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Biochimica Et Biophysica Acta.... Dec 2021Cryptophytes are among the few eukaryotes employing phycobiliproteins (PBP) for light harvesting during oxygenic photosynthesis. In contrast to cyanobacterial PBP that...
Cryptophytes are among the few eukaryotes employing phycobiliproteins (PBP) for light harvesting during oxygenic photosynthesis. In contrast to cyanobacterial PBP that are organized in membrane-associated phycobilisomes, those from cryptophytes are soluble within the chloroplast thylakoid lumen. Their light-harvesting capacity is due to covalent linkage of several open-chain tetrapyrrole chromophores (phycobilins). Guillardia theta utilizes the PBP phycoerythrin 545 with 15,16-dihydrobiliverdin (DHBV) in addition to phycoerythrobilin (PEB) as chromophores. The assembly of PBPs in cryptophytes involves the action of PBP-lyases as shown for cyanobacterial PBP. PBP-lyases facilitate the attachment of the chromophore in the right configuration and stereochemistry. Here we present the functional characterization of the eukaryotic S-type PBP lyase GtCPES. We show GtCPES-mediated transfer and covalent attachment of PEB to the conserved Cys82 of the acceptor PBP β-subunit (PmCpeB) of Prochlorococcus marinus MED4. On the basis of the previously solved crystal structure, the GtCPES binding pocket was investigated using site-directed mutagenesis. Thereby, amino acid residues involved in phycobilin binding and transfer were identified. Interestingly, exchange of a single amino acid residue Met67 to Ala extended the substrate specificity to phycocyanobilin (PCB), most likely by enlarging the substrate-binding pocket. Variant GtCPES_M67A binds both PEB and PCB forming a stable, colored complex in vitro and produced in Escherichia coli. GtCPES_M67A is able to mediate PCB transfer to Cys82 of PmCpeB. Based on these findings, we postulate that this single amino acid residue has a crucial role for bilin binding specificity of S-type phycoerythrin lyases but additional factors regulate handover to the target protein.
Topics: Lyases; Phycobiliproteins; Substrate Specificity
PubMed: 34537203
DOI: 10.1016/j.bbabio.2021.148493 -
Journal of Biotechnology Nov 2021Phycobiliproteins are colored, active molecules with potential use in different industries. They are the union of proteins and bilins (Chromophores). The primary source...
Phycobiliproteins are colored, active molecules with potential use in different industries. They are the union of proteins and bilins (Chromophores). The primary source of phycobiliproteins is algae; however, the traditional algae culture has production restrictions. The production in bacterial models can be a more efficient alternative to produce these molecules. However, the lack of knowledge in some steps of the phycobiliprotein metabolic pathway limits this alternative. Porphyridium cruentum is a single cell red alga with a high phycobiliprotein content. Its protein sequences were the basis for phycobilin production in this study. In this study, we cloned and characterized enzymes presumably involved in the chromophore production of P. cruentum. Using sequences obtained from its transcriptome, we characterized two cDNA sequences predicted to code respectively for a ferredoxin-dependent bilin reductase and a bilin lyase-isomerase. We expressed these enzymes in Escherichia coli to obtain in vivo evidence of their enzymatic activity on the substrate biliverdin IXα. Lastly, we analyzed them using thin-layer chromatography, spectrophotometry, and fluorescence spectroscopy. These experiments provided evidence of bilin modification. The expressed bilin lyase-isomerase did not show significant activity over the biliverdin molecule. On the contrary, the expressed ferredoxin-dependent bilin reductase showed activity over the biliverdin.
Topics: Cyanobacteria; Lyases; Phycobilins; Porphyridium; Rhodophyta
PubMed: 34500003
DOI: 10.1016/j.jbiotec.2021.09.001 -
Physical Chemistry Chemical Physics :... Sep 2021Cyanobacteriochromes (CBCRs) are bi-stable photoreceptor proteins with high potential for biotechnological applications. Most of these proteins utilize phycocyanobilin...
Cyanobacteriochromes (CBCRs) are bi-stable photoreceptor proteins with high potential for biotechnological applications. Most of these proteins utilize phycocyanobilin (PCB) as a light-sensing co-factor, which is unique to cyanobacteria, but some variants also incorporate biliverdin (BV). The latter are of particular interest for biotechnology due to the natural abundance and red-shifted absorption of BV. Here, AmI-g2 was investigated, a CBCR capable of binding both PCB and BV. The assembly kinetics and primary photochemistry of AmI-g2 with both chromophores were studied . The assembly reaction with PCB is roughly 10× faster than BV, and the formation of a non-covalent intermediate was identified as the rate-limiting step in the case of BV. This step is fast for PCB, where the formation of the covalent thioether bond between AmI-g2 and PCB becomes rate-limiting. The photochemical quantum yields of the forward and backward reactions of AmI-g2 were estimated and discussed in the context of homologous CBCRs.
Topics: Biliverdine; Cyanobacteria; Kinetics; Photoreceptors, Microbial; Phycobilins; Phycocyanin; Protein Binding; Quantum Theory; Spectrophotometry
PubMed: 34374395
DOI: 10.1039/d1cp02696h -
Microbial Cell Factories Jul 2021The development of multiple gene expression systems, especially those based on the physical signals, such as multiple color light irradiations, is challenging....
BACKGROUND
The development of multiple gene expression systems, especially those based on the physical signals, such as multiple color light irradiations, is challenging. Complementary chromatic acclimation (CCA), a photoreversible process that facilitates the control of cellular expression using light of different wavelengths in cyanobacteria, is one example. In this study, an artificial CCA systems, inspired by type III CCA light-regulated gene expression, was designed by employing a single photosensor system, the CcaS/CcaR green light gene expression system derived from Synechocystis sp. PCC6803, combined with G-box (the regulator recognized by activated CcaR), the cognate cpcG2 promoter, and the constitutively transcribed promoter, the P promoter.
RESULTS
One G-box was inserted upstream of the cpcG2 promoter and a reporter gene, the rfp gene (green light-induced gene expression), and the other G-box was inserted between the P promoter and a reporter gene, the bfp gene (red light-induced gene expression). The Escherichia coli transformants with plasmid-encoded genes were evaluated at the transcriptional and translational levels under red or green light illumination. Under green light illumination, the transcription and translation of the rfp gene were observed, whereas the expression of the bfp gene was repressed. Under red light illumination, the transcription and translation of the bfp gene were observed, whereas the expression of the rfp gene was repressed. During the red and green light exposure cycles at every 6 h, BFP expression increased under red light exposure while RFP expression was repressed, and RFP expression increased under green light exposure while BFP expression was repressed.
CONCLUSION
An artificial CCA system was developed to realize a multiple gene expression system, which was regulated by two colors, red and green lights, using a single photosensor system, the CcaS/CcaR system derived from Synechocystis sp. PCC6803, in E. coli. The artificial CCA system functioned repeatedly during red and green light exposure cycles. These results demonstrate the potential application of this CCA gene expression system for the production of multiple metabolites in a variety of microorganisms, such as cyanobacteria.
Topics: Escherichia coli; Gene Expression; Gene Expression Regulation, Bacterial; Genes, Reporter; Light; Luminescent Proteins; Phycobilins; Phycocyanin; Promoter Regions, Genetic; Synechocystis; Transcription, Genetic; Transformation, Bacterial; Red Fluorescent Protein
PubMed: 34225717
DOI: 10.1186/s12934-021-01621-3 -
Frontiers in Microbiology 2021In the 21st century, we have witnessed three coronavirus outbreaks: SARS in 2003, MERS in 2012, and the ongoing pandemic coronavirus disease 2019 (COVID-19). The search...
In the 21st century, we have witnessed three coronavirus outbreaks: SARS in 2003, MERS in 2012, and the ongoing pandemic coronavirus disease 2019 (COVID-19). The search for efficient vaccines and development and repurposing of therapeutic drugs are the major approaches in the COVID-19 pandemic research area. There are concerns about the evolution of mutant strains (e.g., VUI - 202012/01, a mutant coronavirus in the United Kingdom), which can potentially reduce the impact of the current vaccine and therapeutic drug development trials. One promising approach to counter the mutant strains is the "development of effective broad-spectrum antiviral drugs" against coronaviruses. This study scientifically investigates potent food bioactive broad-spectrum antiviral compounds by targeting main protease (M) and papain-like protease (PL) proteases of coronaviruses (CoVs) using and approaches. The results reveal that phycocyanobilin (PCB) shows potential inhibitor activity against both proteases. PCB had the best binding affinity to Mpro and PLpro with IC values of 71 and 62 μm, respectively. Also, studies with M and PL enzymes of other human and animal CoVs indicate broad-spectrum inhibitor activity of the PCB. As with PCB, other phycobilins, such as phycourobilin (PUB), phycoerythrobilin (PEB), and phycoviolobilin (PVB) show similar binding affinity to SARS-CoV-2 M and PL.
PubMed: 34177827
DOI: 10.3389/fmicb.2021.645713 -
Chemosphere Oct 2021Phyco-remediation of dyestuffs in textile wastewaters is of economic, industrial, and environmental importance. We evaluated the remediation of the textile dye, Direct...
Phyco-remediation of dyestuffs in textile wastewaters is of economic, industrial, and environmental importance. We evaluated the remediation of the textile dye, Direct Green 6 (DG6), by Spirulina platensis, and investigated the novel possibility that DG6 treatment enhances production of the biopolymer, polyhydroxybutyrate (PHB). We showed that both live and dead cells of Spirulina were capable of DG6 remediation, but live cells could be re-used with no loss of remediation efficiency. Furthermore, DG6 remediation by live cells resulted in increased algal biomass and trichome lengths, and stimulated production of valuable metabolites, including PHB, antioxidants, carbohydrates and pigments (phycobilins and astaxanthin). We determined the optimal conditions for DG6 remediation and an artificial neural network (ANN) accurately modeled the experimental data and predicted the concentration of dye as the most and algal turbidity as the least important parameters for DG6 removal efficiency. A DG6 concentration of 60 mg L resulted in the highest simultaneous co-production of PHB (12.7 ± 1.7% DW) and increase of astaxanthin (194%), carotenoids (50%), phenol (51%), carbohydrates (27%) total phycobilin (43%), together with the enhancement of biomass and trichome lengths (95%). Oxidative stress indices and enzyme activities such as peroxidases and laccase (involved in dye removal/antioxidant functions) were also increased by dye dosage. On the basis of our results, we propose that S. platensis may use DG6 dye as a nitrogen/carbon source for co-accumulation of valuable bioplastic and metabolites.
Topics: Azo Compounds; Biomass; Phycobilins; Spirulina; Textiles; Xanthophylls
PubMed: 34162106
DOI: 10.1016/j.chemosphere.2021.130920 -
Angewandte Chemie (International Ed. in... Aug 2021Phytochrome proteins are light receptors that play a pivotal role in regulating the life cycles of plants and microorganisms. Intriguingly, while cyanobacterial...
Phytochrome proteins are light receptors that play a pivotal role in regulating the life cycles of plants and microorganisms. Intriguingly, while cyanobacterial phytochrome Cph1 and cyanobacteriochrome AnPixJ use the same phycocyanobilin (PCB) chromophore to absorb light, their excited-state behavior is very different. We employ multiscale calculations to rationalize the different early photoisomerization mechanisms of PCB in Cph1 and AnPixJ. We found that their electronic S , T , and S potential minima exhibit distinct geometric and electronic structures due to different hydrogen bond networks with the protein environment. These specific interactions influence the S electronic structures along the photoisomerization paths, ultimately leading to internal conversion in Cph1 but intersystem crossing in AnPixJ. This explains why the excited-state relaxation in AnPixJ is much slower (ca. 100 ns) than in Cph1 (ca. 30 ps). Further, we predict that efficient internal conversion in AnPixJ can be achieved upon protonating the carboxylic group that interacts with PCB.
Topics: Bacterial Proteins; Cyanobacteria; Hydrogen Bonding; Molecular Structure; Photochemical Processes; Photoreceptors, Microbial; Phycobilins; Phycocyanin; Phytochrome; Protein Kinases; Stereoisomerism
PubMed: 34097335
DOI: 10.1002/anie.202104853 -
Journal of Biosciences 2021Alzheimer's disease (AD) is a devastating neurodegenerative condition provoking the loss of cognitive and memory performances. Despite huge efforts to develop effective... (Review)
Review
Alzheimer's disease (AD) is a devastating neurodegenerative condition provoking the loss of cognitive and memory performances. Despite huge efforts to develop effective AD therapies, there is still no cure for this neurological condition. Here, we review the main biological properties of Phycocyanobilin (PCB), accounting for its potential uses against AD. PCB, given individually or released in vivo from C-Phycocyanin (C-PC), acts as a bioactive-molecule-mediating antioxidant, is anti-inflammatory and has immunomodulatory activities. PCB/C-PC are able to scavenge reactive oxygen and nitrogen species, to counteract lipid peroxidation and to inhibit enzymes such as NADPH oxidase and COX-2. In animal models of multiple sclerosis and ischemic stroke, these compounds induce remyelination as demonstrated by electron microscopy and the expression of genes such as up-regulation of and down-regulation. These treatments also reduce pro-inflammatory cytokines levels and induce immune suppressive genes. PCB/C-PC protects isolated rat brain mitochondria and inactivate microglia, astrocytes and neuronal apoptosis mediators. Such processes are all involved in the pathogenic cascade of AD, and thus PCB may effectively mitigate the injury in this condition. Furthermore, PCB can be administered safely by oral or parenteral routes and therefore, could be commercially offered as a nutraceutical supplement or as a pharmaceutical drug.
Topics: Alzheimer Disease; Animals; Anti-Inflammatory Agents; Antioxidants; Dietary Supplements; Disease Models, Animal; Gene Expression Regulation; Humans; Immunologic Factors; Microglia; Nerve Tissue Proteins; Oxidative Stress; Phycobilins; Phycocyanin; Rats; Reactive Nitrogen Species; Reactive Oxygen Species; Remyelination
PubMed: 34047285
DOI: No ID Found -
Aquatic Toxicology (Amsterdam,... Jul 2021Selenium, an essential trace element for animals, poses a threat to all forms of life above a threshold concentration. The ubiquitously present cyanobacteria, a major...
Selenium, an essential trace element for animals, poses a threat to all forms of life above a threshold concentration. The ubiquitously present cyanobacteria, a major photosynthetic biotic component of aquatic and other ecosystems, are excellent systems to study the effects of environmental toxicants. The molecular changes that led to beneficial or detrimental effects in response to different doses of selenium oxyanion Se(IV) were analyzed in the filamentous cyanobacterium Anabaena PCC 7120. This organism showed no inhibition in growth up to 15 mg/L sodium selenite, but above this dose i.e. 20-100 mg/L of Se(IV), both growth and photosynthesis were substantially inhibited. Along with the increased accumulation of non-protein thiols, a consistent reduction in levels of ROS was observed at 10 mg/mL dose of Se(IV). High dose of Se(IV) (above 20 mg/L) enhanced endogenous reactive oxygen species (ROS)/lipid peroxidation, and decreased photosynthetic capability. Treatment with 100 mg/L Se(IV) downregulated transcription of several photosynthesis pathways-related genes such as those encoding photosystem I and II proteins, phycobilisome rod-core linker protein, phycocyanobilin, phycoerythrocyanin-associated proteins etc. Interestingly, at a dose range of 10-15 mg/L Se(IV), Anabaena showed an increase in PSII photosynthetic yield and electron transport rate (at PSII), suggesting improved photosynthesis. Se was incorporated into the Anabaena cells, and Se-enriched thylakoid membranes showed higher redox conductivity than the thylakoid membranes from untreated cells. Overall, the data supports that modulation of photosynthetic machinery is one of the crucial mechanisms responsible for the dose-dependent contrasting effect of Se(IV) observed in Anabaena.
Topics: Anabaena; Cnidarian Venoms; Cyanobacteria; Ecosystem; Electron Transport; Oxidation-Reduction; Photosynthesis; Photosystem I Protein Complex; Phycobilins; Phycocyanin; Reactive Oxygen Species; Water Pollutants, Chemical
PubMed: 34015754
DOI: 10.1016/j.aquatox.2021.105839 -
Bioresource Technology Aug 2021The impact of wastewater cultivation was studied on pollutant removal, biomass production, and biosynthesis of high-value metabolites by newly isolated cyanobacteria...
Impact of wastewater cultivation on pollutant removal, biomass production, metabolite biosynthesis, and carbon dioxide fixation of newly isolated cyanobacteria in a multiproduct biorefinery paradigm.
The impact of wastewater cultivation was studied on pollutant removal, biomass production, and biosynthesis of high-value metabolites by newly isolated cyanobacteria namely Acaryochloris marina BERC03, Oscillatoria sp. BERC04, and Pleurocapsa sp. BERC06. During cultivation in urabn wastewater, its pH used to adjust from pH 8.0 to 11, offering contamination-free cultivation, and flotation-based easy harvesting. Besides, wastewater cultivation improved biomass production by 1.3-fold when compared to control along with 3.54-4.2 gL of CO fixation, concomitantly removing suspended organic matter, total nitrogen, and phosphorus by 100%, 53%, and 88%, respectively. Biomass accumulated 26-36% carbohydrates, 15-28% proteins, 38-43% lipids, and 6.3-9.5% phycobilins, where phycobilin yield was improved by 1.6-fold when compared to control. Lipids extracted from the pigment-free biomass were trans-esterified to biodiesel where pigment extraction showed no negative impact on quality of the biodiesel. These strains demonstrated the potential to become feedstock of an integrated biorefinery using urban wastewater as low-cost growth media.
Topics: Biofuels; Biomass; Carbon Dioxide; Cyanobacteria; Environmental Pollutants; Microalgae; Nitrogen; Wastewater
PubMed: 33910117
DOI: 10.1016/j.biortech.2021.125194