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31.
Chlorophyll fluorescence is routinely taken as a quantifiable measure of the redox state of the primary quinone acceptor QA of PSII. The variable fluorescence in thylakoids increases in a single turnover flash (STF) from its low dark level F
o towards a maximum F
mSTF when QA becomes reduced. We found, using twin single turnover flashes (TTFs) that the fluorescence increase induced by the first
twin-partner is followed by a 20–30% increase when the second partner is applied within 20–100 μs after the first one. The
amplitude of the twin response shows a period-of-four oscillation associated with the 4-step oxidation of water in the Kok
cycle (S states) and originates from two different trapped states with a life time of 0.2–0.4 and 2–5 ms, respectively. The
oscillation is supplemented with a binary oscillation associated with the two-electron gate mechanism at the PSII acceptor
side. The F(t) response in high frequency flash trains (1–4 kHz) shows (i) in the first 3–4 flashes a transient overshoot 20–30% above
the F
mSTF = 3*F
o level reached in the 1st flash with a partial decline towards a dip D in the next 2–3 ms, independent of the flash frequency,
and (ii) a frequency independent rise to F
m = 5*F
o in the 3–60 ms time range. The initial overshoot is interpreted to be due to electron trapping in the S0 fraction with QB-nonreducing centers and the dip to the subsequent recovery accompanying the reoxidation of the double reduced acceptor pair
in these RCs after trapping. The rise after the overshoot is, in agreement with earlier findings, interpreted to indicate
a photo-electrochemical control of the chlorophyll fluorescence yield of PSII. It is anticipated that the double exciton and
electron trapping property of PSII is advantageous for the plant. It serves to alleviate the depression of electron transport
in single reduced QB-nonreducing RCs, associated with electrochemically coupled proton transport, by an increased electron trapping efficiency
in these centers. 相似文献
32.
Devaki A. Kelkar 《生物化学与生物物理学报:生物膜》2007,1768(9):2011-2025
The linear peptide gramicidin forms prototypical ion channels specific for monovalent cations and has been extensively used to study the organization, dynamics and function of membrane-spanning channels. In recent times, the availability of crystal structures of complex ion channels has challenged the role of gramicidin as a model membrane protein and ion channel. This review focuses on the suitability of gramicidin as a model membrane protein in general, and the information gained from gramicidin to understand lipid-protein interactions in particular. Special emphasis is given to the role and orientation of tryptophan residues in channel structure and function and recent spectroscopic approaches that have highlighted the organization and dynamics of the channel in membrane and membrane-mimetic media. 相似文献
33.
Mark Pottek Gabriel C. Knop Reto Weiler Karin Dedek 《Journal of visualized experiments : JoVE》2011,(57)
Studying the physiological properties and synaptic connections of specific neurons in the intact tissue is a challenge for those cells that lack conspicuous morphological features or show a low population density. This applies particularly to retinal amacrine cells, an exceptionally multiform class of interneurons that comprise roughly 30 subtypes in mammals1. Though being a crucial part of the visual processing by shaping the retinal output2, most of these subtypes have not been studied up to now in a functional context because encountering these cells with a recording electrode is a rare event.Recently, a multitude of transgenic mouse lines is available that express fluorescent markers like green fluorescent protein (GFP) under the control of promoters for membrane receptors or enzymes that are specific to only a subset of neurons in a given tissue3,4. These pre-labeled cells are therefore accessible to directed microelectrode targeting under microscopic control, permitting the systematic study of their physiological properties in situ. However, excitation of fluorescent markers is accompanied by the risk of phototoxicity for the living tissue. In the retina, this approach is additionally hampered by the problem that excitation light causes appropriate stimulation of the photoreceptors, thus inflicting photopigment bleaching and transferring the retinal circuits into a light-adapted condition. These drawbacks are overcome by using infrared excitation delivered by a mode-locked laser in short pulses of the femtosecond range. Two-photon excitation provides energy sufficient for fluorophore excitation and at the same time restricts the excitation to a small tissue volume minimizing the hazards of photodamage5. Also, it leaves the retina responsive to visual stimuli since infrared light (>850 nm) is only poorly absorbed by photopigments6.In this article we demonstrate the use of a transgenic mouse retina to attain electrophysiological in situ recordings from GFP-expressing cells that are visually targeted by two-photon excitation. The retina is prepared and maintained in darkness and can be subjected to optical stimuli which are projected through the condenser of the microscope (Figure 1). Patch-clamp recording of light responses can be combined with dye filling to reveal the morphology and to check for gap junction-mediated dye coupling to neighboring cells, so that the target cell can by studied on different experimental levels. 相似文献
34.
S.?E.?Jorge?Villar H.?G.?M.?EdwardsEmail author M.?R.?Worland 《Origins of life and evolution of the biosphere》2005,35(5):489-506
Raman spectra have been obtained for extremophiles from several geological environments; selected examples have been taken
from hot and cold deserts comprising psychrophiles, thermophiles and halophiles. The purpose of this study is the assessment
of the effect of the wavelength of the laser excitation on the ability to determine unique information from the Raman spectra
about the specificity of detection of biomolecules produced as a result of the survival strategies adopted by organisms in
extreme terrestrial environments. It was concluded that whereas FT-Raman spectroscopy at 1064 nm gave good quality results
the time required to record the data was relatively large compared with other wavelengths of excitation but that better access
to the CH stretching region for organic molecules was given. Shorter wavelength excitation of biomolecules in the blue-green
regions of the visible spectrum using a conventional dispersive spectrometer was more rapid but very dependent upon the type
of chemical compound being studied; most relevant biomolecules fluoresced at these wavelengths but carotenoids exhibited a
resonance effect which resulted in an improved detection capability. Minerals and geological materials, in contrast, were
best studied at these visible wavelengths. In general, the best compromise system for the excitation of the Raman spectra
of both geological and biological materials was provided using a 785 nm laser coupled with a dispersive spectrometer, especially
for accessing the 1800–200 cm−1 wavenumber shift region where much of the definitive analytical information resides. This work will have conclusions relevant
to the use of miniaturised Raman spectrometers for the detection of biomolecules in extraterrestrial planetary exploration. 相似文献
35.
This review centers on the structural and functional organization of the light-harvesting system in the peripheral antenna of Photosystem I (LHC I) and its energy coupling to the Photosystem I (PS I) core antenna network in view of recently available structural models of the eukaryotic Photosystem I–LHC I complex, eukaryotic LHC II complexes and the cyanobacterial Photosystem I core. A structural model based on the 3D homology of Lhca4 with LHC II is used for analysis of the principles of pigment arrangement in the LHC I peripheral antenna, for prediction of the protein ligands for the pigments that are unique for LHC I and for estimates of the excitonic coupling in strongly interacting pigment dimers. The presence of chlorophyll clusters with strong pigment–pigment interactions is a structural feature of PS I, resulting in the characteristic red-shifted fluorescence. Analysis of the interactions between the PS I core antenna and the peripheral antenna leads to the suggestion that the specific function of the red pigments is likely to be determined by their localization with respect to the reaction center. In the PS I core antenna, the Chl clusters with a different magnitude of low energy shift contribute to better spectral overlap of Chls in the reaction center and the Chls of the antenna network, concentrate the excitation around the reaction center and participate in downhill enhancement of energy transfer from LHC II to the PS I core. Chlorophyll clusters forming terminal emitters in LHC I are likely to be involved in photoprotection against excess energy. 相似文献
36.
Rubin JE 《Journal of computational neuroscience》2005,18(3):333-342
Typically, excitatory synaptic coupling is thought of as an influence that accelerates and propagates firing in neuronal networks. This paper reviews recent results explaining how, contrary to these expectations, the presence of excitatory synaptic coupling can drastically slow oscillations in a network and how localized, sustained activity can arise in a network with purely excitatory coupling, without sustained inputs. These two effects stem from interactions of excitatory coupling with two different forms of intrinsic neuronal dynamics, and both serve to highlight the fact that the influence of synaptic coupling in a network depends strongly on the intrinsic properties of cells in the network.This work was partially supported by the National Science Foundation, under award DMS-0414023 相似文献
37.
Nicotinamide cofactors play numerous roles in cellular metabolic and biosynthetic reactions and intracellular signaling events. Recently, nicotinamide cofactors have been implicated in the function of cellular biological clocks. To gain insight into the possible roles of nicotinamide cofactors in complex time-related events, we have developed a rapid and sensitive method for extraction of NAD(P)(H) from cultured cells, separation of analytes by capillary electrophoresis, and detection by multiphoton excitation of fluorescence. Extraction and quantitation steps have been systematically characterized for optimal pH, detergent, temperature, sonication, filtration, efficiency, accuracy, and reproducibility. The method is suitable for extractions at 2- to 3-h intervals over 1 day or more or as frequently as every hour for shorter durations. Natively fluorescent NAD(P)H are assayed directly, and nonfluorescent NAD(P) are enzymatically reduced to their fluorescent counterparts before analysis. The method yields accurate values for cellular NADP, NADPH, and total NAD(H) levels and relative information on cellular NADH concentration; modification of the procedure allows full quantitation of all relevant species. We conclude that these assays are more suitable than any yet published for tracking variations in nicotinamide cofactor levels over periods of 1 day or more. 相似文献
38.
Reaction centers (RCs) of purple bacteria are uniquely suited objects to study the mechanisms of the photosynthetic conversion of light energy into chemical energy. A recently introduced method of higher order derivative spectroscopy [I.K. Mikhailyuk, H. Lokstein, A.P. Razjivin, A method of spectral subband decomposition by simultaneous fitting the initial spectrum and a set of its derivatives, J. Biochem. Biophys. Methods 63 (2005) 10-23] was used to analyze the NIR absorption spectra of RC preparations from Rhodobacter (R.) sphaeroides strain 2R and Blastochloris (B.) viridis strain KH, containing bacteriochlorophyll (BChl) a and b, respectively. Q(y) bands of individual RC porphyrin components (BChls and bacteriopheophytins, BPheo) were identified. The results indicate that the upper exciton level P(y+) of the photo-active BChl dimer in RCs of R. sphaeroides has an absorption maximum of 810nm. The blue shift of a complex integral band at approximately 800nm upon oxidation of the RC is caused primarily by bleaching of P(y+), rather than by an electrochromic shift of the absorption band(s) of the monomeric BChls. Likewise, the disappearance of a band peaking at 842nm upon oxidation of RCs from B. viridis indicates that this band has to be assigned to P(y+). A blue shift of an absorption band at approximately 830nm upon oxidation of RCs of B. viridis is also essentially caused by the disappearance of P(y+), rather than by an electrochromic shift of the absorption bands of monomeric BChls. Absorption maxima of the monomeric BChls, B(B) and B(A) are at 802 and 797nm, respectively, in RCs of R. sphaeroides at room temperature. BPheo co-factors H(B) and H(A) peak at 748 and 758nm, respectively, at room temperature. For B. viridis RCs the spectral positions of H(B) and H(A) were found to be 796 and 816nm, respectively, at room temperature. 相似文献
39.
Slipped frameshift intermediates can occur when DNA polymerase slows or stalls at sites of DNA lesions. However, this phenomenon is much less common when unmodified DNA is replicated. In order to study the effect of templating bases on the alignment of primer-templates, NMR structural investigation has been performed on primer-template oligonucleotide models which mimic the situation that dNTP has just been incorporated opposite template. NMR evidence reveals the occurrence of misalignment when dGTP is incorporated opposite template T with a downstream nucleotide C. Depending on the template sequence, further extension of the primer can lead to realignment. 相似文献
40.
TÜNDE TÓTH OTTÓ ZSIROS MIHÁLY KIS GYŐZŐ GARAB LÁSZLÓ KOVÁCS 《Plant, cell & environment》2012,35(12):2075-2086
Despite intense research, the mechanism of Cd2+ toxicity on photosynthesis is still elusive because of the multiplicity of the inhibitory effects and different barriers in plants. The quick Cd2+ uptake in Synechocystis PCC 6803 permits the direct interaction of cadmium with the photosynthetic machinery and allows the distinction between primary and secondary effects. We show that the CO2‐dependent electron transport is rapidly inhibited upon exposing the cells to 40 µm Cd2+ (50% inhibition in ~15 min). However, during this time we observe only symptoms of photosystem I acceptor side limitation and a build of an excitation pressure on the reaction centres, as indicated by light‐induced P700 redox transients, O2 polarography and changes in chlorophyll a fluorescence parameters. Inhibitory effects on photosystem II electron transport and the degradation of the reaction centre protein D1 can only be observed after several hours, and only in the light, as revealed by chlorophyll a fluorescence transients, thermoluminescence and immunoblotting. Despite the marked differences in the manifestations of these short‐ and long‐term effects, they exhibit virtually the same Cd2+ concentration dependence. These data strongly suggest a cascade mechanism of the toxic effect, with a primary effect in the dark reactions. 相似文献