The aim and novelty of this paper are found in assessing the influence of inhibitors and antibiotics on intact cell MALDI-TOF mass spectra of the cyanobacterium Synechococcus sp. UPOC S4 and to check the impact on reliability of identification. Defining the limits of this method is important for its use in biology and applied science. The compounds included inhibitors of respiration, glycolysis, citrate cycle, and proteosynthesis. They were used at 1-10 μM concentrations and different periods of up to 3 weeks. Cells were also grown without inhibitors in a microgravity because of expected strong effects. Mass spectra were evaluated using controls and interpreted in terms of differential peaks and their assignment to protein sequences by mass. Antibiotics, azide, and bromopyruvate had the greatest impact. The spectral patterns were markedly altered after a prolonged incubation at higher concentrations, which precluded identification in the database of reference spectra. The incubation in microgravity showed a similar effect. These differences were evident in dendrograms constructed from the spectral data. Enzyme inhibitors affected the spectra to a smaller extent. This study shows that only a long-term presence of antibiotics and strong metabolic inhibitors in the medium at 10-5 M concentrations hinders the correct identification of cyanobacteria by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF).
- MeSH
- antibakteriální látky toxicita MeSH
- antimycin A analogy a deriváty toxicita MeSH
- azidy toxicita MeSH
- buněčné dýchání účinky léků MeSH
- chloramfenikol toxicita MeSH
- citrátový cyklus účinky léků MeSH
- deoxyglukosa toxicita MeSH
- fluoracetáty toxicita MeSH
- glykolýza účinky léků MeSH
- malonáty toxicita MeSH
- proteosyntéza účinky léků MeSH
- pyruváty toxicita MeSH
- reprodukovatelnost výsledků MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice metody MeSH
- stav beztíže MeSH
- streptomycin toxicita MeSH
- Synechococcus chemie účinky léků izolace a purifikace metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
The Black Sea is the largest meromictic sea with a reservoir of anoxic water extending from 100 to 1000 m depth. These deeper layers are characterised by a poorly understood fluorescence signal called "deep red fluorescence", a chlorophyll a- (Chl a) like signal found in deep dark oceanic waters. In two cruises, we repeatedly found up to 103 cells ml-1 of picocyanobacteria at 750 m depth in these waters and isolated two phycoerythrin-rich Synechococcus sp. strains (BS55D and BS56D). Tests on BS56D revealed its high adaptability, involving the accumulation of Chl a in anoxic/dark conditions and its capacity to photosynthesise when re-exposed to light. Whole-genome sequencing of the two strains showed the presence of genes that confirms the putative ability of our strains to survive in harsh mesopelagic environments. This discovery provides new evidence to support early speculations associating the "deep red fluorescence" signal to viable picocyanobacteria populations in the deep oxygen-depleted oceans, suggesting a reconsideration of the ecological role of a viable stock of Synechococcus in dark deep waters.
- MeSH
- chlorofyl a metabolismus MeSH
- ekosystém MeSH
- fluorescence MeSH
- fotosyntéza MeSH
- fykoerythrin metabolismus MeSH
- fylogeneze MeSH
- genom bakteriální MeSH
- oceány a moře MeSH
- Synechococcus chemie klasifikace izolace a purifikace metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Černé moře MeSH
- oceány a moře MeSH
Photosystem II (PSII) catalyses the photoinduced oxygen evolution and, by producing reducing equivalents drives, in concert with PSI, the conversion of carbon dioxide to sugars. Our knowledge about the architecture of the reaction centre (RC) complex and the mechanisms of charge separation and stabilisation is well advanced. However, our understanding of the processes associated with the functioning of RC is incomplete: the photochemical activity of PSII is routinely monitored by chlorophyll-a fluorescence induction but the presently available data are not free of controversy. In this work, we examined the nature of gradual fluorescence rise of PSII elicited by trains of single-turnover saturating flashes (STSFs) in the presence of a PSII inhibitor, permitting only one stable charge separation. We show that a substantial part of the fluorescence rise originates from light-induced processes that occur after the stabilisation of charge separation, induced by the first STSF; the temperature-dependent relaxation characteristics suggest the involvement of conformational changes in the additional rise. In experiments using double flashes with variable waiting times (∆τ) between them, we found that no rise could be induced with zero or short ∆τ, the value of which depended on the temperature - revealing a previously unknown rate-limiting step in PSII.
- MeSH
- chlorofyl a metabolismus MeSH
- fluorescence * MeSH
- fotosyntéza MeSH
- fotosystém II (proteinový komplex) antagonisté a inhibitory metabolismus MeSH
- Spinacia oleracea metabolismus MeSH
- Synechococcus metabolismus MeSH
- Synechocystis metabolismus MeSH
- teplota MeSH
- tylakoidy metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The assimilation of N-NO3- requires more energy than that of N-NH4+ . This becomes relevant when energy is limiting and may impinge differently on cell energy budget depending on depth, time of the day and season. We hypothesize that N-limited and energy-limited cells of the oceanic cyanobacterium Synechococcus sp. differ in their response to the N source with respect to growth, elemental stoichiometry and carbon allocation. Under N limitation, cells retained almost absolute homeostasis of elemental and organic composition, and the use of NH4+ did not stimulate growth. When energy was limiting, however, Synechococcus grew faster in NH4+ than in NO3- and had higher C (20%), N (38%) and S (30%) cell quotas. Furthermore, more C was allocated to protein, whereas the carbohydrate and lipid pool size did not change appreciably. Energy limitation also led to a higher photosynthetic rate relative to N limitation. We interpret these results as an indication that, under energy limitation, the use of the least expensive N source allowed a spillover of the energy saved from N assimilation to the assimilation of other nutrients. The change in elemental stoichiometry influenced C allocation, inducing an increase in cell protein, which resulted in a stimulation of photosynthesis and growth.
- MeSH
- adenosintrifosfát metabolismus MeSH
- amoniové sloučeniny farmakologie MeSH
- bakteriální proteiny metabolismus MeSH
- biomasa MeSH
- dusičnany farmakologie MeSH
- dusík metabolismus MeSH
- energetický metabolismus * účinky léků MeSH
- fosfor metabolismus MeSH
- fotosyntéza účinky léků MeSH
- kyslík metabolismus MeSH
- lipidy analýza MeSH
- sacharidy analýza MeSH
- síra metabolismus MeSH
- Synechococcus cytologie účinky léků růst a vývoj metabolismus MeSH
- uhlík metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
The carbon-concentrating mechanisms (CCMs) of cyanobacteria counteract the low CO2 affinity and CO2:O2 selectivities of the Rubisco of these photolithotrophs and the relatively low oceanic CO2 availability. CCMs have a significant energy cost; if light is limiting, the use of N sources whose assimilation demands less energy could permit a greater investment of energy into CCMs and inorganic C (Ci) assimilation. To test this, we cultured Synechococcus sp. UTEX LB 2380 under either N or energy limitation, in the presence of NO3- or NH4+. When growth was energy-limited, NH4+-grown cells had a 1.2-fold higher growth rate, 1.3-fold higher dissolved inorganic carbon (DIC)-saturated photosynthetic rate, 19% higher linear electron transfer, 80% higher photosynthetic 1/K1/2(DIC), 2.0-fold greater slope of the linear part of the photosynthesis versus DIC curve, 3.5-fold larger intracellular Ci pool, and 2.3-fold higher Zn quota than NO3--grown cells. When energy was not limiting growth, there were not differences between NH4+- and NO3--grown cells, except for higher linear electron transfer and larger intracellular Ci pool.We conclude that, when energy limits growth, cells that use the cheaper N source divert energy from N assimilation to C acquisition and assimilation; this does not happen when energy is not limiting.
The structure of monomeric and trimeric photosystem I (PS I) of Thermosynechococcus elongatus BP1 (T. elongatus) was investigated by small-angle X-ray scattering (SAXS). The scattering data reveal that the protein-detergent complexes possess radii of gyration of 58 and 78 Å in the cases of monomeric and trimeric PS I, respectively. The results also show that the samples are monodisperse, virtually free of aggregation, and contain empty detergent micelles. The shape of the protein-detergent complexes can be well approximated by elliptical cylinders with a height of 78 Å. Monomeric PS I in buffer solution exhibits minor and major radii of the elliptical cylinder of about 50 and 85 Å, respectively. In the case of trimeric PS I, both radii are equal to about 110 Å. The latter model can be shown to accommodate three elliptical cylinders equal to those describing monomeric PS I. A structure reconstitution also reveals that the protein-detergent complexes are larger than their respective crystal structures. The reconstituted structures are larger by about 20 Å mainly in the region of the hydrophobic surfaces of the monomeric and trimeric PS I complexes. This seeming contradiction can be resolved by the addition of a detergent belt constituted by a monolayer of dodecyl-β-D-maltoside molecules. Assuming a closest possible packing, a number of roughly 1024 and 1472 detergent molecules can be determined for monomeric and trimeric PS I, respectively. Taking the monolayer of detergent molecules into account, the solution structure can be almost perfectly modeled by the crystal structures of monomeric and trimeric PS I.
- MeSH
- bakteriální proteiny chemie MeSH
- detergenty chemie MeSH
- difrakce rentgenového záření * MeSH
- fotosystém I (proteinový komplex) chemie metabolismus MeSH
- maloúhlový rozptyl * MeSH
- molekulární modely MeSH
- multimerizace proteinu * MeSH
- roztoky MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice MeSH
- Synechococcus metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
Cyanobacteria Synechococcus sp. PCC 7942 and Synechocystis sp. PCC 6803 show similar changes in the metabolic response to changed CO2 conditions but exhibit significant differences at the transcriptomic level. This study employs a systems biology approach to investigate the difference in metabolic regulation of Synechococcus sp. PCC 7942 and Synechocystis sp. PCC 6803. Presented multi-level kinetic model for Synechocystis sp. PCC 6803 is a new approach integrating and analysing metabolomic, transcriptomic and fluxomics data obtained under high and ambient CO2 levels. Modelling analysis revealed that higher number of different isozymes in Synechocystis 6803 improves homeostatic stability of several metabolites, especially 3PGA by 275%, against changes in gene expression, compared to Synechococcus sp. PCC 7942. Furthermore, both cyanobacteria have the same amount of phosphoglycerate mutases but Synechocystis 6803 exhibits only ~20% differences in their mRNA levels after shifts from high to ambient CO2 level, in comparison to ~500% differences in the case of Synechococcus sp. PCC 7942. These and other data imply that the biochemical control dominates over transcriptional regulation in Synechocystis 6803 to acclimate central carbon metabolism in the environment of variable inorganic carbon availability without extra cost carried by large changes in the proteome.
- MeSH
- metabolismus MeSH
- metabolomika MeSH
- oxid uhličitý metabolismus MeSH
- regulace genové exprese enzymů * MeSH
- regulace genové exprese u bakterií * MeSH
- stanovení celkové genové exprese MeSH
- Synechococcus genetika metabolismus MeSH
- Synechocystis genetika metabolismus MeSH
- systémová biologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
When photosystem II (PSII) is exposed to excess light, singlet oxygen ((1)O(2)) formed by the interaction of molecular oxygen with triplet chlorophyll. Triplet chlorophyll is formed by the charge recombination of triplet radical pair (3)[P680(•+)Pheo(•-)] in the acceptor-side photoinhibition of PSII. Here, we provide evidence on the formation of (1)O(2) in the donor side photoinhibition of PSII. Light-induced (1)O(2) production in Tris-treated PSII membranes was studied by electron paramagnetic resonance (EPR) spin-trapping spectroscopy, as monitored by TEMPONE EPR signal. Light-induced formation of carbon-centered radicals (R(•)) was observed by POBN-R adduct EPR signal. Increased oxidation of organic molecules at high pH enhanced the formation of TEMPONE and POBN-R adduct EPR signals in Tris-treated PSII membranes. Interestingly, the scavenging of R(•) by propyl gallate significantly suppressed (1)O(2). Based on our results, it is concluded that (1)O(2) formation correlates with R(•) formation on the donor side of PSII due to oxidation of organic molecules (lipids and proteins) by long-lived P680(•+)/TyrZ(•). It is proposed here that the Russell mechanism for the recombination of two peroxyl radicals formed by the interaction of R(•) with molecular oxygen is a plausible mechanism for (1)O(2) formation in the donor side photoinhibition of PSII.
- MeSH
- chemické modely MeSH
- elektronová paramagnetická rezonance metody MeSH
- fotochemie metody MeSH
- fotosystém II (proteinový komplex) fyziologie MeSH
- koncentrace vodíkových iontů MeSH
- kyslík chemie MeSH
- propylgalan chemie MeSH
- singletový kyslík * MeSH
- spin trapping metody MeSH
- Spinacia oleracea MeSH
- světlo MeSH
- Synechococcus metabolismus MeSH
- uhlík chemie MeSH
- volné radikály MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
