Different light spectra from light-emitting diodes (LEDs) trigger species-specific adaptive responses in plants. We exposed Artemisia argyi (A. argyi) to four LED spectra: white (the control group), monochromatic red light (R), monochromatic blue light (B), or a mixture of R and B light of photon flux density ratio is 3 (RB), with equivalent photoperiod (14 h) and light intensity (160 μmol s-1 m-2). R light accelerated photomorphogenesis but decreased biomass, while B light significantly increased leaf area and short-term exposure (7 days) to B light increased total phenols and flavonoids. HPLC identified chlorogenic acid, 3,5-dicaffeoylquinic acid, gallic acid, jaceosidin, eupatilin, and taxol compounds, with RB and R light significantly accumulating chlorogenic acid, 3,5-dicaffeoylquinic acid, and gallic acid, and B light promoting jaceosidin, eupatilin, and taxol. OJIP measurements showed that B light had the least effect on the effective quantum yield ΦPSII, with higher rETR(II), Fv/Fm, qL and PIabs, followed by RB light. R light led to faster photomorphology but lower biomass than RB and B lights and produced the most inadaptability, as shown by reduced ΦPSII and enlarged ΦNPQ and ΦNO. Overall, short-term B light promoted secondary metabolite production while maintaining effective quantum yield and less energy dissipation.
来自发光二极管(LED)的不同光光谱会引发植物物种特有的适应性反应。我们在光周期(14 小时)和光照强度(160 μmol s-1 m-2)相同的情况下,将蒿属植物暴露在四种 LED 光谱下:白光(对照组)、单色红光(R)、单色蓝光(B)或光通量密度比为 3 的 R 光和 B 光混合物(RB)。R 光加速了光形态发生,但降低了生物量,而 B 光则显著增加了叶面积,B 光的短期照射(7 天)增加了总酚和黄酮类化合物。高效液相色谱鉴定出绿原酸、3,5-二咖啡酰奎宁酸、没食子酸、栀子苷、木犀草素和紫杉醇化合物,其中 RB 和 R 光显著增加绿原酸、3,5-二咖啡酰奎宁酸和没食子酸,B 光促进栀子苷、木犀草素和紫杉醇。OJIP 测量显示,B 光对有效量子产率 ΦPSII 的影响最小,rETR(II)、Fv/Fm、qL 和 PIabs 较高,其次是 RB 光。与 RB 光和 B 光相比,R 光的光形态变化较快,但生物量较低,产生的不适应性最强,表现为 ΦPSII 降低,ΦNPQ 和 ΦNO 扩大。总体而言,短期 B 光促进了次生代谢物的产生,同时保持了有效的量子产率和较少的能量耗散。
{"title":"Plant morphology, secondary metabolites and chlorophyll fluorescence of Artemisia argyi under different LED environments.","authors":"Pengfei Su, Shuangshuang Ding, Dacheng Wang, Wenjie Kan, Meng Yuan, Xue Chen, Caiguo Tang, Jinyan Hou, Lifang Wu","doi":"10.1007/s11120-023-01026-w","DOIUrl":"10.1007/s11120-023-01026-w","url":null,"abstract":"<p><p>Different light spectra from light-emitting diodes (LEDs) trigger species-specific adaptive responses in plants. We exposed Artemisia argyi (A. argyi) to four LED spectra: white (the control group), monochromatic red light (R), monochromatic blue light (B), or a mixture of R and B light of photon flux density ratio is 3 (RB), with equivalent photoperiod (14 h) and light intensity (160 μmol s<sup>-1</sup> m<sup>-2</sup>). R light accelerated photomorphogenesis but decreased biomass, while B light significantly increased leaf area and short-term exposure (7 days) to B light increased total phenols and flavonoids. HPLC identified chlorogenic acid, 3,5-dicaffeoylquinic acid, gallic acid, jaceosidin, eupatilin, and taxol compounds, with RB and R light significantly accumulating chlorogenic acid, 3,5-dicaffeoylquinic acid, and gallic acid, and B light promoting jaceosidin, eupatilin, and taxol. OJIP measurements showed that B light had the least effect on the effective quantum yield ΦPSII, with higher rETR(II), Fv/Fm, qL and PIabs, followed by RB light. R light led to faster photomorphology but lower biomass than RB and B lights and produced the most inadaptability, as shown by reduced ΦPSII and enlarged ΦNPQ and ΦNO. Overall, short-term B light promoted secondary metabolite production while maintaining effective quantum yield and less energy dissipation.</p>","PeriodicalId":20130,"journal":{"name":"Photosynthesis Research","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10197053/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9502724","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-03-01Epub Date: 2023-04-24DOI: 10.1007/s11120-023-01019-9
Irina V Elanskaya, Alexander A Bulychev, Evgeny P Lukashev, Elena M Muronets, Eugene G Maksimov
Flavodiiron proteins Flv1/Flv3 accept electrons from photosystem (PS) I. In this work we investigated light adaptation mechanisms of Flv1-deficient mutant of Synechocystis PCC 6803, incapable to form the Flv1/Flv3 heterodimer. First seconds of dark-light transition were studied by parallel measurements of light-induced changes in chlorophyll fluorescence, P700 redox transformations, fluorescence emission at 77 K, and OCP-dependent fluorescence quenching. During the period of Calvin cycle activation upon dark-light transition, the linear electron transport (LET) in wild type is supported by the Flv1/Flv3 heterodimer, whereas in Δflv1 mutant activation of LET upon illumination is preceded by cyclic electron flow that maintains State 2. The State 2-State 1 transition and Orange Carotenoid Protein (OCP)-dependent non-photochemical quenching occur independently of each other, begin in about 10 s after the illumination of the cells and are accompanied by a short-term re-reduction of the PSI reaction center (P700+). ApcD is important for the State 2-State 1 transition in the Δflv1 mutant, but S-M rise in chlorophyll fluorescence was not completely inhibited in Δflv1/ΔapcD mutant. LET in Δflv1 mutant starts earlier than the S-M rise in chlorophyll fluorescence, and the oxidation of plastoquinol (PQH2) pool promotes the activation of PSII, transient re-reduction of P700+ and transition to State 1. An attempt to induce state transition in the wild type under high intensity light using methyl viologen, highly oxidizing P700 and PQH2, was unsuccessful, showing that oxidation of intersystem electron-transport carriers might be insufficient for the induction of State 2-State 1 transition in wild type of Synechocystis under high light.
{"title":"Roles of ApcD and orange carotenoid protein in photoinduction of electron transport upon dark-light transition in the Synechocystis PCC 6803 mutant deficient in flavodiiron protein Flv1.","authors":"Irina V Elanskaya, Alexander A Bulychev, Evgeny P Lukashev, Elena M Muronets, Eugene G Maksimov","doi":"10.1007/s11120-023-01019-9","DOIUrl":"10.1007/s11120-023-01019-9","url":null,"abstract":"<p><p>Flavodiiron proteins Flv1/Flv3 accept electrons from photosystem (PS) I. In this work we investigated light adaptation mechanisms of Flv1-deficient mutant of Synechocystis PCC 6803, incapable to form the Flv1/Flv3 heterodimer. First seconds of dark-light transition were studied by parallel measurements of light-induced changes in chlorophyll fluorescence, P700 redox transformations, fluorescence emission at 77 K, and OCP-dependent fluorescence quenching. During the period of Calvin cycle activation upon dark-light transition, the linear electron transport (LET) in wild type is supported by the Flv1/Flv3 heterodimer, whereas in Δflv1 mutant activation of LET upon illumination is preceded by cyclic electron flow that maintains State 2. The State 2-State 1 transition and Orange Carotenoid Protein (OCP)-dependent non-photochemical quenching occur independently of each other, begin in about 10 s after the illumination of the cells and are accompanied by a short-term re-reduction of the PSI reaction center (P700<sup>+</sup>). ApcD is important for the State 2-State 1 transition in the Δflv1 mutant, but S-M rise in chlorophyll fluorescence was not completely inhibited in Δflv1/ΔapcD mutant. LET in Δflv1 mutant starts earlier than the S-M rise in chlorophyll fluorescence, and the oxidation of plastoquinol (PQH<sub>2</sub>) pool promotes the activation of PSII, transient re-reduction of P700<sup>+</sup> and transition to State 1. An attempt to induce state transition in the wild type under high intensity light using methyl viologen, highly oxidizing P700 and PQH<sub>2</sub>, was unsuccessful, showing that oxidation of intersystem electron-transport carriers might be insufficient for the induction of State 2-State 1 transition in wild type of Synechocystis under high light.</p>","PeriodicalId":20130,"journal":{"name":"Photosynthesis Research","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9394688","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-03-01Epub Date: 2023-11-30DOI: 10.1007/s11120-023-01059-1
M Kis, T Szabó, J Tandori, P Maróti
In photosynthetic bacteria, the absorbed light drives the canonical cyclic electron transfer between the reaction center and the cytochrome bc1 complexes via the pools of mobile electron carriers. If kinetic or structural barriers hinder the participation of the bc1 complex in the cyclic flow of electrons, then the pools of mobile redox agents must supply the electrons for the multiple turnovers of the reaction center. These conditions were achieved by continuous high light excitation of intact cells of bacterial strains Rba. sphaeroides and Rvx. gelatinosus with depleted donor side cytochromes c2 (cycA) and tetraheme cytochrome subunit (pufC), respectively. The gradual oxidation by ferricyanide further reduced the availability of electron donors to pufC. Electron transfer through the reaction center was tracked by absorption change and by induction and relaxation of the fluorescence of the bacteriochlorophyll dimer. The rate constants of the electron transfer (~ 3 × 103 s‒1) from the mobile donors of Rvx. gelatinosus bound either to the RC (pufC) or to the tetraheme subunit (wild type) were similar. The electrons transferred through the reaction center dimer were supplied entirely by the donor pool; their number amounted to about 5 in wild type Rvx. gelatinosus and decreased to 1 in pufC oxidized by ferricyanide. Fluorescence yield was measured as a function of the oxidized fraction of the dimer and its complex shape reveals the contribution of two competing processes: the migration of the excitation energy among the photosynthetic units and the availability of electron donors to the oxidized dimer. The experimental results were simulated and rationalized by a simple kinetic model of the two-electron cycling of the acceptor side combined with aperiodic one-electron redox function of the donor side.
{"title":"Roadmap of electrons from donor side to the reaction center of photosynthetic purple bacteria with mutated cytochromes.","authors":"M Kis, T Szabó, J Tandori, P Maróti","doi":"10.1007/s11120-023-01059-1","DOIUrl":"10.1007/s11120-023-01059-1","url":null,"abstract":"<p><p>In photosynthetic bacteria, the absorbed light drives the canonical cyclic electron transfer between the reaction center and the cytochrome bc<sub>1</sub> complexes via the pools of mobile electron carriers. If kinetic or structural barriers hinder the participation of the bc<sub>1</sub> complex in the cyclic flow of electrons, then the pools of mobile redox agents must supply the electrons for the multiple turnovers of the reaction center. These conditions were achieved by continuous high light excitation of intact cells of bacterial strains Rba. sphaeroides and Rvx. gelatinosus with depleted donor side cytochromes c<sub>2</sub> (cycA) and tetraheme cytochrome subunit (pufC), respectively. The gradual oxidation by ferricyanide further reduced the availability of electron donors to pufC. Electron transfer through the reaction center was tracked by absorption change and by induction and relaxation of the fluorescence of the bacteriochlorophyll dimer. The rate constants of the electron transfer (~ 3 × 10<sup>3</sup> s<sup>‒1</sup>) from the mobile donors of Rvx. gelatinosus bound either to the RC (pufC) or to the tetraheme subunit (wild type) were similar. The electrons transferred through the reaction center dimer were supplied entirely by the donor pool; their number amounted to about 5 in wild type Rvx. gelatinosus and decreased to 1 in pufC oxidized by ferricyanide. Fluorescence yield was measured as a function of the oxidized fraction of the dimer and its complex shape reveals the contribution of two competing processes: the migration of the excitation energy among the photosynthetic units and the availability of electron donors to the oxidized dimer. The experimental results were simulated and rationalized by a simple kinetic model of the two-electron cycling of the acceptor side combined with aperiodic one-electron redox function of the donor side.</p>","PeriodicalId":20130,"journal":{"name":"Photosynthesis Research","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10991045/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138461794","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-03-01Epub Date: 2024-01-10DOI: 10.1007/s11120-023-01064-4
Franz-Josef Schmitt, Anne Hüls, Marcus Moldenhauer, Thomas Friedrich
Halomicronema hongdechloris, the first cyanobacterium reported to produce the red-shifted chlorophyll f (Chl f) upon acclimation to far-red light, demonstrates remarkable adaptability to diverse light conditions. The photosystem II (PS II) of this organism undergoes reversible changes in its Chl f content, ranging from practically zero under white-light culture conditions to a Chl f: Chl a ratio of up to 1:8 when exposed to far-red light (FRL) of 720-730 nm for several days. Our ps time- and wavelength-resolved fluorescence data obtained after excitation of living H. hongdechloris cells indicate that the Soret band of a far-red (FR) chlorophyll involved in charge separation absorbs around 470 nm. At 10 K, the fluorescence decay at 715-720 nm is still fast with a time constant of 165 ps indicating an efficient electron tunneling process. There is efficient excitation energy transfer (EET) from 715-720 nm to 745 nm with the latter resulting from FR Chl f, which mainly functions as light-harvesting pigment upon adaptation to FRL. From there, excitation energy reaches the primary donor in the reaction center of PS II with an energetic uphill EET mechanism inducing charge transfer. The fluorescence data are well explained with a secondary donor PD1 represented by a red-shifted Chl a molecule with characteristic fluorescence around 715 nm and a more red-shifted FR Chl f with fluorescence around 725 nm as primary donor at the ChlD1 or PD2 position.
卤米龙(Halomicronema hongdechloris)是第一个被报道在适应远红光后产生红移叶绿素f(Chl f)的蓝藻,它对不同光照条件的适应能力非常强。这种生物的光系统 II(PS II)的叶绿素 f 含量会发生可逆变化,从白光培养条件下的几乎为零,到暴露在 720-730 纳米的远红光(FRL)下数天后叶绿素 f:叶绿素 a 的比例高达 1:8。我们对活的 H. hongdechloris 细胞进行激发后获得的 ps 时间和波长分辨荧光数据表明,参与电荷分离的远红光(FR)叶绿素的 Soret 波段吸收波长约为 470 nm。在 10 K 时,715-720 nm 处的荧光衰减仍然很快,时间常数为 165 ps,这表明电子隧道过程非常有效。从 715-720 纳米到 745 纳米之间存在高效的激发能量转移(EET),后者来自 FR Chl f,它在适应 FRL 后主要发挥光收集色素的作用。从这里,激发能量到达 PS II 反应中心的初级供体,通过高能上坡 EET 机制诱导电荷转移。在 ChlD1 或 PD2 的位置上,以具有 715 nm 附近特征荧光的红移 Chl a 分子为代表的次级供体 PD1 和具有 725 nm 附近荧光的更红移 FR Chl f 作为初级供体,可以很好地解释荧光数据。
{"title":"How electron tunneling and uphill excitation energy transfer support photochemistry in Halomicronema hongdechloris.","authors":"Franz-Josef Schmitt, Anne Hüls, Marcus Moldenhauer, Thomas Friedrich","doi":"10.1007/s11120-023-01064-4","DOIUrl":"10.1007/s11120-023-01064-4","url":null,"abstract":"<p><p>Halomicronema hongdechloris, the first cyanobacterium reported to produce the red-shifted chlorophyll f (Chl f) upon acclimation to far-red light, demonstrates remarkable adaptability to diverse light conditions. The photosystem II (PS II) of this organism undergoes reversible changes in its Chl f content, ranging from practically zero under white-light culture conditions to a Chl f: Chl a ratio of up to 1:8 when exposed to far-red light (FRL) of 720-730 nm for several days. Our ps time- and wavelength-resolved fluorescence data obtained after excitation of living H. hongdechloris cells indicate that the Soret band of a far-red (FR) chlorophyll involved in charge separation absorbs around 470 nm. At 10 K, the fluorescence decay at 715-720 nm is still fast with a time constant of 165 ps indicating an efficient electron tunneling process. There is efficient excitation energy transfer (EET) from 715-720 nm to 745 nm with the latter resulting from FR Chl f, which mainly functions as light-harvesting pigment upon adaptation to FRL. From there, excitation energy reaches the primary donor in the reaction center of PS II with an energetic uphill EET mechanism inducing charge transfer. The fluorescence data are well explained with a secondary donor P<sub>D1</sub> represented by a red-shifted Chl a molecule with characteristic fluorescence around 715 nm and a more red-shifted FR Chl f with fluorescence around 725 nm as primary donor at the Chl<sub>D1</sub> or P<sub>D2</sub> position.</p>","PeriodicalId":20130,"journal":{"name":"Photosynthesis Research","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139404076","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The objectives of this study were to measure the chlorophyll fluorescence (ChlF) parameters of Barbula indica (Hook.) Spreng and Conocephalum conicum (L.) Dumort subjected to various light intensities (LI) as a reflection of their adaptability to their habitats. The electron transport rate (ETR) of all plants under 500 μmol m-2 s-1 photosynthetic photon flux density (PPFD) was significantly higher than other LI treatments, implying that these plants could be grown under a specific and optimal light intensity adapted to 500 PPFD conditions. As LI increased from 50 to 2,000 PPFD, we observed in all plants increased non-photochemical quenching (NPQ) and photo-inhibitory quenching (qI) and decreased photosystem II efficiency (ΦPSII), potential quantum efficiency of PSII (Fv/Fm), actual PSII efficiency (ΔF/Fm'%), and Fv/Fm%. In addition, energy-dependent quenching (qE), the light protection system (qE + qZ + qT), and qI increased as ΦPSII decreased and photo-inhibition% increased under 1000, 1500, and 2000 PPFD conditions, suggesting that these plants had higher photo-protective ability under high LI treatments to maintain higher photosynthetic system performance. B. indica plants remained photochemically active and maintained higher qE under 300, 500, and 1000 PPFD, whereas C. conicum qZ + qT exhibited higher photo-protection under 500, 1000, and 1500 PPFD conditions. These ChlF indices can be used for predicting photosynthetic responses to light induction in different bryophytes and provide a theoretical basis for ecological monitoring.
{"title":"Photo-protection and photo-inhibition during light induction in Barbula indica and Conocephalum conicum under different light gradients.","authors":"Chung-I Chen, Kuan-Hung Lin, Meng-Yuan Huang, Kuei-Yu Yao, Chau-Ching Huang, Tzu-Chao Lin, En-Liang Chu, Jia-Dong Yang, Ching-Wen Wang","doi":"10.1007/s11120-023-01030-0","DOIUrl":"10.1007/s11120-023-01030-0","url":null,"abstract":"<p><p>The objectives of this study were to measure the chlorophyll fluorescence (ChlF) parameters of Barbula indica (Hook.) Spreng and Conocephalum conicum (L.) Dumort subjected to various light intensities (LI) as a reflection of their adaptability to their habitats. The electron transport rate (ETR) of all plants under 500 μmol m<sup>-2</sup> s<sup>-1</sup> photosynthetic photon flux density (PPFD) was significantly higher than other LI treatments, implying that these plants could be grown under a specific and optimal light intensity adapted to 500 PPFD conditions. As LI increased from 50 to 2,000 PPFD, we observed in all plants increased non-photochemical quenching (NPQ) and photo-inhibitory quenching (q<sub>I</sub>) and decreased photosystem II efficiency (ΦPSII), potential quantum efficiency of PSII (F<sub>v</sub>/F<sub>m</sub>), actual PSII efficiency (ΔF/F<sub>m</sub>'%), and F<sub>v</sub>/F<sub>m</sub>%. In addition, energy-dependent quenching (q<sub>E</sub>), the light protection system (q<sub>E</sub> + q<sub>Z</sub> + q<sub>T</sub>), and q<sub>I</sub> increased as ΦPSII decreased and photo-inhibition% increased under 1000, 1500, and 2000 PPFD conditions, suggesting that these plants had higher photo-protective ability under high LI treatments to maintain higher photosynthetic system performance. B. indica plants remained photochemically active and maintained higher q<sub>E</sub> under 300, 500, and 1000 PPFD, whereas C. conicum q<sub>Z</sub> + q<sub>T</sub> exhibited higher photo-protection under 500, 1000, and 1500 PPFD conditions. These ChlF indices can be used for predicting photosynthetic responses to light induction in different bryophytes and provide a theoretical basis for ecological monitoring.</p>","PeriodicalId":20130,"journal":{"name":"Photosynthesis Research","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10037051","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-03-01Epub Date: 2023-07-07DOI: 10.1007/s11120-023-01035-9
Sarah M Mäusle, Neva Agarwala, Viktor G Eichmann, Holger Dau, Dennis J Nürnberg, Gary Hastings
Microsecond time-resolved step-scan FTIR difference spectroscopy was used to study photosystem I (PSI) from Thermosynechococcus vestitus BP-1 (T. vestitus, formerly known as T. elongatus) at 77 K. In addition, photoaccumulated (P700+-P700) FTIR difference spectra were obtained at both 77 and 293 K. The FTIR difference spectra are presented here for the first time. To extend upon these FTIR studies nanosecond time-resolved infrared difference spectroscopy was also used to study PSI from T. vestitus at 296 K. Nanosecond infrared spectroscopy has never been used to study PSI samples at physiological temperatures, and here it is shown that such an approach has great value as it allows a direct probe of electron transfer down both branches in PSI. In PSI at 296 K, the infrared flash-induced absorption changes indicate electron transfer down the B- and A-branches is characterized by time constants of 33 and 364 ns, respectively, in good agreement with visible spectroscopy studies. These time constants are associated with forward electron transfer from A1- to FX on the B- and A-branches, respectively. At several infrared wavelengths flash-induced absorption changes at 296 K recover in tens to hundreds of milliseconds. The dominant decay phase is characterized by a lifetime of 128 ms. These millisecond changes are assigned to radical pair recombination reactions, with the changes being associated primarily with P700+ rereduction. This conclusion follows from the observation that the millisecond infrared spectrum is very similar to the photoaccumulated (P700+-P700) FTIR difference spectrum.
使用微秒时间分辨阶跃扫描傅立叶变换红外差分光谱法研究了 77 K 时热鞘藻细胞 BP-1(Thermosynechococcus vestitus BP-1,以前称为 T.elongatus)的光系统 I(PSI)。在这些傅立叶变换红外光谱研究的基础上,我们还使用了纳秒时间分辨红外差分光谱来研究开氏 296 度时的 T. vestitus PSI。纳秒红外光谱从未被用于研究生理温度下的 PSI 样品,而这里的研究表明,这种方法具有很大的价值,因为它可以直接探测 PSI 中两个分支的电子传递情况。在 296 K 的 PSI 中,红外闪光灯诱导的吸收变化表明电子沿 B 支和 A 支向下转移的时间常数分别为 33 和 364 ns,这与可见光谱研究结果十分吻合。这些时间常数分别与 B 支链和 A 支链上从 A1 到 FX 的正向电子转移有关。在 296 K 的几个红外波长下,闪烁诱导的吸收变化在几十到几百毫秒内恢复。主要衰减阶段的寿命为 128 毫秒。这些毫秒级的变化归因于自由基对重组反应,主要与 P700+ 的还原有关。这一结论来自于毫秒级红外光谱与光累积(P700+-P700)傅立叶变换红外差分光谱非常相似的观察结果。
{"title":"Nanosecond time-resolved infrared spectroscopy for the study of electron transfer in photosystem I.","authors":"Sarah M Mäusle, Neva Agarwala, Viktor G Eichmann, Holger Dau, Dennis J Nürnberg, Gary Hastings","doi":"10.1007/s11120-023-01035-9","DOIUrl":"10.1007/s11120-023-01035-9","url":null,"abstract":"<p><p>Microsecond time-resolved step-scan FTIR difference spectroscopy was used to study photosystem I (PSI) from Thermosynechococcus vestitus BP-1 (T. vestitus, formerly known as T. elongatus) at 77 K. In addition, photoaccumulated (P700<sup>+</sup>-P700) FTIR difference spectra were obtained at both 77 and 293 K. The FTIR difference spectra are presented here for the first time. To extend upon these FTIR studies nanosecond time-resolved infrared difference spectroscopy was also used to study PSI from T. vestitus at 296 K. Nanosecond infrared spectroscopy has never been used to study PSI samples at physiological temperatures, and here it is shown that such an approach has great value as it allows a direct probe of electron transfer down both branches in PSI. In PSI at 296 K, the infrared flash-induced absorption changes indicate electron transfer down the B- and A-branches is characterized by time constants of 33 and 364 ns, respectively, in good agreement with visible spectroscopy studies. These time constants are associated with forward electron transfer from A<sub>1</sub><sup>-</sup> to F<sub>X</sub> on the B- and A-branches, respectively. At several infrared wavelengths flash-induced absorption changes at 296 K recover in tens to hundreds of milliseconds. The dominant decay phase is characterized by a lifetime of 128 ms. These millisecond changes are assigned to radical pair recombination reactions, with the changes being associated primarily with P700<sup>+</sup> rereduction. This conclusion follows from the observation that the millisecond infrared spectrum is very similar to the photoaccumulated (P700<sup>+</sup>-P700) FTIR difference spectrum.</p>","PeriodicalId":20130,"journal":{"name":"Photosynthesis Research","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10991071/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9763937","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-03-01Epub Date: 2023-06-16DOI: 10.1007/s11120-023-01031-z
Igor N Stadnichuk, Pavel M Krasilnikov
The chromophorylated PBLcm domain of the ApcE linker protein in the cyanobacterial phycobilisome (PBS) serves as a bottleneck for Förster resonance energy transfer (FRET) from the PBS to the antennal chlorophyll of photosystem II (PS II) and as a redirection point for energy distribution to the orange protein ketocarotenoid (OCP), which is excitonically coupled to the PBLcm chromophore in the process of non-photochemical quenching (NPQ) under high light conditions. The involvement of PBLcm in the quenching process was first directly demonstrated by measuring steady-state fluorescence spectra of cyanobacterial cells at different stages of NPQ development. The time required to transfer energy from the PBLcm to the OCP is several times shorter than the time it takes to transfer energy from the PBLcm to the PS II, ensuring quenching efficiency. The data obtained provide an explanation for the different rates of PBS quenching in vivo and in vitro according to the half ratio of OCP/PBS in the cyanobacterial cell, which is tens of times lower than that realized for an effective NPQ process in solution.
{"title":"Relationship between non-photochemical quenching efficiency and the energy transfer rate from phycobilisomes to photosystem II.","authors":"Igor N Stadnichuk, Pavel M Krasilnikov","doi":"10.1007/s11120-023-01031-z","DOIUrl":"10.1007/s11120-023-01031-z","url":null,"abstract":"<p><p>The chromophorylated PBLcm domain of the ApcE linker protein in the cyanobacterial phycobilisome (PBS) serves as a bottleneck for Förster resonance energy transfer (FRET) from the PBS to the antennal chlorophyll of photosystem II (PS II) and as a redirection point for energy distribution to the orange protein ketocarotenoid (OCP), which is excitonically coupled to the PBLcm chromophore in the process of non-photochemical quenching (NPQ) under high light conditions. The involvement of PBLcm in the quenching process was first directly demonstrated by measuring steady-state fluorescence spectra of cyanobacterial cells at different stages of NPQ development. The time required to transfer energy from the PBLcm to the OCP is several times shorter than the time it takes to transfer energy from the PBLcm to the PS II, ensuring quenching efficiency. The data obtained provide an explanation for the different rates of PBS quenching in vivo and in vitro according to the half ratio of OCP/PBS in the cyanobacterial cell, which is tens of times lower than that realized for an effective NPQ process in solution.</p>","PeriodicalId":20130,"journal":{"name":"Photosynthesis Research","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9635001","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-03-01Epub Date: 2024-02-05DOI: 10.1007/s11120-023-01070-6
A S Benditkis, A A Ashikhmin, A A Moskalenko, A A Krasnovsky
Measurement of photosensitized luminescence of singlet oxygen has been applied to studies of singlet oxygen generation and quenching by C40 carotenoids (neurosporene, lycopene, rhodopin, and spirilloxanthin) with long chain of conjugated double bonds (CDB) using hexafluorobenzene as a solvent. It has been found that neurosporene, lycopene, and rhodopin are capable of the low efficient singlet oxygen generation in aerated solutions upon photoexcitation in the spectral region of their main absorption maxima. The quantum yield of this process was estimated to be (1.5-3.0) × 10-2. This value is near the singlet oxygen yields in solutions of ζ-carotene (7 CDB) and phytoene (3 CDB) and many-fold smaller than in solutions of phytofluene (5 CDB) (Ashikhmin et al. Biochemistry (Mosc) 85:773-780, https://doi.org/10.1134/S0006297920070056 , 2020, Biochemistry (Mosc) 87:1169-1178, 2022, https://doi.org/10.1134/S00062979221001082022 ). Photogeneration of singlet oxygen was not observed in spirilloxanthin solutions. A correlation was found between the singlet oxygen yields and the quantum yields and lifetimes of the fluorescence of the carotenoid molecules. All carotenoids were shown to be strong physical quenchers of singlet oxygen. The rate constants of 1O2 quenching by the carotenoids with long chain of CDB (9-13) were close to the rate constant of the diffusion-limited reactions ≈1010 M-1 s-1, being many-fold greater than the rate constants of 1O2 quenching by the carotenoids with the short chain of CDB (3-7) phytoene, phytofluene, and ζ-carotene studied in prior papers of our group (Ashikhmin et al. 2020, 2022). To our knowledge, the quenching rate constants of rhodopin and spirilloxanthin have been obtained in this paper for the first time. The mechanisms of 1O2 photogeneration by carotenoids in solution and in the LH2 complexes of photosynthetic cells, as well as the efficiencies of their protective action are discussed.
{"title":"Photogeneration and quenching of singlet molecular oxygen by bacterial C<sub>40</sub> carotenoids with long chain of conjugated double bonds.","authors":"A S Benditkis, A A Ashikhmin, A A Moskalenko, A A Krasnovsky","doi":"10.1007/s11120-023-01070-6","DOIUrl":"10.1007/s11120-023-01070-6","url":null,"abstract":"<p><p>Measurement of photosensitized luminescence of singlet oxygen has been applied to studies of singlet oxygen generation and quenching by C<sub>40</sub> carotenoids (neurosporene, lycopene, rhodopin, and spirilloxanthin) with long chain of conjugated double bonds (CDB) using hexafluorobenzene as a solvent. It has been found that neurosporene, lycopene, and rhodopin are capable of the low efficient singlet oxygen generation in aerated solutions upon photoexcitation in the spectral region of their main absorption maxima. The quantum yield of this process was estimated to be (1.5-3.0) × 10<sup>-2</sup>. This value is near the singlet oxygen yields in solutions of ζ-carotene (7 CDB) and phytoene (3 CDB) and many-fold smaller than in solutions of phytofluene (5 CDB) (Ashikhmin et al. Biochemistry (Mosc) 85:773-780, https://doi.org/10.1134/S0006297920070056 , 2020, Biochemistry (Mosc) 87:1169-1178, 2022, https://doi.org/10.1134/S00062979221001082022 ). Photogeneration of singlet oxygen was not observed in spirilloxanthin solutions. A correlation was found between the singlet oxygen yields and the quantum yields and lifetimes of the fluorescence of the carotenoid molecules. All carotenoids were shown to be strong physical quenchers of singlet oxygen. The rate constants of <sup>1</sup>O<sub>2</sub> quenching by the carotenoids with long chain of CDB (9-13) were close to the rate constant of the diffusion-limited reactions ≈10<sup>10</sup> M<sup>-1</sup> s<sup>-1</sup>, being many-fold greater than the rate constants of <sup>1</sup>O<sub>2</sub> quenching by the carotenoids with the short chain of CDB (3-7) phytoene, phytofluene, and ζ-carotene studied in prior papers of our group (Ashikhmin et al. 2020, 2022). To our knowledge, the quenching rate constants of rhodopin and spirilloxanthin have been obtained in this paper for the first time. The mechanisms of <sup>1</sup>O<sub>2</sub> photogeneration by carotenoids in solution and in the LH2 complexes of photosynthetic cells, as well as the efficiencies of their protective action are discussed.</p>","PeriodicalId":20130,"journal":{"name":"Photosynthesis Research","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139692678","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-02-08DOI: 10.1007/s11120-024-01075-9
Colin Gates, Gennady Ananyev, Fatima Foflonker, Debashish Bhattacharya, G Charles Dismukes
The green algal genus Picochlorum is of biotechnological interest because of its robust response to multiple environmental stresses. We compared the metabolic performance of P. SE3 and P. oklahomense to diverse microbial phototrophs and observed exceptional performance of photosystem II (PSII) in light energy conversion in both Picochlorum species. The quantum yield (QY) for O2 evolution is the highest of any phototroph yet observed, 32% (20%) by P. SE3 (P. okl) when normalized to total PSII subunit PsbA (D1) protein, and 80% (75%) normalized per active PSII, respectively. Three factors contribute: (1) an efficient water oxidizing complex (WOC) with the fewest photochemical misses of any organism; (2) faster reoxidation of reduced (PQH2)B in P. SE3 than in P. okl. (period-2 Fourier amplitude); and (3) rapid reoxidation of the plastoquinol pool by downstream electron carriers (Cyt b6f/PETC) that regenerates PQ faster in P. SE3. This performance gain is achieved without significant residue changes around the QB site and thus points to a pull mechanism involving faster PQH2 reoxidation by Cyt b6f/PETC that offsets charge recombination. This high flux in P. SE3 may be explained by genomically encoded plastoquinol terminal oxidases 1 and 2, whereas P. oklahomense has neither. Our results suggest two distinct types of PSII centers exist, one specializing in linear electron flow and the other in PSII-cyclic electron flow. Several amino acids within D1 differ from those in the low-light-descended D1 sequences conserved in Viridiplantae, and more closely match those in cyanobacterial high-light D1 isoforms, including changes near tyrosine Yz and a water/proton channel near the WOC. These residue changes may contribute to the exceptional performance of Picochlorum at high-light intensities by increasing the water oxidation efficiency and the electron/proton flux through the PSII acceptors (QAQB).
{"title":"Exceptional Quantum Efficiency Powers Biomass Production in Halotolerant Algae Picochlorum sp.<sup />","authors":"Colin Gates, Gennady Ananyev, Fatima Foflonker, Debashish Bhattacharya, G Charles Dismukes","doi":"10.1007/s11120-024-01075-9","DOIUrl":"https://doi.org/10.1007/s11120-024-01075-9","url":null,"abstract":"<p><p>The green algal genus Picochlorum is of biotechnological interest because of its robust response to multiple environmental stresses. We compared the metabolic performance of P. SE3 and P. oklahomense to diverse microbial phototrophs and observed exceptional performance of photosystem II (PSII) in light energy conversion in both Picochlorum species. The quantum yield (QY) for O<sub>2</sub> evolution is the highest of any phototroph yet observed, 32% (20%) by P. SE3 (P. okl) when normalized to total PSII subunit PsbA (D1) protein, and 80% (75%) normalized per active PSII, respectively. Three factors contribute: (1) an efficient water oxidizing complex (WOC) with the fewest photochemical misses of any organism; (2) faster reoxidation of reduced (PQH<sub>2</sub>)<sub>B</sub> in P. SE3 than in P. okl. (period-2 Fourier amplitude); and (3) rapid reoxidation of the plastoquinol pool by downstream electron carriers (Cyt b<sub>6</sub>f/PETC) that regenerates PQ faster in P. SE3. This performance gain is achieved without significant residue changes around the Q<sub>B</sub> site and thus points to a pull mechanism involving faster PQH<sub>2</sub> reoxidation by Cyt b<sub>6</sub>f/PETC that offsets charge recombination. This high flux in P. SE3 may be explained by genomically encoded plastoquinol terminal oxidases 1 and 2, whereas P. oklahomense has neither. Our results suggest two distinct types of PSII centers exist, one specializing in linear electron flow and the other in PSII-cyclic electron flow. Several amino acids within D1 differ from those in the low-light-descended D1 sequences conserved in Viridiplantae, and more closely match those in cyanobacterial high-light D1 isoforms, including changes near tyrosine Y<sub>z</sub> and a water/proton channel near the WOC. These residue changes may contribute to the exceptional performance of Picochlorum at high-light intensities by increasing the water oxidation efficiency and the electron/proton flux through the PSII acceptors (Q<sub>A</sub>Q<sub>B</sub>).</p>","PeriodicalId":20130,"journal":{"name":"Photosynthesis Research","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-02-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139703166","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-02-02DOI: 10.1007/s11120-023-01069-z
Karim A. Walters, Kevin E. Redding, John H. Golbeck
The homodimeric Type I reaction center (RC) from Heliomicrobium modesticaldum lacks the PsaC subunit found in Photosystem I and instead uses the interpolypeptide [4Fe–4S] cluster FX as the terminal electron acceptor. Our goal was to identify which of the small mobile dicluster ferredoxins encoded by the H. modesticaldum genome are capable of accepting electrons from the heliobacterial RC (HbRC) and pyruvate:ferredoxin oxidoreductase (PFOR), a key metabolic enzyme. Analysis of the genome revealed seven candidates: HM1_1462 (PshB1), HM1_1461 (PshB2), HM1_2505 (Fdx3), HM1_0869 (FdxB), HM1_1043, HM1_0357, and HM1_2767. Heterologous expression in Escherichia coli and studies using time-resolved optical spectroscopy revealed that only PshB1, PshB2, and Fdx3 are capable of accepting electrons from the HbRC and PFOR. Modeling studies using AlphaFold show that only PshB1, PshB2, and Fdx3 should be capable of docking on PFOR at a positively charged patch that overlays a surface-proximal [4Fe–4S] cluster. Proteomic analysis of wild-type and gene deletion strains ΔpshB1, ΔpshB2, ΔpshB1pshB2, and Δfdx3 grown under nitrogen-replete conditions revealed that Fdx3 is undetectable in the wild-type, ΔpshB1, and Δfdx3 strains, but it is present in the ΔpshB2 and ΔpshB1pshB2 strains, implying that Fdx3 may substitute for PshB2. When grown under nitrogen-deplete conditions, Fdx3 is present in the wild-type and all deletion strains except for Δfdx3. None of the knockout strains demonstrated significant impairment during chemotrophic dark growth on pyruvate, photoheterotrophic light growth on pyruvate, or phototrophic growth on acetate+CO2, indicating a high degree of redundancy among these three electron transfer proteins. Loss of both PshB1 and PshB2, but not FdxB, resulted in poor growth under N2-fixing conditions.
H. modesticaldum 的同源二聚体 I 型反应中心(RC)缺乏光系统 I 中的 PsaC 亚基,而是使用多肽间[4Fe-4S]簇 FX 作为终端电子受体。我们的目标是确定 H. modesticaldum 基因组编码的小型移动二簇铁氧还蛋白中哪些能够接受来自日光细菌 RC(HbRC)和丙酮酸:铁氧还蛋白氧化还原酶(PFOR)(一种关键的代谢酶)的电子。对基因组的分析发现了七个候选基因:HM1_1462(PshB1)、HM1_1461(PshB2)、HM1_2505(Fdx3)、HM1_0869(FdxB)、HM1_1043、HM1_0357 和 HM1_2767。在大肠杆菌中的异源表达和使用时间分辨光学光谱的研究表明,只有 PshB1、PshB2 和 Fdx3 能够接受来自 HbRC 和 PFOR 的电子。利用 AlphaFold 进行的建模研究表明,只有 PshB1、PshB2 和 Fdx3 能够与 PFOR 上覆盖表面近端 [4Fe-4S] 簇的带正电补丁对接。对在氮充足条件下生长的野生型和基因缺失株ΔpshB1、ΔpshB2、ΔpshB1pshB2和Δfdx3进行的蛋白质组分析表明,在野生型中检测不到Fdx3、但在ΔpshB2 和 ΔpshB1pshB2 株系中却存在,这意味着 Fdx3 可能替代了 PshB2。在缺氮条件下生长时,野生型和除Δfdx3以外的所有基因缺失菌株中都存在 Fdx3。在丙酮酸的趋化暗生、丙酮酸的光异养光生或乙酸+CO2的光生过程中,没有一个基因敲除菌株表现出明显的生长障碍,这表明这三种电子传递蛋白之间存在高度冗余。缺失 PshB1 和 PshB2,但不缺失 FdxB,会导致固氮条件下的生长不良。
{"title":"Identification and characterization of the low molecular mass ferredoxins involved in central metabolism in Heliomicrobium modesticaldum","authors":"Karim A. Walters, Kevin E. Redding, John H. Golbeck","doi":"10.1007/s11120-023-01069-z","DOIUrl":"https://doi.org/10.1007/s11120-023-01069-z","url":null,"abstract":"<p>The homodimeric Type I reaction center (RC) from <i>Heliomicrobium modesticaldum</i> lacks the PsaC subunit found in Photosystem I and instead uses the interpolypeptide [4Fe–4S] cluster F<sub>X</sub> as the terminal electron acceptor. Our goal was to identify which of the small mobile dicluster ferredoxins encoded by the <i>H. modesticaldum</i> genome are capable of accepting electrons from the heliobacterial RC (HbRC) and pyruvate:ferredoxin oxidoreductase (PFOR), a key metabolic enzyme. Analysis of the genome revealed seven candidates: HM1_1462 (PshB1), HM1_1461 (PshB2), HM1_2505 (Fdx3), HM1_0869 (FdxB), HM1_1043, HM1_0357, and HM1_2767. Heterologous expression in <i>Escherichia coli</i> and studies using time-resolved optical spectroscopy revealed that only PshB1, PshB2, and Fdx3 are capable of accepting electrons from the HbRC and PFOR. Modeling studies using AlphaFold show that only PshB1, PshB2, and Fdx3 should be capable of docking on PFOR at a positively charged patch that overlays a surface-proximal [4Fe–4S] cluster. Proteomic analysis of wild-type and gene deletion strains <i>ΔpshB1</i>, <i>ΔpshB2</i>, <i>ΔpshB1pshB2,</i> and Δ<i>fdx3</i> grown under nitrogen-replete conditions revealed that Fdx3 is undetectable in the wild-type, <i>ΔpshB1, and</i> Δ<i>fdx3</i> strains, but it is present in the <i>ΔpshB2</i> and <i>ΔpshB1pshB2</i> strains, implying that Fdx3 may substitute for PshB2. When grown under nitrogen-deplete conditions, Fdx3 is present in the wild-type and all deletion strains except for Δ<i>fdx3.</i> None of the knockout strains demonstrated significant impairment during chemotrophic dark growth on pyruvate, photoheterotrophic light growth on pyruvate, or phototrophic growth on acetate+CO<sub>2</sub>, indicating a high degree of redundancy among these three electron transfer proteins. Loss of both PshB1 and PshB2, but not FdxB, resulted in poor growth under N<sub>2</sub>-fixing conditions.</p>","PeriodicalId":20130,"journal":{"name":"Photosynthesis Research","volume":null,"pages":null},"PeriodicalIF":3.7,"publicationDate":"2024-02-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139662191","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}