低叶绿素b水稻突变体类囊体膜的比较蛋白质组学

采用蓝绿温和胶凝胶电泳(blue-nativepolyacrylamidegel-electrophoresis,BN-PAGE),以及改进的第二向SDS-PAGE分离了水稻低叶绿素b突变体ZH249-Y和野生型ZH249-W类囊体膜蛋白复合物,系统比较了突变体和野生型各复合物亚基的表达差异.结果显示,第一向BN-PAGE分离了PSⅠ-LHCⅠ、LHCⅠ缺失的PSⅠ、ATP合成酶、细胞色素b6f、CP43缺失的PSⅡ及LHCⅡ六种复合物.上述各复合物经第二相SDS-Urea-PAGE分离后,利用胶内酶解,高效液相层析分离肽段,电喷雾串联质谱鉴定了复合物的亚基.结合免疫印迹研究,证明和野生型相比,突变体光系统Ⅱ捕光天线复合体的表达量适度下降,但光系统Ⅰ捕光天线破坏严重,同时光系统Ⅱ核心蛋白和ATP合成酶的表达量上升.研究结果对揭示低叶绿素b水稻突变体较高光化学效率和光稳定性的分子基础提供了线索,同时也表明,改进的BN/SDS-PAGE双向电泳不仅可以有效地分离膜蛋白复合物及亚基,也可以进行不同生理条件下,或野生型和突变体之间膜蛋白质组的比较研究.     

龙须菜叶绿素-蛋白复合物的分离及鉴定

以海洋经济海藻--龙须菜(Gracilarialemaneiformis)为材料,机械破碎与超声波相结合破碎这种含胶量非常多的细胞,蔗糖密度梯度超速离心纯化其类囊体膜,去垢剂Triton X-100增溶纯化的类囊体膜,再用蔗糖密度梯度超速离心方法分离其叶绿素-蛋白质复合物.P700差示光谱鉴定分离的光系统Ⅰ(PSⅠ)颗粒,并且检测到了具有DCIP光还原活性的光系统Ⅱ(PSⅡ)颗粒.结果表明,尽管海藻类囊体膜增溶困难,但只要条件合适,可以得到具有活性的光系统颗粒.     

溶液离子强度对光系统Ⅰ和光系统Ⅱ结构性质及分离的影响（英文）

本文运用现代分析手段系统考察了溶液离子强度对菠菜来源光系统Ⅰ(PSⅠ)和光系统(PSⅡ)结构性质的影响,研究的结构性质包括:低温荧光光谱、放(耗)氧活性、聚集尺寸、聚集形貌、Zeta电位和热稳定性等.结果表明,溶液离子强度对PSⅠ和PSⅡ的放(耗)氧活性、聚集尺寸和热稳定性具有显著影响.此外,根据测试结果的分析得知,"筛分效应"在光系统Ⅰ的超滤分离过程中起决定性作用.     

NaCl胁迫加重强光胁迫下超大甜椒叶片的光系统Ⅱ和光系统Ⅰ的光抑制

快速叶绿素荧光动力学可以在无损情况下探知叶片光合机构的损伤程度,快速叶绿素荧光测定和分析技术(JIP-test)将测量值转化为多种具有生物学意义的参数,因而被广泛应用于植物光合机构对环境的响应机制研究.该文研究了超大甜椒(Capsicum annuum)幼苗在强光及不同NaCl浓度胁迫下的荧光响应情况.与单纯强光胁迫相比,NaCl胁迫引起了叶绿素荧光诱导曲线的明显改变,光系统Ⅱ(PSⅡ)光抑制加重,同时PSⅡ反应中心和受体侧受到明显影响,而且高NaCl浓度胁迫下PSⅡ供体侧受伤害明显,同时PSⅠ反应中心活性(P700+)在盐胁迫下明显降低.这些结果表明,NaCl胁迫会增强强光对超大甜椒光系统的光抑制,并且浓度越高抑制越明显,但对PSⅠ的抑制作用低于PSⅡ.高NaCl浓度胁迫易对PSⅡ供体侧造成破坏,且PSⅠ光抑制严重.     

低温光抑制恢复过程中黄瓜叶片PSⅡ活性及其电子传递对PSⅠ的影响

以“津春4号”黄瓜为试材,通过测定黄瓜叶片叶绿素荧光快速诱导动力学曲线和对820 nm光的吸收曲线,结合叶绿素荧光淬灭分析,研究低温光胁迫(4℃,200 μmol·m-2·s-1)6 h后,黄瓜叶片在常温(25℃)不同光强(0、15、200μmol·m-2·s-1)下PS Ⅰ和PS Ⅱ活性的恢复,以及恢复过程中PS Ⅰ与PS Ⅱ的相互作用.结果表明:低温光胁迫6h后,PS Ⅰ和PS Ⅱ发生不同程度的光抑制.在常温恢复阶段,PS Ⅱ活性快速恢复且对光强不敏感;PS Ⅰ活性在弱光下(15 μmol·m-2·s-1)快速恢复,在较强光(200 μmol·m-2·s-1)下恢复较慢.在低温光抑制恢复过程中,常温下PS Ⅱ活性恢复较快可能导致PS Ⅱ向PS Ⅰ的线性电子传递过快,进而抑制PS Ⅰ的活性恢复.因此,在进行黄瓜抗冷性育种时,不应该仅追求较高的PS Ⅱ抗性和较快的PS Ⅱ恢复速度,还应该注意两个光系统活性的协调.在生产中,应当在低温逆境发生及其之后较长一段时间内采取措施降低叶表面光照强度,以利于对植株光合机构的保护和光合活性的恢复.     

低温弱光胁迫对日光温室栽培杏树光系统功能的影响

以温室栽培的金太阳杏为材料,测定了金太阳杏叶片光合速率(P_n)、光系统Ⅱ(PSⅡ)光下实际光化学效率(Φ_PSⅡ)、光化学猝灭系数(q_P)和开放的PSⅡ反应中心的激发能捕获效率(F_v^′/F_m^′), 探讨了低温弱光(7 ℃、200 μmol·m^-2·s^-1 PFD)对叶片光系统Ⅰ(PSⅠ)和PSⅡ的抑制作用.结果表明：温室栽培的金太阳杏叶光合作用的最适温度在25 ℃左右.光下7 ℃的低温可使叶片净光合速率(P_n)大幅下降,造成激发压(1-q_P)增大,进而引起光抑制.低温弱光条件使PSⅠ和PSⅡ功能受到破坏，与单纯低温胁迫(7 ℃,黑暗)处理相比,经低温、弱光(7 ℃, 200 μmol·m^-2·s^-1PFD)胁迫2 h后,PSⅠ活性下降了28.26%,而PSⅡ最大光化学效率（F_v/F_m）没有发生显著变化,表明低温弱光条件下PSⅠ比PSⅡ 更易发生光抑制.     

溶液离子强度对光系统Ⅰ和光系统Ⅱ结构性质及分离的影响

本文运用现代分析手段系统考察了溶液离子强度对菠菜来源光系统Ⅰ(PSⅠ)和光系统(PSⅡ)结构性质的影响,研究的结构性质包括：低温荧光光谱、放(耗)氧活性、聚集尺寸、聚集形貌、Zeta电位和热稳定性等．结果表明,溶液离子强度对PSⅠ和PSⅡ的放(耗)氧活性、聚集尺寸和热稳定性具有显著影响．此外,根据测试结果的分析得知,“筛分效应”在光系统Ⅰ的超滤分离过程中起决定性作用．     

光合作用中光收缩现象的电化学本质

光合水裂解反应的电化学模型要求:PSⅡ的放氧反应和PSⅠ的NADPH还原反应必须同时发生.因此静态时相互分开的PSⅡ、Cytb6 f和PSⅠ,在光合作用中必然会相互紧密结合在一起,形成超结构的光合PSⅡ Cytb6 f PSⅠ聚集体,从而导致类囊体结构的变形.     

两种常绿阔叶植物越冬光系统功能转变的特异性

通过叶绿素荧光和P700氧化还原动力学同步测定,研究大叶黄杨(Euonymus japonicus)和锦熟黄杨(Buxus spervirens L.)的阳生叶和阴生叶在北京地区越冬进程,光系统Ⅱ(PSⅡ)和光系统Ⅰ(PSⅠ)功能转变机制的特异性.结果表明,入冬前0℃以上低温条件下,各叶片PSⅡ有效光量子效率Y(Ⅱ)(Effective quantum yield of PSⅡ)处于同一水平,但阳生叶Y(Ⅰ)(Effective quantum yield of PSⅠ)均高于阴生叶,同时各叶片Fo(Minimal fluorescence)和OJIP水平不完全相同:大叶黄杨两种叶片差异不显著,锦熟黄杨阳生叶显著低于其阴生叶;同步测定P700氧化还原变化表明,两种阴生叶在0-20 ms的P700氧化过程分两个阶段,尤其是锦熟黄杨2 ms后的氧化速率显著降低,而两种阳生叶0-20 ms基本保持同一氧化速率;两种阴生叶的两个光系统量子效率比Y(Ⅰ)/Y(Ⅱ)＜1,两种阳生叶Y(Ⅰ)/Y(Ⅱ)=1.入冬后,各类叶片PSⅡ受到不同程度光抑制,而PS Ⅰ光量子效率Y(Ⅰ)均先增加后减小,Y(Ⅰ)/Y(Ⅱ)发生不同程度增加,Y(Ⅱ)和两个光系统的平衡能力均依次为锦熟黄杨阴生叶＞锦熟黄杨阳生叶＞大叶黄杨阴生叶＞大叶黄杨阳生叶;冬季大叶黄杨阴生叶J相的相对强度高于锦熟黄杨阴生叶,而两种阳生叶OJIP动力学变化几乎消失;同步测定P700氧化还原变化表明,锦熟黄杨阴生叶2 ms即达到Pm(Maximal P700 change),其他叶片0-20 ms保持同一氧化速率,阳生叶Fo、P700氧化速率和Pm均低于阴生叶.返青后,各叶片两个光系统功能逐渐恢复.可见,冬季低温或低温强光逆境会导致阳生叶和阴生叶的两个光系统功能和互动机制发生不同转变.     

铀在小麦幼苗中的积累分布及其对叶片光系统活性的影响

分别采用不同浓度的铀[0、5、20、50、100mg.L-1 UO2(NO3)2.6H2O]对五叶期的‘西科麦3号’小麦幼苗于水培条件下处理7d,分析小麦对铀的吸收积累情况,并通过快速叶绿素荧光诱导动力学OJIP曲线及820nm光吸收曲线,分析铀对叶片光系统Ⅱ(PSⅡ)和光系统Ⅰ(PSⅠ)活性的影响。结果表明:(1)小麦对铀的富集系数和转移系数较小,吸收的铀主要集中在根部。(2)铀胁迫显著降低了小麦叶片捕光色素叶绿素b的含量,并显著影响小麦叶片两个光系统的活性;铀显著抑制PSⅡ反应中心的活性,但是对PSⅡ的电子供体侧和受体侧电子传递活性及PSⅠ的活性则表现为促进作用。(3)低浓度的铀处理会影响小麦叶片中两个光合系统之间的平衡,对PSⅠ性能的促进作用显著大于PSⅡ。     

Demonstration of Two Sites of Inhibition of Electron Transport by Protein Phosphorylation in Wheat Thylakoids

The effect of protein phosphorylation on electron transportactivities of thylakoids isolated from wheat leaves was investigated.Protein phosphorylation resulted in a reduction in the apparentquantum yield of whole chain and photosystem II (PSII) electrontransport but had no effect on photosystem I (PSI) activity.The affinity of the D1 reaction centre polypeptide of PSII tobind atrazine was diminished upon phosphorylation, however,this did not reduce the light-saturated rate of PSII electrontransport. Phosphorylation also produced an inhibition of thelight-saturated rate of electron transport from water or durohydroquinoneto methyl viologen with no similar effect being observed onthe light-saturated rate of either PSII or PSI alone. This suggeststhat phosphorylation produces an inhibition of electron transportat a site, possibly the cytochrome b₆/f complex, between PSIIand PSI. This inhibition of whole-chain electron transport wasalso observed for thylakoids isolated from leaves grown underintermittent light which were deficient in polypeptides belongingto the light-harvesting chlorophyll-protein complex associatedwith photosystem II (LHCII). Consequently, this phenomenon isnot associated with phosphorylation of LCHII polypeptides. Apossible role for cytochrome b₆/f complexes in the phosphorylation-inducedinhibition of whole chain electron transport is discussed. Key words: Electron transport, light harvesting, photosystem 2, protein phosphorylation, thylakoid membranes, wheat (Triticum aestivum)     

Photo-inhibition and the Effects of Protein Phosphorylation on Electron Transport in Wheat Thylakoids

The kinetics of changes in photosystem I (PSI), photosystemII (PSII), and whole chain (PSII and PSI) electron transport,chlorophyll fluorescence parameters, the capacity to bind atrazineand the polypeptide profiles of thylakoids isolated from wheatleaves on exposure to a photon flux density of 2000 µmolm^–2 s^–1 were determined. Severe and similar levelsof photo-inhibitory damage to both PSII and whole chain electrontransport occurred and were correlated with decreases in theratio of variable to maximal fluorescence, the proportionalcontribution of the rapid a phase of the fluorescence kineticsand the capacity to bind atrazine. Severe photo-inhibition ofelectron transport was not associated with a major loss of chlorophyllor total thylakoid protein. However, a small decrease in a 70kDa polypeptide together with increases in a number of low molecularmass polypeptides (8–24 kDa) occurred. Phosphorylation of thylakoid polypeptides alleviated photo-inhibitionof PSII electron transport but stimulated photoinhibitory damageto whole chain electron transport. The consequences of suchphosphorylation-induced effects on photoinhibition in vivo areconsidered. Key words: Chlorophyll fluorescence, electron transport, photo-inhibition, protein phosphorylation, thylakoid membranes, wheat (Triticum aestivum)     

Repressible chloroplast gene expression in Chlamydomonas: A new tool for the study of the photosynthetic apparatus

A repressible/inducible chloroplast gene expression system has been used to conditionally inhibit chloroplast protein synthesis in the unicellular alga Chlamydomonas reinhardtii. This system allows one to follow the fate of photosystem II and photosystem I and their antennae upon cessation of chloroplast translation. The main results are that the levels of the PSI core proteins decrease at a slower rate than those of PSII. Amongst the light-harvesting complexes, the decrease of CP26 proceeds at the same rate as for the PSII core proteins whereas it is significantly slower for CP29, and for the antenna complexes of PSI this rate is comprised between that of CP26 and CP29. In marked contrast, the components of trimeric LHCII, the major PSII antenna, persist for several days upon inhibition of chloroplast translation. This system offers new possibilities for investigating the biosynthesis and turnover of individual photosynthetic complexes in the thylakoid membranes. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: Keys to Produce Clean Energy.     

Core protein phosphorylation facilitates the repair of photodamaged photosystem II at high light

Phosphorylation of photosystem II (PSII) reaction center protein D1 has been hypothesised to function as a signal for the migration of photodamaged PSII core complex from grana membranes to stroma lamellae for concerted degradation and replacement of the photodamaged D1 protein. Here, by using the mutants with impaired capacity (stn8) or complete lack (stn7 stn8) in phosphorylation of PSII core proteins, the role of phosphorylation in PSII photodamage and repair was investigated. We show that the lack of PSII core protein phosphorylation disturbs the disassembly of PSII supercomplexes at high light, which is a prerequisite for efficient migration of damaged PSII complexes from grana to stroma lamellae for repair. This results in accumulation of photodamaged PSII complexes, which in turn results, upon prolonged exposure to high light (HL), in general oxidative damage of photosynthetic proteins in the thylakoid membrane.     

Concerning a dual function of coupled cyclic electron transport in leaves

Coupled cyclic electron transport is assigned a role in the protection of leaves against photoinhibition in addition to its role in ATP synthesis. In leaves, as in reconstituted thylakoid systems, cyclic electron transport requires “poising,” i.e. availability of electrons at the reducing side of photosystem I (PSI) and the presence of some oxidized plastoquinone between photosystem II (PSII) and PSI. Under self-regulatory poising conditions that are established when carbon dioxide limits photosynthesis at high light intensities, and particularly when stomata are partially or fully closed as a result of water stress, coupled cyclic electron transport controls linear electron transport by helping to establish a proton gradient large enough to decrease PSII activity and electron flow to PSI. This brings electron donation by PSII, and electron consumption by available electron acceptors, into a balance in which PSI becomes more oxidized than it is during fast carbon assimilation. Avoidance of overreduction of the electron transport chain is a prerequisite for the efficient protection of the photosynthetic apparatus against photoinactivation.     

Effects of Drought Stress on the Photosynthesis in Maize

To clarify how the components of the entire photosynthetic electron transport chain in response to drought stress in maize. The activities of photosystem II (PSII), photosystem I (PSI), and the electron transport chain between PSII and PSI of maize were investigated by prompt fluorescence (PF), delayed fluorescence (DF) and 820 nm modulated reflection (MR). Maize (Zea mays L.) plants were subjected to different levels of soil water availability including control, moderate and severe drought stress. A significant decrease in ?E₀, Ψ₀ and PI_ABS was found in maize treated with moderate drought stress. A significant increase in ABS/RC was observed, but there were no significant change in the fast MR phase and the amplitude of DF under moderate drought stress compared to the control. Under severe drought stress, the exchange capacity between Q_A to Q_B, reoxidation capacity of plastoquinol, and the oxidation and re-reduction rates of PC and P₇₀₀ all decreased. These results demonstrated that moderate drought stress reduced the photochemical activity of PSII from Q_A to PQH₂, while the photochemical activity of PSI was unscathed. However, severe drought stress inhibited the entire electron transport chain from the donor side of PSII to PSI-end electron acceptors. In addition, the photochemical activity of PSII is more sensitive to drought stress than PSI.     

Changes in cyclic and respiratory electron transport by the movement of phycobilisomes in the cyanobacterium Synechocystis sp. strain PCC 6803

Phycobilisomes (PBS) are the major accessory light-harvesting complexes in cyanobacteria and their mobility affects the light energy distribution between the two photosystems. We investigated the effect of PBS mobility on state transitions, photosynthetic and respiratory electron transport, and various fluorescence parameters in Synechocystis sp. strain PCC 6803, using glycinebetaine to immobilize and couple PBS to photosystem II (PSII) or photosystem I (PSI) by applying under far-red or green light, respectively. The immobilization of PBS at PSII inhibited the increase in cyclic electron flow, photochemical and non-photochemical quenching, and decrease in respiration that occurred during the movement of PBS from PSII to PSI. In contrast, the immobilization of PBS at PSI inhibited the increase in respiration and photochemical quenching and decrease in cyclic electron flow and non-photochemical quenching that occurred when PBS moved from PSI to PSII. Linear electron transport did not change during PBS movement but increased or decreased significantly during longer illumination with far-red or green light, respectively. This implies that PBS movement is completed in a short time but it takes longer for the overall photosynthetic reactions to be tuned to a new state.     

Response of soybean photosynthesis and chloroplast membrane function to canopy development and mutual shading

The effect of natural shading on photosynthetic capacity and chloroplast thylakoid membrane function was examined in soybean (Glycine max. cv Young) under field conditions using a randomized complete block design. Seedlings were thinned to 15 plants per square meter at 20 days after planting. Leaves destined to function in the shaded regions of the canopy were tagged during early expansion at 40 days after planting. To investigate the response of shaded leaves to an increase in available light, plants were removed from certain plots at 29 or 37 days after tagging to reduce the population from 15 to three plants per square meter and alter the irradiance and spectral quality of light. During the transition from a sun to a shade environment, maximum photosynthesis and chloroplast electron transport of control leaves decreased by two- to threefold over a period of 40 days followed by rapid senescence and abscission. Senescence and abscission of tagged leaves were delayed by more than 4 weeks in plots where plant populations were reduced to three plants per square meter. Maximum photosynthesis and chloroplast electron transport activity were stabilized or elevated in response to increased light when plant populations were reduced from 15 to three plants per square meter. Several chloroplast thylakoid membrane components were affected by light environment. Cytochrome f and coupling factor protein decreased by 40% and 80%, respectively, as control leaves became shaded and then increased when shaded leaves acclimated to high light. The concentrations of photosystem I (PSI) and photosystem II (PSII) reaction centers were not affected by light environment or leaf age in field grown plants, resulting in a constant PSII/PSI ratio of 1.6 ± 0.3. Analysis of the chlorophyll-protein composition revealed a shift in chlorophyll from PSI to PSII as leaves became shaded and a reversal of this process when shaded leaves were provided with increased light. These results were in contrast to those of soybeans grown in a growth chamber where the PSII/PSI ratio as well as cytochrome f and coupling factor protein levels were dependent on growth irradiance. To summarize, light environment regulated both the photosynthetic characteristics and the timing of senescence in soybean leaves grown under field conditions.     

Location,expression and orientation of the putative chlororespiratory enzymes,Ndh and IMMUTANS,in higher-plant plastids

The chloroplasts of many plants contain not only the photosynthetic electron transport chain, but also two enzymes, Ndh and IMMUTANS, which might participate in a chloroplast respiratory chain. IMMUTANS encodes a protein with strong similarities to the mitochondrial alternative oxidase and hence is likely to be a plastoquinol oxidase. The Ndh complex is a homologue of complex I of mitochondria and eubacteria and is considered to be a plastoquinone reductase. As yet these components have not been purified to homogeneity and their expression and orientation within the thylakoid remain ill-defined. Here we show that the IMMUTANS protein, like the Ndh complex, is a minor component of the thylakoid membrane and is localised to the stromal lamellae. Protease digestion of intact and broken thylakoids indicates that both Ndh and IMMUTANS are orientated towards the stromal phase of the membrane in Spinacia oleracea L. Such an orientation is consistent with a role for the Ndh complex in the energisation of the plastid membrane. In expression studies we show that IMMUTANS and the Ndh complex are present throughout the development of both Pisum sativum L. cv Progress No. 9 and Arabidopsis thaliana (L.) Heynh. leaves, from early expansion to early senescence. Interestingly, both the Ndh complex and the IMMUTANS protein accumulate within etiolated leaf tissue, lacking the photosystem II complex, consistent with roles outside photosynthetic electron transport.Abbreviations PQ plastoquinone - PSI, PSII photosystem I, II     

Inhibition of photosynthetic electron transport by palmitoleic acid is partially correlated to loss of thylakoid membrane proteins

Palmitoleic acid (16:1^Δ9), a monounsaturated fatty acid, is found to inhibit electron transport. Inhibition occurs rapidly (within 30 s). The oxidizing side of photosystem (PS) II is ~ 90% inactivated, whereas no inhibition occurs on the reducing side of the PSII complex. PSI activity is ∼ 65% inhibited. Inhibition of electron transport is not correlated to lipid peroxidation. 16:1 causes the loss of proteins from the thylakoid membrane which is exacerbated by the light. The loss of five proteins, viz. plastocyanin (PC), manganese stabilizing protein (MSP), cytochrome f (Cyt f), D1 and D2 is examined. The proteins are found to be lost in the following order: PC (< 1 min), MSP (~5 min), Cyt f (~10 min), D1 (~60 min) and D2 (~60 min). The timing of the loss of a PSI associated protein, PC, overlaps with that of the inhibition of PSI. Also, preventing the loss of PC with spermine (spm) treatment can partially protect PSI from 16:1 inhibition. Thus, the results suggest that the loss of PC is the cause of PSI inhibition. On the other hand, the loss of MSP from the oxygen-evolving complex is not likely to be the cause of the 16:1 inhibition on the oxidizing side of PSII, as the MSP loss occurs later than the inhibition. Also, increased retention of MSP with spm treatment does not relieve the 16:1 inhibition of PSII. Instead, inhibition of PSII by 16:1 is prevented by Mn²⁺ treatment, suggesting that loss of Mn²⁺ ions is the cause of PSII inhibition.