ApcD is necessary for efficient energy transfer from phycobilisomes to photosystem I and helps to prevent photoinhibition in the cyanobacterium Synechococcus sp PCC 7002

Phyrobilisomes (PBS) are the major light-harvesting, protein-pigment complexes in cyanobacteria and red algae. PBS absorb and transfer light energy to photosystem (PS) II as well as PS I, and the distribution of light energy from PBS to the two photosystems is regulated by light conditions through a mechanism known as state transitions. In this study the quantum efficiency of excitation energy transfer from PBS to PS I in the cyanobacterium Synechococcus sp. PCC 7002 was determined, and the results showed that energy transfer from PBS to PS I is extremely efficient. The results further demonstrated that energy transfer from PBS to PS I occurred directly and that efficient energy transfer was dependent upon the allophycocyanin-B alpha subunit, ApcD. In the absence of ApcD, cells were unable to perform state transitions and were trapped in state 1. Action spectra showed that light energy transfer from PBS to PS I was severely impaired in the absence of ApcD. An apcD mutant grew more slowly than the wild type in light preferentially absorbed by phyrobiliproteins and was more sensitive to high light intensity. On the other hand, a mutant lacking ApcF, which is required for efficient energy transfer from PBS to PS II, showed greater resistance to high light treatment. Therefore, state transitions in cyanobacteria have two roles: (1) they regulate light energy distribution between the two photosystems; and (2) they help to protect cells from the effects of light energy excess at high light intensities. (C) 2009 Elsevier B.V. All rights reserved.

Characterization, structure and function of linker polypeptides in phycobilisomes of cyanobacteria and red algae: An overview

Cyanobacteria and red algae have intricate light-harvesting systems comprised of phycobilisomes that are attached to the outer side of the thylakoid membrane. The phycobilisomes absorb light in the wavelength range of 500-650 nm and transfer energy to the chlorophyll for photosynthesis. Phycobilisomes, which biochemically consist of phycobiliproteins and linker polypeptides, are particularly wonderful subjects for the detailed analysis of structure and function due to their spectral properties and their various components affected by growth conditions. The linker potypeptides are believed to mediate both the assembly of phycobiliproteins into the highly ordered arrays in the phycobilisomes and the interactions between the phycobilisomes and the thylakoid membrane. Functionally, they have been reported to improve energy migration by regulating the spectral characteristics of colored phycobiliproteins. In this review, the progress regarding linker polypeptides research, including separation approaches, structures and interactions with phycobiliproteins, as well as their functions in the phycobilisomes, is presented. In addition, some problems with previous work on linkers are also discussed. (c) 2005 Elsevier B.V. All rights reserved.

Interaction between phycobilisomes from Porphyridium cruentum and thylakoid membranes from Gymnodinium sp or spinach

Thylakoid membranes were isolated from Gymnodinium sp. and spinach, whereas the phycobilisomes were isolated and purified from red alga Porphyridium cruentum. The absorption spectra of the purified phycobilisomes (PBS) showed three peaks at 548, 564, and 624 nm, respectively, and the ratio of the fluorescence intensity at the lambda(680)(em) to lambda(80)(em5) that at was about 7.3. All these results demonstrated that the purified PBS remained intact. The thylakoid membranes were incubated with the purified phycobilisomes, and the thylakoid membranes, which harbored the phycobilisomes, were purified by sucrose density gradient centrifugation. Meantime, the conjugates of phycobilisome-thylakoid membranes were constructed using glutaraldehyde and further purified. Their characteristics were studied by measuring the absorption spectra and fluorescence emission spectra. The results showed that the phycobilisomes from Porphyridium cruentum can attach to the thylakoid membranes from Gymnodinium sp. and spinach without covalent cross-linking, but the excited energy transfer did not occur. The conjugate of phycobilisome-thylakoid. membranes with covalent cross-linking exhibits the excited energy transfer between the phycobilisomes and the thylakoid membranes.

The stabilization effect of glutaraldehyde on the Spirulina plateusis phycobilisomes

The spectral properties of the glutaraldehyde-treated phycobilisomes were studied. The results showed that glutaraldehyde was effective in preventing phycobilisomes from dilution-induced dissociation and preserving the intra-phycobilisomes energy transfer.

Langmuir-Blodgett film of phycobilisomes from blue-green alga Spirulina platensis

The phycobilisomes were isolated from blue-green alga Spirulina platensis, and could form monolayer film at air/water interface. The monolayer film of phycobilisomes was transferred to newly cleaved mica, and coated with gold. Scanning tunneling microscope was used to investigate the structure of the Langmuir-Blodgett film of phycobilisomes. It was shown that phycobilisomes in the monolayer arrayed in rows with core attaching on the substrate surface and rods radiating towards the air phase, this phenomenon was similar to the arrangement of phycobilisomes on cytoplasmic surface of thylakoid membrane in vivo. The possible applications of the Langmuir-Blodgett film of phycobilisomes were also discussed.

Spectroscopic properties of the C-phycocyanin-allophycocyanin conjugate and the isolated phycobilisomes from Spirulina platensis

C-phycocyanin (CPC) and allophycocyanin (APC) were purified from Spirulina platensis, then the CPC was attached covalently to the APC by reacting their epsilon-amino groups. The excitation energy could be transferred from the CPC to the APC in the CPC-APC conjugate. Intact phycobilisomes (PBS), consisting of CPC, APC, colourless linker polypeptides, and APC B or L-cm, were isolated from S. platensis. Spectroscopic properties of the isolated PBSs kept at 20 degrees C for various times showed that the connection between the APC and the APC B or L-cm was looser than that between the CPC and the APC in the isolated PBSs. The CPC-APC conjugate was more stable than the isolated PBSs, and the linker polypeptides had a minor influence on the excitation energy transfer characteristic between different phycobiliproteins in the PBS.

Phycobilisomes linker family in cyanobacterial genomes: divergence and evolution

Cyanobacteria are the oldest life form making important contributions to global CO2 fixation on the Earth. Phycobilisomes (PBSs) are the major light harvesting systems of most cyanobacteria species. Recent availability of the whole genome database of cyanobacteria provides us a global and further view on the complex structural PBSs. A PBSs linker family is crucial in structure and function of major light-harvesting PBSs complexes. Linker polypeptides are considered to have the same ancestor with other phycobiliproteins (PBPs), and might have been diverged and evolved under particularly selective forces together. In this paper, a total of 192 putative linkers including 167 putative PBSs-associated linker genes and 25 Ferredoxin-NADP oxidoreductase (FNR) genes were detected through whole genome analysis of all 25 cyanobacterial genomes (20 finished and 5 in draft state). We compared the PBSs linker family of cyanobacteria in terms of gene structure, chromosome location, conservation domain, and polymorphic variants, and discussed the features and functions of the PBSs linker family. Most of PBSs-associated linkers in PBSs linker family are assembled into gene clusters with PBPs. A phylogenetic analysis based on protein data demonstrates a possibility of six classes of the linker family in cyanobacteria. Emergence, divergence, and disappearance of PBSs linkers among cyanobacterial species were due to speciation, gene duplication, gene transfer, or gene loss, and acclimation to various environmental selective pressures especially light.

藻胆体的光谱特性和光能传递

一，螺旋藻藻胆体光谱特性及其光能传递的研究 1，完整藻胆体与解离藻胆体吸收光谱的比较研究 对螺旋藻完整藻胆体和解离藻胆体的吸收光谱中进行了比较研究。随着PBS逐渐解离，其吸收光谱表现出如下变化特点：在紫外区，吸收峰始终位于355nm，尖形峰逐渐变成钝形峰;在红区，完整藻胆体和解离藻胆体都有很强的光吸收，吸收峰呈平顶状，其半带宽逐渐变小，紫外区与红区相对吸收强度比值逐渐变小，四组导数吸收光谱中的小峰数目越来越少。室温荧光发射光谱表明，PBS在低于0.9mol/L的磷酸缓冲液中变得不稳定，并开始逐渐解离，解离的PBS与完整的PBS相比，其荧光发射峰逐渐蓝移。 2，藻胆体在解离过程中荧光发射和光能传递的研究 完整藻胆体的室温荧光发射光谱中只有一个峰，在678nm。说明在完整藻胆体中，光能传递效率高。在77K荧光发射光谱中，完整藻胆体只有一个峰，位于682nm，这是L_(cm)(TE_1)的荧光峰;严重解离的藻胆体的主峰在656nm，是PC的荧光;在679nm有一个小峰，是APC-B的荧光(TE_2)。据此，我们提出螺旋藻藻胆体的光能传递链为：(此处表从略，见全文) 二，螺旋藻藻胆体核心及其与藻蓝蛋白的重组 PC+core混合物，浓缩重组48h后，其室温荧光发射峰位于663nm，与PC的室温荧光发射峰643nm和PC+core混合物（未重组）的室温荧光发射峰648nm相比，说明部分APC与部分PC发生了重组，使部分PC吸收的光能传递给了APC，使荧光发射峰红移;与藻胆体核心室温荧光发射峰664nm相比，则非常接近，说明重组效果较好。PC+core混合物（未重组），其77K荧光发射光谱中有两个峰：654nm,679nm,分别是PC,APC-B的荧光峰，F679/F654的比值为32.0%。我们以F679/F654比值的变化来判断PC与core是否发生了重组。PC+core混合物，经48h浓缩重组后，77K荧光发射光谱中有F657,F679两个峰，F679/F654的比值则为45.9%，比未重组的混合物32.0%升高了，说明部分PC与core发生了重组，部分PC吸收的光能传递给了APC和APC-B，使F679加强，F654减弱。 三，螺旋藻藻胆体一类囊体膜光谱特性与光能传递的研究 藻胆体一类囊体膜的吸收光谱，室温荧光发射光谱和77K荧光发射光谱表明：藻胆蛋白能将捕获的光能传递给叶绿素a，叶绿素a捕获的光能不能逆传给藻胆蛋白。 四，藻胆体一类囊体膜的重组 藻胆体一类囊体膜的吸收光谱说明，一部分被洗下来的PBS能重新结合到类囊体膜上，但并没有达到100%的重组。 五，整体螺旋藻光谱特性及其光能传递的研究 整体螺旋藻光谱特性与PBS-类囊体膜的光谱特性极为相似，表现出同样的规律：PBS的吸收面积与叶绿素a相比，叶绿素a的吸收是主要的。 从PBS-类囊体膜和整体螺旋藻的吸收光谱，室温荧光发射光谱，77K荧光发射光谱的研究中可知，二者表现出极为相似的规律：PBS藻胆蛋白捕获的光能能传递给叶绿素a，叶绿素a捕获的光能不能逆传给PBS藻胆蛋白。主要的捕光物质是叶绿素a。 另外，我们还对Spirulina platensis 6 and Spirulina maxima的藻胆体在解离过程中的荧光发射和光能传递进行了研究，表现规律与Spirulina platensis相同。

藻胆体的光谱特性光能传递

1．对嗜热蓝藻层理鞭枝藻(Mastigocladus laminosus)藻胆体的光谱特性和光能传递进行了研究。其完整藻胆体的吸收峰位于622 nm，室温荧光发射峰位于673 nm。在77K荧光发射光谱中，完整藻胆体的荧光峰只有一个，位于685 nm，是末端发射体1的荧光。在低浓度磷酸缓冲液中发生严重解离的藻胆体，其77K荧光发射光谱中有二个发射峰和一个发射肩。两个荧光发射峰分别位于644 nm和683 nm。前者为主峰，属于C-藻蓝蛋白的荧光，后者是次峰，属于末端发射体2的荧光。荧光发射肩位于660 nm附近，属于别藻蓝蛋白的荧光。据此，提出层理鞭枝藻藻胆体光能传递途径如下： 藻红蓝蛋白→c—藻蓝蛋白→别藻蓝蛋白→端发射体l、末端发射体2： 2．对嗜热蓝藻层理鞭枝藻藻胆体—类囊体膜的光谱特性和光能传递进行了研究。在吸收光谱中，其藻胆体——类囊体膜在可见光区域有5个峰，它们分别位于420 nm、438 nm、490 nm、624 nm和678 nm。420 nm、438 nm和678 nm为叶绿素a的吸收峰位置。490 nm是类胡罗卜素的吸收峰，624 nm是藻胆体的吸收峰。对藻胆体——类囊体膜用580 nm波长的光激发藻胆蛋白时，在室温荧光发射光谱中有一个发射峰和一个发射肩，分别位于657 nm和690 nm，前者属于藻胆蛋白的荧光，后者属于叶绿素a的荧光。这说明藻胆蛋白能将捕获的光能传递给类囊体膜上的叶绿素a。在77K荧光发射光谱中有4个峰，它们分别位于649 nm、660 nm、688 nm和730 nm。前二者属于藻胆蛋白的荧光，后二者属于叶绿素a的荧光。这同样说明藻胆蛋白能将捕获的光能传递给类囊体膜上的叶绿素a。当用436 nm波长光激发叶绿素a时，藻胆体——类囊体膜的室温荧光发射光谱中有两个荧光峰出现，位于685 nm的峰来源于光系统Ⅱ，位于713 nm的峰来源于系统I。这说明叶绿素a捕获的光能不能逆传递给藻胆体中的藻胆蛋白。在77K荧光发射光谱中也只有叶绿素a的荧光峰，位于695 nm的峰来源于光系统Ⅱ，位于730 nm的峰来源于光系统l。此结果同样说明叶绿素a捕获的光能不能逆传递给藻胆蛋白． 3．我们以多变鱼腥藻(Anabaena variabilis)为材料，对其藻胆体核心和藻蓝蛋白进行了重组实验，得到了具有光能传递效率的藻胆体核心——藻蓝蛋白复合物。在吸收光谱中，藻胆体核心有一吸收峰和一个吸收肩，分别位于654 nm和600 nm。藻蓝蛋白的吸收光谱中只有一个峰，位于620 nm．重组样品的吸收光谱有一吸收峰和一吸收肩，分别位于654 nm和620 nm．由于620 nm与654 nm的吸收比远大于核心的600 nm与654 nm的吸收比，因此，可以认为部分藻蓝蛋白已与核心重组。在室温荧光发射光谱中，藻胆体核心只有一个峰，位于676 nm。藻蓝蛋白只有一个峰，位于653 nm。重组样品有一荧光发射峰和一荧光发射肩，分别在669 nm和650 nm附近。669 nm荧光来源于核心，650 nm荧光来源于藻蓝蛋白。重组后的核心的650 nm荧光显著大于未重组的核心，这也说明部分藻蓝蛋白与核心已重组．在77K荧光发射光谱中，藻蓝蛋白只有一个峰，位于655 nm。藻胆体核心有二个峰，分别位于666 nm和686 nm。重组样品有两个荧光发射峰和一荧光发射肩，分别位于666 nm、683 nm和648 nm附近．重组的核心的别藻蓝蛋白的荧光(F666)和藻蓝蛋白的荧光(F648)都强于未重组的核心。这一结果同样说明有藻胆体——藻蓝蛋白复合物生成。 除以上研究工作之外，我们还对多变鱼腥藻藻胆体在解离过程中的光谱特性及光能传递、藻胆体——类囊体膜的光谱特性及光能传递、藻胆体解离重组、藻胆体核心在低浓度磷酸缓冲液中的光谱特性、以及温度对藻胆体核心的影响等进行了研究。研究结果有待整理。 本文编写：PBS：藻胆体;PEB：藻红胆素;PE：藻红蛋白;PUB:藻尿胆素;PEC：藻红蓝蛋白;PCB:藻蓝胆素;PC：藻蓝蛋白;PSⅡ：光系统Ⅱ;APC:别藻蓝蛋白;PS I：光系统I;TE:末端发射体

Factors that effect antioxidant activity of C-phycocyanins from Spirulina platensis

Currently, antioxidants are added in the human diet to prevent free radical-induced cell damage, and there has been an explosive interest in the use of antioxidant nutritional supplements. The effects of different factors on the antioxidant activity of phycocyanins (PCs) were studied. The results showed that PCs generated hydroxyl radicals in the light, while scavenging them in the dark. When PCs were denatured by sodium dodecyl sulfate, urea and in alkaline condition, their ability to generate hydroxyl radicals disappeared and that of scavenging them greatly increased. This showed that the phycobilin moiety is the main part of PC involved in scavenging hydroxyl radicals. Trypsin hydrolysis of PCs showed that the apoprotein portion of the molecule also made a significant contribution to the antioxidant activity.

In vitro assembly of R-phycoerythrin from marine red alga Polysiphonia urceolata

Scanning tunneling microscope was used to investigate the in vitro assembly of R-phycoerythrin (R-PE) from the marine red alga Polysiphonia urceolata. The results showed that R-PE molecules assembled together by disc-to-disc while absorbing on HOPG surface, which just looked like the rods in the phycobilisomes. When the water-soluble R-PE was dissolved in 2% ethanol/water spreading solution, they could form monolayer film at the air/water interface. Similar disc-to-disc array of R-PE was constituted in the two-dimensional Langmuir-Blodgett film by the external force. It could be concluded that, apart from the key role of time linker polypeptides, the in vivo assembly of phycobiliproteins into phycobilisomes is also dependent on the endogenous properties of phycobiliprotein themselves.

A new model of phycobilisome in Spirulina platensis

Phycobilisomes (PBS) were isolated from blue-green alga Spirulina platensis. Scanning tunneling microscope was used to investigate the three-dimensional structure of PBS deposited on freshly cleaved highly oriented pyrolytic graphite (HOPG) in ambient condition at room temperature. The results showed that the rods of PBS radiated from the core to different directions in the space other than arrayed in one plane, which was different from the typical hemi-discoidal model structure. The diameter of PBS was up to 70 nm, and the rod was approximately 50 nm in length. Similar results were observed in Langmuir-Blodgett (LB) film of PBS. The dissociated PBS could reaggregate into rod-like structures and easily form two-dimensional membrane while being absorbed on HOPG, however, no intact PBS was observed. The filling-space model structure of PBS in Spirulina platensis with STM from three-dimensional real space at nanometer scale was found, which showed that this new structural model of PBS surely exists in blue-green algae and red algae. The function of this structural model of PBS was also discussed.