reserves of molecular energy [ATP+NAD(P)H] seemed to overwhelm the cellular machinery to combat oxidative stress and protein denaturation, in preference to other adaptations, while the strain specific differences observed in proteome response appeared to determine the methyl viologen tolerance of individual cyanobacterial strains.This article is part of a Special Issue entitled: Proteomics in India.

Antifungal compounds from cyanobacteria.

Cyanobacteria are photosynthetic prokaryotes found in a range of environments. They are infamous for the production of toxins, as well as bioactive compounds, which exhibit anticancer, antimicrobial and protease inhibition activities. Cyanobacteria produce a broad range of antifungals belonging to structural classes, such as peptides, polyketides and alkaloids. Here, we tested cyanobacteria from a wide variety of environments for antifungal activity. The potent antifungal macrolide scytophycin was detected in Anabaena sp. HAN21/1, Anabaena cf. cylindrica PH133, Nostoc sp. HAN11/1 and Scytonema sp. HAN3/2. To our knowledge, this is the first description of Anabaena strains that produce scytophycins. We detected antifungal glycolipopeptide hassallidin production in Anabaena spp. BIR JV1 and HAN7/1 and in Nostoc spp. 6sf Calc and CENA 219. These strains were isolated from brackish and freshwater samples collected in Brazil, the Czech Republic and Finland. In addition, three cyanobacterial strains, Fischerella sp. CENA 298, Scytonema hofmanni PCC 7110 and Nostoc sp. N107.3, produced unidentified antifungal compounds that warrant further characterization. Interestingly, all of the strains shown to produce antifungal compounds in this study belong to Nostocales or Stigonematales cyanobacterial orders.

Induction and differential expression of certain novel proteins in Anabaena L31 under UV-B radiation stress.

For examining how UV-B radiation alters the proteome of the N2-fixing cyanobacterium, Anabaena L31, we extracted proteins from cultures irradiated with UV-B + white light and controls (white light irradiated) and analyzed the proteins using two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS). Twenty one proteins, including two hypothetical proteins (HPs) were identified and placed in eight functional categories. However several of the proteins were housekeeping proteins involved in key metabolic processes such as carbon, amino acid biosynthesis and energy metabolism, certain proteins seem to have a role in stress (antioxidative enzymes), translation, cellular processes and reductases. Two novel HPs (all3797 and all4050) were characterized in detail. These two were over-expressed after UV-B irradiation and characterized as FAS 1 (all3797) and PRC barrel-like (all4050) proteins. Bioinformatics analysis revealed that the genes of both the HPs have promoter regions as well as transcription binding sites in their upstream region (UTR). Promoters present in all3797 genes suggest their crucial role against UV-B and certain other abiotic stresses. To our knowledge these novel proteins have not been previously reported in any Anabaena strains subjected to UV-B stress. Although we have focused our study on a limited number of proteins, results obtained shed light on the highly complicated but poorly studied aspect of UV-B radiation-mediated changes in the proteome and expression of proteins in cyanobacteria.

Redox-dependent chaperone/peroxidase function of 2-Cys-Prx from the cyanobacterium Anabaena PCC7120: role in oxidative stress tolerance.

BackgroundCyanobacteria, progenitors of plant chloroplasts, provide a suitable model system for plants to study adaptation towards different abiotic stresses. Genome of the filamentous, heterocystous, nitrogen-fixing cyanobacterium Anabaena PCC7120 harbours a single gene (alr4641) encoding a typical 2-Cys-Peroxiredoxins (2-Cys-Prxs). 2-Cys-Prxs are thiol-based peroxidases that also function as molecular chaperones in plants and other systems. The Alr4641 protein from Anabaena PCC7120 shows high level biochemical similarities with the plant 2-Cys-Prx. The physiological role played by the Alr4641 protein in Anabaena was addressed in this study.ResultsIn Anabaena PCC7120, alr4641 transcript /Alr4641 protein was induced in response to abiotic stresses and its promoter was active in the vegetative cells as well as heterocysts. The wild-type Alr4641 protein or Alr4641 lacking the peroxidatic cysteine (Alr4641C56S) or the resolving cysteine (Alr4641C178S) existed as higher oligomers in their native form. The wild-type or the mutant Alr4641 proteins showed similar chaperone activity, but only the wild-type protein exhibited peroxidase activity indicating that unlike peroxidase activity, chaperone activity was not dependent on cysteines. In contrast to other 2-Cys-Prxs, chaperone/peroxidase activity of Alr4641 was dependent on its redox state and not oligomerization status. Alr4641 could protect plasmid DNA from oxidative damage and physically associate with NADPH-dependent thioredoxin reductase (NTRC). Like 2-Cys-Prxs from plants (e.g. rice), Alr4641 could detoxify various peroxides using NTRC as reductant. On exposure to H2O2, recombinant Anabaena PCC7120 strain over-expressing Alr4641 (An4641+) showed reduced content of reactive oxygen species (ROS), intact photosynthetic functions and consequently better survival than the wild-type Anabaena PCC7120, indicating that Alr4641 can protect Anabaena from oxidative stress.ConclusionsThe peroxidase/chaperone function of Alr4641, its inherent transcriptional/translational induction under different abiotic stresses and localization in both vegetative cells and heterocysts could be an adaptive strategy to battle various oxidative stresses that Anabaena encounters during its growth. Moreover, the recombinant Anabaena strain over expressing Alr4641 showed higher resistance to oxidative stress, suggesting its potential to serve as stress-tolerant biofertilizers in rice fields.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-015-0444-2) contains supplementary material, which is available to authorized users.

Draft genome sequence of calothrix strain 336/3, a novel h2-producing cyanobacterium isolated from a finnish lake.

We announce the draft genome sequence of Calothrix strain 336/3, an N2-fixing heterocystous filamentous cyanobacterium isolated from a natural habitat. Calothrix 336/3 produces higher levels of hydrogen than Nostoc punctiforme PCC 73102 and Anabaena strain PCC 7120 and, therefore, is of interest for potential technological applications.

First report of an Anabaena Bory strain containing microcystin-LR in a freshwater body in Africa

In South Africa, little is known about the production of microcystin by the genus Anabaena Bory. In April 2012, during a cyanobacterial bloom event in Theewaterskloof Dam, Western Cape province, the plankton was sampled on 10 occasions. The dominant algae belonged to the genus Anabaena, a family of filamentous cyanobacteria known to produce cyanotoxins such as anatoxin-a, harmful to humans and the aquatic foodweb. The specimens isolated lacked the characteristic akinetes and/or heterocysts associated with this genus. Therefore the 16S rRNA gene was Sanger sequenced and a maximum parsimony tree was constructed, confirming its identity as Anabaena ucrainica (Schkorbatow) M. Watanabe. Enzyme-linked immunosorbent assay (ELISA) confirmed the presence of microcystin-LR in the isolated A. ucrainica field sample, while PCR analysis and sequencing further confirmed the presence of mcy genes in this species. It was speculated from the data that prevailing low water-column temperatures and strong gusty winds may have resulted in the lack of akinete or heterocyst production. The Anabaena strain isolated from Theewaterskloof Dam is the first report of a strain containing microcystin-LR belonging to this genus in a freshwater body in Africa.

Sustained photobiological hydrogen production in the presence of N2 by nitrogenase mutants of the heterocyst-forming cyanobacterium Anabaena

Heterocyst-forming cells of the cyanobacterium Anabaena sp. strain PCC 7120 ΔHup, lacking an uptake hydrogenase, photobiologically produce H2 by nitrogenase. Under N2-rich atmosphere, the nitrogenase activity declines in a rather short time due to the sufficiency of combined nitrogen. From the parental ΔHup strain, site-directed double-crossover variants, dc-Q193S and dc-R284H, were created with amino acid substitutions presumed to be located in the vicinity of the FeMo-cofactor of nitrogenase. Unlike the case for the ΔHup strain, H2 production activities of the variants were not decreased by the presence of high concentrations of N2 and they continuously produced H2 over 21 days with occasional headspace gas replacement. This property of N2 insensitivity is a potentially useful strategy for reducing the cost of the culture gas in future practical applications of sustainable biofuel production. This Anabaena strain has only the Mo-containing nitrogenase which reduces acetylene to ethylene, but the dc-Q193S variant also produced ethane at low but measurable rates along with greater rates of ethylene production.

Inactivation of agmatinase expressed in vegetative cells alters arginine catabolism and prevents diazotrophic growth in the heterocyst-forming cyanobacterium Anabaena.

Arginine decarboxylase produces agmatine, and arginase and agmatinase are ureohydrolases that catalyze the production of ornithine and putrescine from arginine and agmatine, respectively, releasing urea. In the genome of the filamentous, heterocyst-forming cyanobacterium Anabaena sp. strain PCC 7120, ORF alr2310 putatively encodes an ureohydrolase. Cells of Anabaena supplemented with [14C]arginine took up and catabolized this amino acid generating a set of labeled amino acids that included ornithine, proline, and glutamate. In an alr2310 deletion mutant, an agmatine spot appeared and labeled glutamate increased with respect to the wild type, suggesting that Alr2310 is an agmatinase rather than an arginase. As determined in cell-free extracts, agmatinase activity could be detected in the wild type but not in the mutant. Thus, alr2310 is the Anabaena speB gene encoding agmatinase. The Δalr2310 mutant accumulated large amounts of cyanophycin granule polypeptide, lacked nitrogenase activity, and did not grow diazotrophically. Growth tests in solid media showed that agmatine is inhibitory for Anabaena, especially under diazotrophic conditions, suggesting that growth of the mutant is inhibited by non-metabolized agmatine. Measurements of incorporation of radioactivity from [14C]leucine into macromolecules showed, however, a limited inhibition of protein synthesis in the Δalr2310 mutant. Analysis of an Anabaena strain producing an Alr2310-GFP (green fluorescent protein) fusion showed expression in vegetative cells but much less in heterocysts, implying compartmentalization of the arginine decarboxylation pathway in the diazotrophic filaments of this heterocyst-forming cyanobacterium.

Synthesis of ZnO nanoparticles using the cell extract of the cyanobacterium, Anabaena strain L31 and its conjugation with UV-B absorbing compound shinorine

In the present work, we describe a cheap, unexplored and simple procedure for the synthesis of zinc oxide nanoparticles (ZnONPs) using the cell extract of the cyanobacterium, Anabaena strain L31. An attempt was also made to conjugate synthesized ZnONPs with a UV-absorbing water soluble compound shinorine. UV–vis spectroscopy, X-ray diffraction (XRD), Fourier transform infra-red (FTIR) spectroscopy, transmission electron microscopy (TEM) and TEM–selected area electron diffraction (SAED) analyses were made to elucidate the formation and characterization of ZnONPs and ZnONPs-shinorine conjugate. The synthesized ZnONPs were characterized by a sharp peak at 370nm in UV–vis spectrum. TEM images showed the formation of spherical shaped nanoparticles with an average size of 80nm. Results of selective area electron diffraction (SAED) pattern showed a set of rings which suggested uniform shape with hexagonal structure of ZnONPs. XRD spectra confirmed the crystalline structure of particles. Conjugation of ZnONPs with shinorine was successfully achieved at pH 7.0 and 10mM concentration of shinorine. The conjugate showed a zeta potential value of −3.75mV as compared to +30.25mV of ZnONPs. The change in zeta potential value of ZnONPs–shinorine conjugate was attributed to the changes in the surface functionalities after conjugation. The generation of in vivo reactive oxygen species (ROS) by Anabaena strain L31 with treatment of ZnONPs–shinorine conjugate showed approximately 75% less ROS generation as compared to ZnONPs. Properties exhibited by the ZnONPs–shinorine conjugate suggest that it may be used as a potential agent in developing environmental-friendly sunscreen filters of biological origin.

Hassallidins, antifungal glycolipopeptides, are widespread among cyanobacteria and are the end-product of a nonribosomal pathway.

Cyanobacteria produce a wide variety of cyclic peptides, including the widespread hepatotoxins microcystins and nodularins. Another class of peptides, cyclic glycosylated lipopeptides called hassallidins, show antifungal activity. Previously, two hassallidins (A and B) were reported from an epilithic cyanobacterium Hassallia sp. and found to be active against opportunistic human pathogenic fungi. Bioinformatic analysis of the Anabaena sp. 90 genome identified a 59-kb cryptic inactive nonribosomal peptide synthetase gene cluster proposed to be responsible for hassallidin biosynthesis. Here we describe the hassallidin biosynthetic pathway from Anabaena sp. SYKE748A, as well as the large chemical variation and common occurrence of hassallidins in filamentous cyanobacteria. Analysis demonstrated that 20 strains of the genus Anabaena carry hassallidin synthetase genes and produce a multitude of hassallidin variants that exhibit activity against Candida albicans. The compounds discovered here were distinct from previously reported hassallidins A and B. The IC50 of hassallidin D was 0.29-1.0 µM against Candida strains. A large variation in amino acids, sugars, their degree of acetylation, and fatty acid side chain length was detected. In addition, hassallidins were detected in other cyanobacteria including Aphanizomenon, Cylindrospermopsis raciborskii, Nostoc, and Tolypothrix. These compounds may protect some of the most important bloom-forming and globally distributed cyanobacteria against attacks by parasitic fungi.

Planktic cyanobacteria from freshwater localities in Thuathien-Hue province, Vietnam. III. Phylogenetic inference based on partial phycocyanin sequences, morphological and toxicological characters

The morphology, toxicity and partial sequences of beta and alpha subunits including the intergenic spacer (cpcBA-IGS) of the phycocyanin operon were studied in 21 strains of nonaxenic cyanobacterial cultures. The cultures were established from water samples collected in the Thuathien Hue province in Vietnam, 2004. Based on testing for the presence of the mcyA gene, microcystin-production of Microcystis aeruginosa, M. botrys and M. panniformis was confirmed. The presence of the mcyA gene in a strain of Microcystis flos-aquae and a strain of Planktothrix spiroides (HA308) revealed their toxic potential. The phylogeny of the Vietnamese cyanobacteria was inferred using sequences encoding the cpcBA genes. There was no correlation between toxic characters based on ELISA, HPLC and mcyA gene and the phylogeny based on partial cpcBA sequences. Although the resulting tree topology assisted in resolving the phylogeny of some strains of Anabaena, Dolichospermum and Planktothrix, taxonomical problems still exist at the species level for Lyngbya and Microcystis. Future morphological and molecular studies will have to solve these issues.

Polyphasic characterization of Nostoc commune (Cyanobacteria, Nostocaceae) isolated from rice growing agro-ecosystems of Dima Hasao district of Assam, North-East India

Two strains of Nostoc commune have been isolated from the soils of two different rice-growing agro-ecosystems, viz. flat and terrace paddy fields of Dima Hasao district of the state of Assam, North-East India. Phenotypic characterization was made for both the strains and their growth, pigments (chlorophyll a, total carotenoid content and phycobiliproteins) and biochemical properties (total carbohydrate and soluble proteins) were studied. Phylogenetic comparison was made utilizing 16S rRNA gene sequences. Both strains presented higher phycocyanin content than other biliprotein pigments. Total carotenoid content (TCC) was higher in the strain isolated from flat paddy field, while the isolate from terrace paddy field was richer in phycobiliproteins. 16S rRNA gene sequences of the isolated N. commune strains were compared with available sequences of other strains of Nostoc and Anabaena from various geographical locations. Gene sequences were clustered according to their geographical origin, which also reflected the disputed taxonomic position of the Nostocacean genera Nostoc and Anabaena.

Phylogeography of Cylindrospermopsin and Paralytic Shellfish Toxin-Producing Nostocales Cyanobacteria from Mediterranean Europe (Spain)

Planktonic Nostocales cyanobacteria represent a challenge for microbiological research because of the wide range of cyanotoxins that they synthesize and their invasive behavior, which is presumably enhanced by global warming. To gain insight into the phylogeography of potentially toxic Nostocales from Mediterranean Europe, 31 strains of Anabaena (Anabaena crassa, A. lemmermannii, A. mendotae, and A. planctonica), Aphanizomenon (Aphanizomenon gracile, A. ovalisporum), and Cylindrospermopsis raciborskii were isolated from 14 freshwater bodies in Spain and polyphasically analyzed for their phylogeography, cyanotoxin production, and the presence of cyanotoxin biosynthesis genes. The potent cytotoxin cylindrospermopsin (CYN) was produced by all 6 Aphanizomenon ovalisporum strains at high levels (5.7 to 9.1 μg CYN mg(-1) [dry weight]) with low variation between strains (1.5 to 3.9-fold) and a marked extracellular release (19 to 41% dissolved CYN) during exponential growth. Paralytic shellfish poisoning (PSP) neurotoxins (saxitoxin, neosaxitoxin, and decarbamoylsaxitoxin) were detected in 2 Aphanizomenon gracile strains, both containing the sxtA gene. This gene was also amplified in non-PSP toxin-producing Aphanizomenon gracile and Aphanizomenon ovalisporum. Phylogenetic analyses supported the species identification and confirmed the high similarity of Spanish Anabaena and Aphanizomenon strains with other European strains. In contrast, Cylindrospermopsis raciborskii from Spain grouped together with American strains and was clearly separate from the rest of the European strains, raising questions about the current assumptions of the phylogeography and spreading routes of C. raciborskii. The present study confirms that the nostocalean genus Aphanizomenon is a major source of CYN and PSP toxins in Europe and demonstrates the presence of the sxtA gene in CYN-producing Aphanizomenon ovalisporum.

Antioxidant and iron-chelating activities of cyanobacterial exopolymers with potential for wound healing

Progress of wound healing is critically dependent on the balance between oxidants and antioxidants at the wound site, and transition metals such as iron can exacerbate ROS generation. In the present study, cyanobacterial exopolymers from three strains of Anabaena and Tolypothrix tenuis have been characterized for their antiradical and Fe2+-chelating activity. All the four exopolymers exhibited antioxidant activities against O2·, H2O2, OH·, and NO·, with the exopolymer from Anabaena oryzae showing strong inhibition of NO· and ·OH radicals followed by that from Anabaena anomala. Correlation analysis of antioxidant activities and sulphate, uronic and phenolic content of the exopolymers showed a strong correlation of sulphate content to superoxide scavenging and activity against nitric oxide radicals. H2O2 scavenging was related to the presence of phenolics in the preparation which also contributed to the reducing power. Iron chelation had a strong bearing upon the overall reducing power and superoxide control.

Oxidative stress management in the filamentous, heterocystous, diazotrophic cyanobacterium, Anabaena PCC7120.

Reactive oxygen species (ROS) are inevitably generated as by-products of respiratory/photosynthetic electron transport in oxygenic photoautotrophs. Unless effectively scavenged, these ROS can damage all cellular components. The filamentous, heterocystous, nitrogen-fixing strains of the cyanobacterium, Anabaena, serve as naturally abundant contributors of nitrogen biofertilizers in tropical rice paddy fields. Anabaena strains are known to tolerate several abiotic stresses, such as heat, UV, gamma radiation, desiccation, etc., that are known to generate ROS. ROS are detoxified by specific antioxidant enzymes like superoxide dismutases (SOD), catalases and peroxiredoxins. The genome of Anabaena PCC7120 encodes two SODs, two catalases and seven peroxiredoxins, indicating the presence of an elaborate antioxidant enzymatic machinery to defend its cellular components from ROS. This article summarizes recent findings and depicts important perspectives in oxidative stress management in Anabaena PCC7120.

N‐terminal processing of membrane‐targeted MnSOD and formation of multiple active superoxide dismutase dimers in the nitrogen‐fixing cyanobacterium Anabaena sp. strain PCC7120

Anabaena sp. strain PCC7120 expresses a 30 kDa manganese-dependent superoxide dismutase (MnSOD) comprising a hydrophobic region (signal peptide + linker peptide) attached to a catalytic unit. Bioinformatics predicted cleavage of the signal peptide at (25)CQPQ by signal peptidase and of the linker peptide by an Arg-C-like protease at the Arg52/Arg59 residue. The three predicted forms of MnSOD were immunodetected in Anabaena, with the 30 kDa MnSOD found exclusively in the membrane and the shorter 27 and 24 kDa forms found both in the membrane and soluble fractions. The corresponding sodA gene was truncated for (a) the first eight residues, or, (b) the signal peptide, or (c) the entire hydrophobic region, or (d) the Arg52/Arg59 residues were modified to serine. Overexpression of these MnSOD variants in recombinant Anabaena strains revealed that (a) the 30 kDa membrane-targeted MnSOD was cleaved by membrane-localized signal peptidase either during or after its transport through the membrane to release the 27 kDa form, either in the cytosol or in the periplasmic/thylakoid lumen, (b) the 27 kDa form was further cleaved to the 24 kDa form by Arg-C-like protease, both in the cytosol and in the periplasmic/thylakoid lumen, (c) deletion of signal peptide localized the MnSOD forms in the cytosol, and (d) alteration of the signal/linker peptide cleavage sites interfered with MnSOD localization and processing. Homo/heterodimerization of the 24 and 27 kDa forms of MnSOD and the cytosolic iron-dependent SOD results in multiple SOD activities, from a single MnSOD gene (sodA), in different cellular compartments of Anabaena.

The occurrence of cyanobacteria and microcystins in the Hoan Kiem Lake and the Nui Coc reservoir (North Vietnam)

The occurrence of increasing blooms of toxic cyanobacteria in freshwaters has received much attention due to the ability of many cyanobacteria to produce potent cyanotoxins. In this paper, the occurrence of dominant cyanobacteria and the concentration of microcystins (MCs) analysis were investigated monthly from July 2008 to April 2009 in the Hoan Kiem Lake and from February to April 2009 in the Nui Coc reservoir. Concentrations of intracellular MCs from water, bloom samples, and isolated strains were quantified by using high performance liquid chromatography (HPLC). During the study period, the microscopic examination of the phytoplankton samples showed the dominance of the genus Microcystis in the water environment of the Hoan Kiem Lake and the Nui Coc reservoir. The toxin analysis by HPLC demonstrated the presence of two MC variants: MC-LR and MC-RR in water samples. Total concentrations of the toxins in filtered samples from surface water ranged from non-detected to 0.91 μg L−1 at Nui Coc reservoir and they ranged from 2.1 to 46.0 μg L−1 at Hoan Kiem Lake. The results of the HPLC analysis confirmed the production of MCs in bloom samples (ranged from 115.9 to 184.6 μg L−1 in the Hoan Kiem Lake and from 726.5 to 1116 μg L−1 in the Nui Coc reservoir) and isolated strains of Anabaena sp. and Microcystis with the concentration of MC ranging from 152 to 396.2 μg g−1 dry mass, respectively.

New N-acyl-d-glucosamine 2-epimerases from cyanobacteria with high activity in the absence of ATP and low inhibition by pyruvate

N-Acetylneuraminic acid, an important component of glycoconjugates with various biological functions, can be produced from N-acetyl-d-glucosamine (GlcNAc) and pyruvate using a one-pot, two-enzyme system consisting of N-acyl-d-glucosamine 2-epimerase (AGE) and N-acetylneuraminate lyase (NAL). In this system, the epimerase catalyzes the conversion of GlcNAc into N-acetyl-d-mannosamine (ManNAc). However, all currently known AGEs have one or more disadvantages, such as a low specific activity, substantial inhibition by pyruvate and strong dependence on allosteric activation by ATP. Therefore, four novel AGEs from the cyanobacteria Acaryochloris marina MBIC 11017, Anabaena variabilis ATCC 29413, Nostoc sp. PCC 7120, and Nostoc punctiforme PCC 73102 were characterized. Among these enzymes, the AGE from the Anabaena strain showed the most beneficial characteristics. It had a high specific activity of 117±2Umg−1 at 37°C (pH 7.5) and an up to 10-fold higher inhibition constant for pyruvate as compared to other AGEs indicating a much weaker inhibitory effect. The investigation of the influence of ATP revealed that the nucleotide has a more pronounced effect on the Km for the substrate than on the enzyme activity. At high substrate concentrations (≥200mM) and without ATP, the enzyme reached up to 32% of the activity measured with ATP in excess.

Optimization of photobioreactor growth conditions for a cyanobacterium expressing mosquitocidal Bacillus thuringiensis Cry proteins

An Anabaena strain (PCC 7120#11) that was genetically engineered to express Bacillus thuringiensis subsp. israelensis cry genes has shown good larvicidal activity against Anopheles arabiensis, a major vector of malaria in Africa. Response surface methodology was used to evaluate the relationship between key growth factors and the volumetric productivity of PCC 7120#11 in an indoor, flat-plate photobioreactor. The interaction of input CO2 concentration and airflow rate had a statistically significant effect on the volumetric productivity of PCC 7120#11, as did the interaction of airflow rate and photosynthetic photon flux density. Model-based numerical optimization indicated that the optimal factor level combination for maximizing PCC 7120#11 volumetric productivity was a photosynthetic photon flux density of 154μmolm−2s−1 and air enriched with 3.18% (v/v) CO2 supplied at a flow rate of 1.02 vessel volumes per minute. At the levels evaluated in the study, none of the growth factors had a significant effect on the median lethal concentration of PCC 7120#11 against An. arabiensis larvae. This finding is important because loss of mosquitocidal activity under growth conditions that maximize volumetric productivity would impact on the feasibility of using PCC 7120#11 in malaria vector control programs. The study showed the usefulness of response surface methodology for determination of the optimal growth conditions for a cyanobacterium that is genetically engineered to have larvicidal activity against malaria vectors.

Enzymatic and non-enzymatic defense mechanisms against ultraviolet-B radiation in two Anabaena species

Enzymatic and non-enzymatic defense strategies against ultraviolet-B radiation (UV-B, 280–315nm) were studied in Anabaena doliolum and Anabaena strain L31, two of the most common strains of Indian cyanobacteria. Upon UV-B irradiation, both strains showed a 2–5-fold increase in antioxidative enzymes, such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and peroxidase (POD), as compared to non-irradiated control cultures. These enzymes scavenge damaging reactive oxygen species (ROS), generated by UV-B radiation inside the cells. In addition, these organisms also synthesize mycosporine-like amino acids (MAAs) which are able to carry out UV-screening and/at the same time as UV-quenching. The identification and characterization of three types of MAAs from both Anabaena species were performed using absorption spectroscopy, high performance liquid chromatography (HPLC), electro spray ionization-mass spectrometry (ESI-MS), Fourier Transform Infrared (FTIR) spectroscopy and Nuclear Magnetic Resonance (NMR) spectroscopy. Shinorine was found to be the most common MAA in both Anabaena species while porphyra-334 and mycosporine-glycine were present only in A. doliolum. The results of the present investigation clearly demonstrate that both enzymatic and non-enzymatic defense mechanisms are being employed by A. doliolum and Anabaena strain L31 to counteract the damaging effects of UV-B radiation.