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Molecular & Cellular Proteomics : MCP Apr 2022The filamentous cyanobacterium Anabaena sp. PCC 7120 can differentiate into heterocysts to fix atmospheric nitrogen. During cell differentiation, cellular morphology and...
The filamentous cyanobacterium Anabaena sp. PCC 7120 can differentiate into heterocysts to fix atmospheric nitrogen. During cell differentiation, cellular morphology and gene expression undergo a series of significant changes. To uncover the mechanisms responsible for these alterations, we built protein-protein interaction (PPI) networks for these two cell types by cofractionation coupled with mass spectrometry. We predicted 280 and 215 protein complexes, with 6322 and 2791 high-confidence PPIs in vegetative cells and heterocysts, respectively. Most of the proteins in both types of cells presented similar elution profiles, whereas the elution peaks of 438 proteins showed significant changes. We observed that some well-known complexes recruited new members in heterocysts, such as ribosomes, diflavin flavoprotein, and cytochrome c oxidase. Photosynthetic complexes, including photosystem I, photosystem II, and phycobilisome, remained in both vegetative cells and heterocysts for electron transfer and energy generation. Besides that, PPI data also reveal new functions of proteins. For example, the hypothetical protein Alr4359 was found to interact with FraH and Alr4119 in heterocysts and was located on heterocyst poles, thereby influencing the diazotrophic growth of filaments. The overexpression of Alr4359 suspended heterocyst formation and altered the pigment composition and filament length. This work demonstrates the differences in protein assemblies and provides insight into physiological regulation during cell differentiation.
Topics: Anabaena; Bacterial Proteins; Biology; Cell Differentiation; Gene Expression Regulation, Bacterial
PubMed: 35288331
DOI: 10.1016/j.mcpro.2022.100224 -
Proceedings of the National Academy of... Nov 2020Cyanobacteriochromes (CBCRs) are photoswitchable linear tetrapyrrole (bilin)-based light sensors in the phytochrome superfamily with a broad spectral range from the near...
Cyanobacteriochromes (CBCRs) are photoswitchable linear tetrapyrrole (bilin)-based light sensors in the phytochrome superfamily with a broad spectral range from the near UV through the far red (330 to 760 nm). The recent discovery of far-red absorbing CBCRs (frCBCRs) has garnered considerable interest from the optogenetic and imaging communities because of the deep penetrance of far-red light into mammalian tissue and the small size of the CBCR protein scaffold. The present studies were undertaken to determine the structural basis for far-red absorption by JSC1_58120g3, a frCBCR from the thermophilic cyanobacterium sp. JSC-1 that is a representative member of a phylogenetically distinct class. Unlike most CBCRs that bind phycocyanobilin (PCB), a phycobilin naturally occurring in cyanobacteria and only a few eukaryotic phototrophs, JSC1_58120g3's far-red absorption arises from incorporation of the PCB biosynthetic intermediate 18,18-dihydrobiliverdin (18,18-DHBV) rather than the more reduced and more abundant PCB. JSC1_58120g3 can also yield a far-red-absorbing adduct with the more widespread linear tetrapyrrole biliverdin IXα (BV), thus circumventing the need to coproduce or supplement optogenetic cell lines with PCB. Using high-resolution X-ray crystal structures of 18,18-DHBV and BV adducts of JSC1_58120g3 along with structure-guided mutagenesis, we have defined residues critical for its verdin-binding preference and far-red absorption. Far-red sensing and verdin incorporation make this frCBCR lineage an attractive template for developing robust optogenetic and imaging reagents for deep tissue applications.
Topics: Bacterial Proteins; Biliverdine; Cyanobacteria; Light; Photoreceptor Cells; Photoreceptors, Microbial; Phycobilins; Phycocyanin; Phytochrome; Porphyrins
PubMed: 33106421
DOI: 10.1073/pnas.2016047117 -
Frontiers in Plant Science 2023Phycobilisomes serve as a light-harvesting antenna of both photosystem I (PSI) and II (PSII) in cyanobacteria, yet direct energy transfer from phycobilisomes to PSI is...
Phycobilisomes serve as a light-harvesting antenna of both photosystem I (PSI) and II (PSII) in cyanobacteria, yet direct energy transfer from phycobilisomes to PSI is not well documented. Here we recorded picosecond time-resolved fluorescence at wavelengths of 605-760 nm in isolated photosystem I (PSI), phycobilisomes and intact cells of a PSII-deficient mutant of sp. PCC 6803 at 77 K to study excitation energy transfer and trapping. By means of a simultaneous target analysis of the kinetics of isolated complexes and whole cells, the pathways and dynamics of energy transfer and were established. We establish that the timescale of the slowest equilibration between different terminal emitters in the phycobilisome is ≈800 ps. It was estimated that the terminal emitter in about 40% of the phycobilisomes transfers its energy with a rate constant of 42 ns to PSI. This energy transfer rate is higher than the rates of equilibration within the phycobilisome - between the rods and the core or between the core cylinders - and is evidence for the existence of specific phycobilisome-PSI interactions. The rest of the phycobilisomes remain unconnected or slowly transferring energy to PSI.
PubMed: 38078099
DOI: 10.3389/fpls.2023.1293813 -
Biomacromolecules Dec 2021Environment-triggered protein conformational changes have garnered wide interest in both fundamental research, for deciphering in vivo acclimatory responses, and...
Environment-triggered protein conformational changes have garnered wide interest in both fundamental research, for deciphering in vivo acclimatory responses, and practical applications, for designing stimuli-responsive probes. Here, we propose a protein-chromophore regulatory mechanism that allows for manipulation of C-phycocyanin (C-PC) from by environmental pH and UV irradiation. Using small-angle X-ray scattering, a pH-mediated C-PC assembly-disassembly pathway, from monomers to nonamers, was unraveled. Such flexible protein matrices impart tunability to the embedded tetrapyrroles, whose photochemical behaviors were found to be modulated by protein assembly states. UV irradiation on C-PC triggers pH-dependent singlet oxygen (O) generation and conformational changes. Intermolecular photo-crosslinking occurs at pH 5.0 via dityrosine species, which bridges solution-based C-PC oligomers into unprecedented dodecamers and 24-mers. These supramolecular assemblies impart C-PC at pH 5.0, which significantly enhanced O yield, fluorescence, and photostability relative to those at other pH values, a finding that makes C-PC appealing for tumor-targeted photodynamic therapy.
Topics: Hydrogen-Ion Concentration; Photochemotherapy; Phycocyanin
PubMed: 34767353
DOI: 10.1021/acs.biomac.1c01095 -
Plant Phenomics (Washington, D.C.) 2023Phycobilisomes and chlorophyll-a () play important roles in the photosynthetic physiology of red macroalgae and serve as the primary light-harvesting antennae and...
Phycobilisomes and chlorophyll-a () play important roles in the photosynthetic physiology of red macroalgae and serve as the primary light-harvesting antennae and reaction center for photosystem II. is an economically important red macroalga widely cultivated in East Asian countries. The contents and ratios of 3 main phycobiliproteins and are visible traits to evaluate its commercial quality. The traditional analytical methods used for measuring these components have several limitations. Therefore, a high-throughput, nondestructive, optical method based on hyperspectral imaging technology was developed for phenotyping the pigments phycoerythrin (PE), phycocyanin (PC), allophycocyanin (APC), and in thalli in this study. The average spectra from the region of interest were collected at wavelengths ranging from 400 to 1000 nm using a hyperspectral camera. Following different preprocessing methods, 2 machine learning methods, partial least squares regression (PLSR) and support vector machine regression (SVR), were performed to establish the best prediction models for PE, PC, APC, and contents. The prediction results showed that the PLSR model performed the best for PE ( = 0.96, MAPE = 8.31%, RPD = 5.21) and the SVR model performed the best for PC ( = 0.94, MAPE = 7.18%, RPD = 4.16) and APC ( = 0.84, MAPE = 18.25%, RPD = 2.53). Two models (PLSR and SVR) performed almost the same for (PLSR: = 0.92, MAPE = 12.77%, RPD = 3.61; SVR: = 0.93, MAPE = 13.51%, RPD =3.60). Further validation of the optimal models was performed using field-collected samples, and the result demonstrated satisfactory robustness and accuracy. The distribution of PE, PC, APC, and contents within a thallus was visualized according to the optimal prediction models. The results showed that hyperspectral imaging technology was effective for fast, accurate, and noninvasive phenotyping of the PE, PC, APC, and contents of in situ. This could benefit the efficiency of macroalgae breeding, phenomics research, and other related applications.
PubMed: 37040513
DOI: 10.34133/plantphenomics.0012 -
F1000Research 2021contains several bioactive molecules such as phenol, flavonoid and phycocyanin pigments. This study unveils total phenol, flavonoid, antioxidant activity,...
contains several bioactive molecules such as phenol, flavonoid and phycocyanin pigments. This study unveils total phenol, flavonoid, antioxidant activity, phycocyanin content and evaluated encapsulation efficiency from intervention on . intervention aims to reduce unpleasant odors from that will increase consumption and increase bioactive compounds. The intervention was carried out by soaking a control sample (SP) in with a ratio of 1:4 (w/v) and it was then dried (DSB) and microencapsulated by freeze drying methods (MSB) using a combination of maltodextrin and gelatin. Total flavonoid and phenolic analysis with curve fitting analysis used a linear regression approach. Antioxidant activity of samples was analysed with the 2,2'-azino-bis-3-3thylbenzthiazoline-6-sulphonic acid (ABTS) method. Data were analysed using ANOVA at significance level (p < 0.05) followed by Tukey test models using SPSS v.22. The result of this study indicated that intervention treatment (DSB) has the potential to increase bioactive compounds such as total phenol, antioxidant activity and phycocyanin, and flavonoid content. Intervention of on (DSB) significantly increases total phenol by 49.5% and phycocyanin by 40.7%. This is due to the phenol and azulene compounds in which have a synergistic effect on phenol and phycocyanin in . Microencapsulation using a maltodexrin and gelatin coating is effective in phycocyanin protection and antioxidant activity with an encapsulation efficiency value of 71.58% and 80.5%. The intervention of on improved the total phenol and phycocyanin content and there is potential for a pharmaceutical product for a functional food and pharmaceutical product.
Topics: Antioxidants; Flavonoids; Gelatin; Ocimum basilicum; Phenol; Phenols; Phycocyanin; Powders; Spirulina
PubMed: 35083034
DOI: 10.12688/f1000research.52394.3 -
Metabolic Engineering Communications Dec 2021Phycocyanin (PC) is a soluble phycobiliprotein found within the light-harvesting phycobilisome complex of cyanobacteria and red algae, and is considered a high-value...
Phycocyanin (PC) is a soluble phycobiliprotein found within the light-harvesting phycobilisome complex of cyanobacteria and red algae, and is considered a high-value product due to its brilliant blue colour and fluorescent properties. However, commercially available PC has a relatively low temperature stability. Thermophilic species produce more thermostable variants of PC, but are challenging and energetically expensive to cultivate. Here, we show that the PC operon from the thermophilic cyanobacterium BP-1 () is functional in the mesophile sp. PCC 6803. Expression of in an 'Olive' mutant strain of lacking endogenous PC resulted in high yields of thermostable PC (112 ± 1 mg g DW) comparable to that of endogenous PC in wild-type cells. Heterologous PC also improved the growth of the Olive mutant, which was further supported by evidence of a functional interaction with the endogenous allophycocyanin core of the phycobilisome complex. The thermostability properties of the heterologous PC were comparable to those of PC from , and could be purified from the Olive mutant using a low-cost heat treatment method. Finally, we developed a scalable model to calculate the energetic benefits of producing PC from in cultures. Our model showed that the higher yields and lower cultivation temperatures of resulted in a 3.5-fold increase in energy efficiency compared to , indicating that producing thermostable PC in non-native hosts is a cost-effective strategy for scaling to commercial production.
PubMed: 34168957
DOI: 10.1016/j.mec.2021.e00175 -
Journal of the American Chemical Society May 2023The phycobilisome is the primary light-harvesting antenna in cyanobacterial and red algal oxygenic photosynthesis. It maintains near-unity efficiency of energy transfer...
The phycobilisome is the primary light-harvesting antenna in cyanobacterial and red algal oxygenic photosynthesis. It maintains near-unity efficiency of energy transfer to reaction centers despite relying on slow exciton hopping along a relatively sparse network of highly fluorescent phycobilin chromophores. How the complex maintains this high efficiency remains unexplained. Using a two-dimensional electronic spectroscopy polarization scheme that enhances energy transfer features, we directly watch energy flow in the phycobilisome complex of sp. PCC 6803 from the outer phycocyanin rods to the allophycocyanin core. The observed downhill flow of energy, previously hidden within congested spectra, is faster than timescales predicted by Förster hopping along single rod chromophores. We attribute the fast, 8 ps energy transfer to interactions between rod-core linker proteins and terminal rod chromophores, which facilitate unidirectionally downhill energy flow to the core. This mechanism drives the high energy transfer efficiency in the phycobilisome and suggests that linker protein-chromophore interactions have likely evolved to shape its energetic landscape.
Topics: Phycobilisomes; Photosynthesis; Energy Transfer; Synechocystis
PubMed: 37200045
DOI: 10.1021/jacs.3c01799 -
Nature Communications Mar 2023Cryo-electron tomography is a major tool used to study the structure of protein complexes in situ. However, the throughput of tilt-series image data collection is still...
Cryo-electron tomography is a major tool used to study the structure of protein complexes in situ. However, the throughput of tilt-series image data collection is still quite low. Here, we show that GisSPA, a GPU accelerated program, can translationally and rotationally localize the target protein complex in cellular lamellae, as prepared with a focused ion beam, using single cryo-electron microscopy images without tilt-series, and reconstruct the protein complex at near-atomic resolution. GisSPA allows high-throughput data collection without the acquisition of tilt-series images and reconstruction of the tomogram, which is essential for high-resolution reconstruction of asymmetric or low-symmetry protein complexes. We demonstrate the power of GisSPA with 3.4-Å and 3.9-Å resolutions of resolving phycobilisome and tetrameric photosystem II complex structures in cellular lamellae, respectively. In this work, we present GisSPA as a practical tool that facilitates high-resolution in situ protein structure determination.
Topics: Cryoelectron Microscopy; Electron Microscope Tomography; Image Processing, Computer-Assisted
PubMed: 36922493
DOI: 10.1038/s41467-023-36175-y -
Proceedings of the National Academy of... Apr 2023Terrestrial ecosystems and human societies depend on oxygenic photosynthesis, which began to reshape our atmosphere approximately 2.5 billion years ago. The earliest...
Terrestrial ecosystems and human societies depend on oxygenic photosynthesis, which began to reshape our atmosphere approximately 2.5 billion years ago. The earliest known organisms carrying out oxygenic photosynthesis are the cyanobacteria, which use large complexes of phycobiliproteins as light-harvesting antennae. Phycobiliproteins rely on phycocyanobilin (PCB), a linear tetrapyrrole (bilin) chromophore, as the light-harvesting pigment that transfers absorbed light energy from phycobilisomes to the chlorophyll-based photosynthetic apparatus. Cyanobacteria synthesize PCB from heme in two steps: A heme oxygenase converts heme into biliverdin IXα (BV), and the ferredoxin-dependent bilin reductase (FDBR) PcyA then converts BV into PCB. In the current work, we examine the origins of this pathway. We demonstrate that PcyA evolved from pre-PcyA proteins found in nonphotosynthetic bacteria and that pre-PcyA enzymes are active FDBRs that do not yield PCB. Pre-PcyA genes are associated with two gene clusters. Both clusters encode bilin-binding globin proteins, phycobiliprotein paralogs that we designate as BBAGs (bilin biosynthesis-associated globins). Some cyanobacteria also contain one such gene cluster, including a BBAG, two V4R proteins, and an iron-sulfur protein. Phylogenetic analysis shows that this cluster is descended from those associated with pre-PcyA proteins and that light-harvesting phycobiliproteins are also descended from BBAGs found in other bacteria. We propose that PcyA and phycobiliproteins originated in heterotrophic, nonphotosynthetic bacteria and were subsequently acquired by cyanobacteria.
Topics: Humans; Phylogeny; Phycobiliproteins; Oxidoreductases; Ecosystem; Bile Pigments; Cyanobacteria
PubMed: 37071675
DOI: 10.1073/pnas.2300770120