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1.
Excess light triggers protective nonradiative dissipation of excitation energy in photosystem II through the formation of a trans-thylakoid pH gradient that in turn stimulates formation of zeaxanthin and antheraxanthin. These xanthophylls when combined with protonation of antenna pigment-protein complexes may increase nonradiative dissipation and, thus, quench chlorophyll a fluorescence. Here we measured, in parallel, the chlorophyll a fluorescence lifetime and intensity to understand the mechanism of this process. Increasing the xanthophyll concentration in the presence of a pH gradient (quenched conditions) decreases the fractional intensity of a fluorescence lifetime component centered at approximately 2 ns and increases a component at approximately 0.4 ns. Uncoupling the pH gradient (unquenched conditions) eliminates the 0.4-ns component. Changes in the xanthophyll concentration do not significantly affect the fluorescence lifetimes in either the quenched or unquenched sample conditions. However, there are differences in fluorescence lifetimes between the quenched and unquenched states that are due to pH-related, but nonxanthophyll-related, processes. Quenching of the maximal fluorescence intensity correlates with both the xanthophyll concentration and the fractional intensity of the 0.4-ns component. The unchanged fluorescence lifetimes and the proportional quenching of the maximal and dark-level fluorescence intensities indicate that the xanthophylls act on antenna, not reaction center processes. Further, the fluorescence quenching is interpreted as the combined effect of the pH gradient and xanthophyll concentration, resulting in the formation of a quenching complex with a short (approximately 0.4 ns) fluorescence lifetime.  相似文献   

2.
Photosystem (PS) II centers, which split water into oxygen, protons, and electrons during photosynthesis, require light but are paradoxically inactivated by it. Prolonged light exposure concomitantly decreased both the functional fraction of PSII reaction centers and the integral PSII chlorophyll (Chl) a fluorescence lifetime in leaf segments of Capsicum annuum L. Acceleration of photoinactivation of PSII by a pretreatment with the inhibitors/uncoupler lincomycin, DTT, or nigericin further reduced PSII Chl a fluorescence lifetimes. A global analysis of fluorescence lifetime distributions revealed the presence of at least two distinct populations of photoinactivated PSII centers, one at 1.25 ns, and the other at 0.58 ns. Light treatment first increased the 1.25-ns component, a weak quencher, at the expense of a component at 2.22 ns corresponding to functional PSII centers. The 0.58-ns component, a strong quencher, emerged later than the 1.25-ns component. The strongly quenching PSII reaction centers could serve to avoid further damage to themselves and protect their functional neighbors by acting as strong energy sinks.  相似文献   

3.
The role of carotenoids in quenching of chlorophyll fluorescence in the major light-harvesting complex of photosystem II has been studied with a view to understanding the molecular basis of the control of photoprotective nonradiative energy dissipation by the xanthophyll cycle in vivo. The control of chlorophyll fluorescence quenching in the isolated complex has been investigated in terms of the number of the conjugated double bonds for a series of carotenoids ranging from n = 5-19, giving an estimated first excited singlet state energy from 20,700 cm-1 to 10,120 cm-1. At pH 7.8 the addition of exogenous carotenoids with >=10 conjugated double bonds (including zeaxanthin) stimulated fluorescence quenching relative to the control with no added carotenoid, whereas those with n 相似文献   

4.
In a previous study, we measured the redox potential of the primary electron acceptor pheophytin (Phe) a of photosystem (PS) II in the chlorophyll d-dominated cyanobacterium Acaryochloris marina and a chlorophyll a-containing cyanobacterium, Synechocystis. We obtained the midpoint redox potential (E(m)) values of -478 mV for A. marina and -536 mV for Synechocystis. In this study, we measured the redox potentials of the primary electron acceptor quinone molecule (Q(A)), i.e., E(m)(Q(A)/Q(A)(-)), of PS II and the energy difference between [P680·Phe a(-)·Q(A)] and [P680·Phe a·Q(A)(-)], i.e., ΔG(PhQ). The E(m)(Q(A)/Q(A)(-)) of A. marina was determined to be +64 mV without the Mn cluster and was estimated to be -66 to -86 mV with a Mn-depletion shift (130-150 mV), as observed with other organisms. The E(m)(Phe a/Phe a(-)) in Synechocystis was measured to be -525 mV with the Mn cluster, which is consistent with our previous report. The Mn-depleted downshift of the potential was measured to be approximately -77 mV in Synechocystis, and this value was applied to A. marina (-478 mV); the E(m)(Phe a/Phe a(-)) was estimated to be approximately -401 mV. These values gave rise to a ΔG(PhQ) of -325 mV for A. marina and -383 mV for Synechocystis. In the two cyanobacteria, the energetics in PS II were conserved, even though the potentials of Q(A)(-) and Phe a(-) were relatively shifted depending on the special pair, indicating a common strategy for electron transfer in oxygenic photosynthetic organisms.  相似文献   

5.
Antibody to the Euglena light-harvesting chlorophyll a/b binding protein of photosystem II (LHCPII) immunoprecipitated 207-, 161-, 122-, and 110-kDa proteins from total Euglena proteins pulse-labeled for 10 min with [35S]sulfate. The 25.6- and 27.2-kDa LHCPII were barely detectable in the immunoprecipitate. During a 40-min chase with unlabeled sulfate, the amount of radioactivity in the high molecular mass proteins decreased, and the amount of radioactivity in the 25.6- and 27.2-kDa LHCPII increased with kinetics consistent with a precursor-product relationship. The half-life of the high molecular mass proteins was ≈20 min. The major proteins immunoprecipitated from a nuclease-treated rabbit reticulocyte cell-free translation system programmed with Euglena whole cell or poly(A)+ RNA had molecular masses corresponding to the molecular masses of the proteins immunoprecipitated from the pulse-labeled in vivo translation products. RNAs of 6.6 and 8.3 kilobases were the only Euglena whole cell and poly(A)+ RNAs that hybridized to a 0.7-kilobase EcoRI-BamHI fragment of plasmid pAB165, which contains a portion of the coding sequence for Arabidopsis LHCPII. RNAs of this size are more than sufficient to code for proteins of 207 kDa. Taken together, these findings demonstrate that the LHCPIIs of Euglena are initially synthesized as slowly processed precursors with molecular masses of 207, 161, 122, and 110 kDa.  相似文献   

6.
Coral bleaching has been defined as a general phenomenon, whereby reef corals turn visibly pale because of the loss of their symbiotic dinoflagellates and/or algal pigments during periods of exposure to elevated seawater temperatures. During the summer of 1997, seawater temperatures in the Florida Keys remained at or above 30 degrees C for more than 6 weeks, and extensive coral bleaching was observed. Bleached colonies of the dominant Caribbean reef-building species, Montastrea faveolata and Montastrea franksi, were sampled over a depth gradient from 1 to 17 m during this period of elevated temperature and contained lower densities of symbiotic dinoflagellates in deeper corals than seen in previous "nonbleaching" years. Fluorescence analysis by pulse-amplitude modulation fluorometry revealed severe damage to photosystem II (PSII) in remaining symbionts within the corals, with greater damage indicated at deeper depths. Dinoflagellates with the greatest loss in PSII activity also showed a significant decline in the D1 reaction center protein of PSII, as measured by immunoblot analysis. Laboratory experiments on the temperature-sensitive species Montastrea annularis, as well as temperature-sensitive and temperature-tolerant cultured symbiotic dinoflagellates, confirmed the temperature-dependent loss of PSII activity and concomitant decrease in D1 reaction center protein seen in symbionts collected from corals naturally bleached on the reef. In addition, variation in PSII repair was detected, indicating that perturbation of PSII protein turnover rates during photoinhibition at elevated temperatures underlies the physiological collapse of symbionts in corals susceptible to heat-induced bleaching.  相似文献   

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10.
The photosystem II core complex (PSII-CC) is the smallest subunit of the oxygenic photosynthetic apparatus that contains core antennas and a reaction center, which together allow for rapid energy transfer and charge separation, ultimately leading to efficient solar energy conversion. However, there is a lack of consensus on the interplay between the energy transfer and charge separation dynamics of the core complex. Here, we report the application of two-dimensional electronic-vibrational (2DEV) spectroscopy to the spinach PSII-CC at 77 K. The simultaneous temporal and spectral resolution afforded by 2DEV spectroscopy facilitates the separation and direct assignment of coexisting dynamical processes. Our results show that the dominant dynamics of the PSII-CC are distinct in different excitation energy regions. By separating the excitation regions, we are able to distinguish the intraprotein dynamics and interprotein energy transfer. Additionally, with the improved resolution, we are able to identify the key pigments involved in the pathways, allowing for a direct connection between dynamical and structural information. Specifically, we show that C505 in CP43 and the peripheral chlorophyll ChlzD1 in the reaction center are most likely responsible for energy transfer from CP43 to the reaction center.

Photosynthesis is the process through which solar energy is converted into chemical energy (13). Photosystem II (PSII), a pigment–protein complex found in cyanobacteria, algae, and land plants, is the site of water splitting and is therefore crucial for photosynthetic function (46). It is connected with a large light-harvesting antenna system that collects solar energy and transfers the energy to the reaction center (RC), where charge separation (CS) occurs. Unlike the antenna system of purple bacteria that has a clear energy funnel, the PSII antenna system has a more complicated composition and a very complex energy landscape (47). These features allow for regulation that responds to rapid environmental fluctuations and protect the organisms in, for example, excess light, while maintaining highly efficient electronic energy transfer (EET) under optimal conditions (8). To understand the intricate interactions between the subunits that allow for the robustness of this photosynthetic system, the first step is to understand how the antenna system is connected to the RC. The PSII core complex (PSII-CC) is the smallest unit in which the RC is connected to the antenna proteins. It is a dimeric pigment–protein complex in which each monomer contains an RC and two core antenna proteins, namely, CP43 and CP47 (1, 7). These core antennas not only harvest solar energy but also act as the crucial bridge between the peripheral light-harvesting antenna system and the RC. Fig. 1A shows the pigment arrangement of the PSII-CC. The RC, consisting of the D1 and D2 branches, binds the following pigments: 1) two special pair chlorophyll a (PD1 and PD2), 2) two accessory chlorophyll a (ChlD1 and ChlD2), 3) two pheophytin a (PheoD1 and PheoD2), and 4) two peripheral chlorophyll a (ChlzD1 and ChlzD2) (9, 10). Despite the similarity between the D1 and D2 branches, CS occurs only along the D1 branch (11, 12). CP43, one of the two core antenna proteins, contains 13 chlorophyll a (Chls) and is located closer to the D1 active branch. CP47 contains 16 Chls and is located closer to the D2 branch (10). Together, these proteins provide highly effective EET and CS, which are key to the high quantum yield of CS in the RC.Open in a separate windowFig. 1.(A) Pigment arrangement of monomeric PSII-CC (whereas it is typically found as a dimer) depicted based on the cryoelectron microscopy structure (3JCU) reported by Wei et al. (10). The pigments of CP43, RC, and CP47 are shown in green, blue, and red, respectively. (B) Corresponding excitonic energy levels of monomeric PSII-CC color coded to match pigments in A (5557). The gray shaded regions in the background represent the three groups based on similar characteristic dynamics. Note that the boundaries between the groups provide only a rough separation region as the dynamical behaviors change gradually along ωexc. The asterisk (for the RC state) indicates an optically dark state.Despite the importance of the PSII-CC, its early time dynamics is not fully understood—specifically the competition between EET and CS (5, 7). This is largely due to the highly congested excitonic manifold (Fig. 1B) and ultrafast EET timescales, which challenge ultrafast spectroscopic techniques. Two distinct models have been put forth to try to describe the function of the PSII-CC. These two models are the “exciton/radical pair equilibrium” (ERPE) model (1317) and the “transfer-to-trap limited” (TTTL) model (1822). An early fluorescence decay experiment (13, 14) suggested that rapid EET allows the excitonic states to reach an equilibrium between the core antennas and the RC before CS occurs (kEET ≫ kCS), which is the basis for the ERPE model. This model was later supported by improved time-resolved fluorescence (15) and transient absorption experiments (16). However, a major discrepancy in this model arose with the measurement of the X-ray crystal structure of the PSII-CC (18). It was suggested that the large distances (center-to-center distance, >20 Å) between antenna and RC pigments resolved in the crystal structure would mean that ultrafast EET between the antenna proteins and the RC is unlikely. A model was then put forth that instead suggested that the EET from the core antenna to the RC is slow compared to CS (kEET ≪ kCS), and therefore, the EET to the trap becomes a kinetic bottleneck (18). This TTTL model was later supported by transient infrared (IR) (19) and time-resolved fluorescence experiments (20, 21) as well as structure-based simulations (22). Additionally, Kaucikas et al. (23) performed a polarized transient IR experiment on an oriented single PSII-CC crystal. The decay of the polarization-dependent signature (50–100 ps) observed in their experiment suggests that equilibration between different subunits is slow, consistent with the TTTL model. However, it has been pointed out that satisfactory fitting of the spectral evolution to this model does not necessarily imply that it is correct (24, 25), especially as others have shown that the EET dynamics cannot be adequately described by a single hopping scheme (26, 27). A recent two-dimensional electronic spectroscopy (2DES) experiment (28) with improved time resolution has also revealed the existence of ultrafast EET (<100 fs) that was not predicted by theoretical calculations. In their work, Pan et al. (28) attributed the origin of this unexpectedly fast EET pathway to polaron formation. Vibronic effects on the ultrafast EET and CS dynamics of other photosynthetic proteins have also been discussed (2938).The lack of detailed understanding of the PSII-CC early time dynamics, in particular the EET between the core antennas and the RC, highlights the need for further experimental input with the ability to make specific assignments of the dynamical pathways. This, however, requires simultaneous high temporal and spectral resolution, which remains a challenge for ultrafast spectroscopic techniques. Here, we describe the application of two-dimensional electronic-vibrational (2DEV) spectroscopy (3941) to the PSII-CC. The combination of both spectral dimensions provides an improved resolution that allows us to obtain much more detailed dynamical information in complex systems. The excitonic energy landscapes generated by electronic coupling in photosynthetic complexes, combined with site-dependent and charge state–dependent vibrational spectra, allow the resolution along both axes of 2DEV spectra to provide a direct connection between energetic space (via visible excitation) and physical space (via IR detection). This advantage has proven to be useful for the studies of dynamics in photosynthetic pigment–protein complexes (33, 4045). Specifically, the resolution along the electronic excitation axis allows for the separation of the contributions from different pathways, while the resolution along the vibrational detection axis provides a way to identify the protein subunits or even specific states involved in the dynamics. As we will show, this unique feature of 2DEV spectroscopy provides insight into the complex dynamics of the PSII-CC.In the following text, we will show that the sub-100-ps dynamics of the PSII-CC extracted from spinach are highly dependent on the excitation frequency range. The resolution along the detection axis allows different dominant dynamics to be identified. In addition, we will demonstrate how 2DEV spectroscopy allows us to connect the observed dynamics to specific excitonic states. This connection allows us to obtain a more specific pigment assignment for the EET pathways and therefore provides a more detailed understanding of the finely tuned interactions between the RC and the core antennas (specifically CP43, which is closer to the active D1 branch). We will conclude with a comparison between our results and the existing models in order to provide a path forward in the understanding of this critical photosynthetic component.  相似文献   

11.
X-ray absorption spectroscopy was used to measure the damage caused by exposure to x-rays to the Mn(4)Ca active site in single crystals of photosystem II as a function of dose and energy of x-rays, temperature, and time. These studies reveal that the conditions used for structure determination by x-ray crystallography cause serious damage specifically to the metal-site structure. The x-ray absorption spectra show that the structure changes from one that is characteristic of a high-valent Mn(4)(III(2),IV(2)) oxo-bridged Mn(4)Ca cluster to that of Mn(II) in aqueous solution. This damage to the metal site occurs at a dose that is more than one order of magnitude lower than the dose that results in loss of diffractivity and is commonly considered safe for protein crystallography. These results establish quantitative x-ray dose parameters that are applicable to redox-active metalloproteins. This case study shows that a careful evaluation of the structural intactness of the active site(s) by spectroscopic techniques can validate structures derived from crystallography and that it can be a valuable complementary method before structure-function correlations of metalloproteins can be made on the basis of high-resolution x-ray crystal structures.  相似文献   

12.
Background and aimsIncreased oxidative stress is associated with coronary heart disease (CHD). The mitochondrial uncoupling protein-2 (UCP2) negatively regulates reactive oxygen species generation. We have observed that a common variant (−866G>A) in the promoter region of UCP2 is associated with increased CHD risk in healthy men and increased oxidative stress in diabetic men with CHD. The aim of the current study was to test the hypothesis that this variant might interact with smoking (an environmental stress) to influence plasma markers of oxidative stress.Methods and resultsAmongst 453 Caucasian diabetic men there was a significant interaction (p = 0.001) between genotype and smoking in determining plasma Total AntiOxidant Status (TAOS). Current smokers with the −866AA genotype had the lowest TAOS (indicating higher oxidative stress) of all subjects (AA vs. GG: 32.00 ± 17.4% vs. 45.8 ± 12.6%, p = 0.04). In a sub-sample of 20 subjects (10 GG, 10 AA) matched for baseline characteristics, plasma markers of oxidative stress in current smokers were significantly higher in AA compared to GG subjects (TAOS 36.8 ± 9.5% vs. 51.4 ± 9.5%, p = 0.04; F2-isoprostanes 1133.6 ± 701.2 pg ml−1vs. 500.8 ± 64.7 pg ml−1, p = 0.04).ConclusionsThis study demonstrates an interaction between the UCP2 −866G>A variant and smoking to increase oxidative stress in vivo.  相似文献   

13.
The EDITION trials in type 2 diabetes demonstrated comparable glycaemic control with less nocturnal and anytime (24-hour) hypoglycaemia for insulin glargine 300 U/mL (Gla-300) versus glargine 100 U/mL (Gla-100). However, the predefined nocturnal window (0:00–5:59 AM ) may not be the most relevant for clinical practice. This post-hoc analysis compared expansions of the predefined nocturnal interval during basal insulin treatment without prandial insulin. Patient-level, 6-month data, pooled from the EDITION 2 and 3 trials and the EDITION JP 2 trial (N = 1922, basal insulin only) were analysed. Accompanying hypoglycaemia during treatment with Gla-300 was compared to that during treatment with Gla-100, using predefined (0:00–5:59 AM ) and expanded (10:00 PM –5:59 AM , 0:00–7:59 AM , 10:00 PM to pre-breakfast SMPG) windows. Confirmed (≤3.9 mmol/L [≤70 mg/dL]) or severe hypoglycaemic events were reported most frequently between 6:00 AM and 8:00 AM . Windows expanded beyond 6:00 AM included more events than other windows. The percentage of participants with at least one event was lower with Gla-300 than Gla-100 in all windows examined. Expanding the nocturnal interval allows better assessment of the risk of hypoglycaemia associated with basal insulin. The risk of nocturnal hypoglycaemia was consistently lower with Gla-300 versus Gla-100 using all four windows.  相似文献   

14.
15.
B Vavrusa  H Cleve 《Vox sanguinis》1974,26(2):157-162
Abstract. A family from Opava, Czechoslovakia, with a Gc variant is described. The variant has an electrophoretic mobility between Gc 1–1 and Gc 2–2. The notation Gc Opava (Gc Op) is proposed. The variant appears to be determined by an allele at the Gc locus, GcOp. Three family members had the phenotype Gc Op-1; three family members had the phenotype Gc Op-2.  相似文献   

16.
Background and objective: Dendritic cell‐specific intracellular adhesion molecule‐3 grabbing nonintegrin (DC‐SIGN), encoded by the CD209 gene, is a major Mycobacterium tuberculosis receptor on human dendritic cells. The potentially functional ?336A/G polymorphism in the CD209 promoter region has been associated with susceptibility to tuberculosis (TB), but the results have been inconclusive. We performed a meta‐analysis to clarify the relationship between the CD209?336A/G variant and the risk of TB. Methods: Ten studies involving a total of 2598 TB patients and 2614 control subjects were systematically reviewed, and the data were quantitatively synthesized by meta‐analysis. The Q‐test was applied to assess the heterogeneity of associations among the studies, and Egger's regression test was used to assess potential publication bias. Results: No significant association was identified between the CD209?336A/G polymorphism and risk of TB (G allele vs A allele: odds ratio (OR) 1.02, 95% confidence interval (CI) 0.90–1.15). Moreover, no significant association was observed in populations of African ethnicity (OR 1.01, 95% CI 0.87–1.17) or among individuals who were negative for the human immunodeficiency virus (OR 0.98, 95% CI 0.84–1.15). Conclusions: This meta‐analysis has indicated that the CD209?336A/G polymorphism may not contribute to susceptibility to TB.  相似文献   

17.
Conjugation of ubiquitin to proteins is activated during spermatogenesis. Ubiquitination is mediated by ubiquitin-activating enzyme (E1), ubiquitin-conjugating enzymes (UBCs or E2s), and ubiquitin protein ligases (E3s). Since we previously showed that the activated ubiquitination is UBC4 dependent, we characterized Rat100, a UBC4-dependent E3 expressed in the testis. Analysis of expressed sequence tag sequences and immunoblotting showed that Rat100 is actually a 300-kDa protein expressed mainly in the brain and testis and is similar to the human E3 identified by differential display (EDD) protein and the Drosophila hyperplastic discs gene, mutants of which cause a defect in spermatogenesis. Rat100 is induced during postnatal development of the rat testis, peaking at d 25. It is localized only in germ cells and is highly expressed in spermatocytes, moderately in round and slightly in elongating spermatids. In contrast to UBC4 whose removal from a testis extract abrogates much of the conjugation activity, immmunodepletion of Rat100 from the extracts had little effect. Rat100 therefore has a limited subset of substrates, some of which appear associated with the E3 as the immunoprecipitate containing Rat100 supported incorporation of (125)I-ubiquitin into high molecular weight proteins. Thus, Rat100 is the homolog of human EDD and likely of Drosophila hyperplastic discs. This homology, together with our results, suggests that induction of this E3 results in ubiquitination of specific substrates, some of which are important in male germ cell development.  相似文献   

18.
Outward Na+ - K+ cotransport in erythrocytes from essential hypertensive Caucasian subjects was found to be excessively low (Co -) compared to normotensives (Co +) carefully selected for their negative family history of hypertension. Since the frequency of essential hypertension varies widely among different populations and is particularly high in certain coloured peoples, we compared erythrocyte Na+ - K+ cotransport in normotensive and hypertensive subjects in Paris (France) and in Abidjan (Ivory Coast) to seen whether defective cotransport was related to high blood pressure in the African group as well. Of the 66 French unselected normotensives investigated, 26 (39%) were Co - whereas 14 of the 18 Ivory Coast unselected normotensives (79%) were Co -. 64 (80%) of the 80 essential hypertensives examined in France were Co -, but the proportion of Co - subjects among the Ivory Coast hypertensives was even higher. In addition, both hypertensives and normotensives in the African groups often had undetectable outward Na+ effluxes, a rare finding in the French subjects. We suggest that the high incidence of abnormal Na+ - K+ cotransport in the Ivory Coast series may reflect a genetic propensity to hypertension in this population, and that consequently, Na+ - K+ erythrocyte cotransport measurements might prove useful in defining geographic variations in hypertension.  相似文献   

19.
A 83-year-old woman was admitted to our hospital because of intermittent abdominal colicky pain and vomiting for 26 h.The pain localized over the periumbilical area with radiation along the medial side of the thigh.Computed tomography scan with three-dimensional reconstruction revealed a loop of small bowel protruding into the left obturator canal. Incarcerated obturator hernia was diagnosed and emergency laparotomy was arranged immediately. Unfortunately,her family refused surgery because of her worsening ...  相似文献   

20.
A 83-year-old woman was admitted to our hospital because of intermittent abdominal colicky pain and vomiting for 26 h.The pain localized over the periumbilical area with radiation along the medial side of the thigh.Computed tomography scan with three-dimensional reconstruction revealed a loop of small bowel protruding into the left obturator canal. Incarcerated obturator hernia was diagnosed and emergency laparotomy was arranged immediately. Unfortunately,her family refused surgery because of her worsening ...  相似文献   

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