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 ∈-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 Lcm, were isolated from S. platensis. Spectroscopic properties of the isolated PBSs kept at 20 °C for various times showed that the connection between the APC and the APC B or Lcm 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. This revised version was published online in June 2006 with corrections to the Cover Date.     

Photoinhibition induced alterations in energy transfer process in phycobilisomes of PS II in the cyanobacterium, Spirulina platensis

Exposure of algae or plants to irradiance from above the light saturation point of photosynthesis is known as high light stress. This high light stress induces various responses including photoinhibition of the photosynthetic apparatus. The degree of photoinhibition could be clearly determined by measuring the parameters such as absorption and fluorescence of chromoproteins. In cyanobacteria and red algae, most of the photosystem (PS) II associated light harvesting is performed by a membrane attached complex called the phycobilisome (PBS). The effects of high intensity light (1000-4000 micromol photons m(-2) s(-1)) on excitation energy transfer from PBSs to PS II in a cyanobacterium Spirulina platensis were studied by measuring room temperature PC fluorescence emission spectra. High light (3000 micromol photons m(-2) s(-1)) stress had a significant effect on PC fluorescence emission spectra. On the other hand, light stress induced an increase in the ratio of PC fluorescence intensity of PBS indicating that light stress inhibits excitation energy transfer from PBS to PS II. The high light treatment to 3000 micromol photons m(-2) s(-1) caused disappearance of 31.5 kDa linker polypeptide which is known to link PC discs together. In addition we observed the similar decrease in the other polypeptide contents. Our data concludes that the Spirulina cells upon light treatment causes alterations in the phycobiliproteins (PBPs) and affects the energy transfer process within the PBSs.     

High temperature induced alterations in energy transfer in phycobilisomes of the cyanobacterium <Emphasis Type="Italic">Spirulina platensis</Emphasis>

Exposure of intact cells of Spirulina to high temperature (HT) stress (40–60 °C) caused decrease in absorption spectrum and fluorescence emission spectrum. Low temperature emission spectra were altered at phycocyanin (PC) level. Room and low temperature emission spectra of intact phycobilisomes showed that PC was the main target in this cyanobacterium for the altered energy transfer under HT.     

钝顶螺旋藻低中温适应型新品系的选育及RAPD分析

【目的】选育低中温适应型螺旋藻新品系,显著拓展螺旋藻工厂化培植的地理范围及时间、提高产量和降低成本。【方法】以用于工厂化培植的钝顶螺旋藻(Spirulina platensis) ZJU0116为出发品系,用组织匀浆和离心分离法制得其单细胞或原生质球,先以0.6%甲基磺酸乙酯(ethyl methanesulfonate, EMS)处理30 min,再用2.4 kGy的⁶⁰Co γ射线辐照,进行低温逆境筛选、5次冷/热(12 ℃/38 ℃)骤变交替处理、藻丝单体分离培养、温度适应面构建和蛋白质含量等检测及生产培植试验。【结果】获得了1株蛋白质含量与ZJU0116的相当、而温度适应面和平均日产量依次提高10.7%和10.9%的低中温适应型突变体,命名为ZJU0116(LMTA)。光学显微形态与随机扩增的多态性DNA (randomly amplified polymorphic DNA, RAPD)分析结果显示,与其亲本ZJU0116相比,ZJU0116(LMTA)藻丝的螺旋数和长度依次减少54.4%和42.1%,螺距增加28.8%;基因组DNA在随机引物S30的扩增产物中多了1条约470 bp的条带。【结论】ZJU0116(LMTA)在工厂化培植中温度适应性好、生产性状稳定,干藻粉产量可增加10%以上,可为当前螺旋藻产业深度融合“双碳”目标、迈向高质量发展新阶段提供必要支撑。     

Effects of proteinase K on the energy transfer between phycobiliproteins in phycobilisomes

Spectral changes in fluorescence of phycobilisomes (PBS) of A. variabilis treated with proteinase K were studied at room and liquid nitrogen temperature. In control PBS, the relative yield of 77 K fluorescence of F686 was very high, and those of F655 and F666 were low. In PBS treated with proteinase K for less than 1 h, F686 decreased, and F655 and F666 increased. In PBS treated with proteinase K for 2 h, F655 was the main peak of fluorescence emission, F686 was the second peak, the fluorescence emission peak of F666 disappeared. In PBS treated with proteinase K for more than 8 h, F655 showed only one fluorescence emission peak.We suggested that phycobiliporteins in the PBS of A. variabilis constitute an energy transfer chain, shown as follows:{fx91-1}The linkages between APC and APC-B, C-PC and APC, and C-PC and APC-B had different sensitivity towards proteinase K.     

Thermal degradation kinetics of the phycocyanin from Spirulina platensis

The cyanobacterium Spirulina platensis is a source of pigments, such as phycocyanin, which is used in the food, cosmetic and pharmaceutical industries. The thermal degradation kinetics of the liquid extract at pH values of 5, 6 and 7 was studied, evaluating its stability between 50 and 65 °C. The kinetic model was assumed and validated as being of the first order. Between 50 and 55 °C the extract was more stable at pH 6 and between 57 and 65 °C at pH 5, but was shown to be increasingly unstable at pH 7 as the temperature of the treatment increased. The addition of sorbitol between 10 and 50% (w/w) in the treatment at 62 °C for 30 min increased the half-life values of the phycocyanin extract, proving that its de-colorization was related to degradation of the protein chain.     

Effect of High Temperature on Photosynthetic Electron Transport Activities of the Cyanobacterium Spirulina Platensis

The activities of photosystem 2 (PS2) and whole chain electron transport declined in high temperature treated cells at the room temperature beyond 35 °C, while photosystem 1 (PS1) showed increased activity. Thylakoid membrane studies did not exhibit increase in PS1 activity indicating that the enhancement of PS1 activity is due to permeability change of cell membranes. However, the electron transport activity measured from reduced duroquinone to methylviologen which involves intersystem electron transport was extremely sensitive to high temperature. The activity of PS2 at different irradiance, which was accompanied by alterations in absorption and fluorescence emission properties, indicated changes in the energy transfer processes within phycobilisomes. Thus high temperature has multiple target sites in photosynthetic electron transport system of Spirulina platensis.     

Mapping the excitation energy migration pathways in phycobilisomes from the cyanobacterium Acaryochloris marina

In this study, we use ultrafast time-resolved absorption and fluorescence spectroscopies to examine A. marina phycobilisomes isolated from cells grown under light of different intensities and spectral regimes. Investigations were performed at room temperature and at 77?K. The study demonstrates that if complexes are stabilized by high phosphate (900?mM) buffer, there are no differences between them in temporal and spectral properties of fluorescence. However, when the complexes are allowed to disassemble into trimers in low phosphate (50?mM) buffer, differences are clearly observed. The fluorescence properties of intact or disassembled phycobilisomes from cells grown in low intensity white light are unresponsive to variation in phosphate concentration. This antenna complex was further studied in detail with application of femtosecond time-resolved absorption at room temperature. Combined spectroscopic and kinetic analysis of time-resolved fluorescence and absorption data of this antenna allowed us to identify spectrally different forms of phycocyanobilins and to propose a simplified model of how they could be distributed within the phycobilisome structure.     

Cloning and characterization of the gene encoding an esterase from Spirulina platensis

The gene encoding a 23 kDA serine esterase from the cyanobacterium Spirulina platensis has been identified, cloned, characterized and expressed in Escherichia coli. The primary structure of the esterase deduced from the DNA sequence displayed 32% sequence identity with the carboxylesterase (esterase II) encoded by estB of Pseudomonas fluorescens; the highest degree of homology is found in a stretch of 11 identical or highly conserved amino acid residues corresponding to the GXSXG consensus motif found in the catalytic site of many serine proteases, lipases and esterases.     

固定化培养中氮磷浓度对钝顶螺旋藻生长及其代谢产物和叶绿素荧光参数的影响

该研究以钝顶螺旋藻为实验对象,探究培养基中不同氮浓度(5、15、30、45 mmol·L^-1)和磷浓度(0.5、1.5、3.0、4.5 mmol·L^-1)对固定化生物膜培养模式下螺旋藻生长及其代谢产物和叶绿素荧光参数的影响,为生产实际中规模化高效培养螺旋藻提供理论依据。结果表明：(1)随着氮、磷浓度的升高,螺旋藻的生物量密度和产率先增高后降低,并在30 mmol·L^-1氮和3.0 mmol·L^-1磷处理时达到最大值;低氮、低磷条件会影响螺旋藻的藻体结构,表现为单细胞藻丝变短,螺旋变少,导致生物膜细胞密度降低。(2)与生物量变化趋势一致,螺旋藻光合色素含量随着培养基中氮、磷浓度的增加呈先升后降的变化趋势,且均在30 mmol·L^-1氮或者3.0 mmol·L^-1磷条件下达到最大值。(3)培养基中磷浓度固定时,螺旋藻的PSⅡ反应中心最大光能转化效率(F_v/F_m)和电子传递速率(ETR)随着氮浓度增加而增大,氮的缺乏会...     

螺旋藻有效成分超临界萃取与分离研究

使用超临界CO2萃取技术对钝顶螺旋藻粉进行萃取与分离,结果表明:超临界CO2萃取的最优条件为压力20Mpa,夹带剂使用量100mL/100g,温度50℃,萃取时间2h;利用凯氏定氮法及气相色谱法分别对超临界萃取处理过的螺旋藻中蛋白质和多糖进行研究与分析,结果表明:螺旋藻粉中蛋白质含量为49.39%,螺旋藻多糖中的单糖含量占多糖粗品的9.25%,主要组成为鼠李糖、木糖、甘露糖、葡萄糖、半乳糖等。     

Cloning and expression of the genes for ribulose-1,5-bisphosphate carboxylase from Spirulina platensis

The genes for the large and small subunits of ribulose-1,5-bisphosphate carboxylase have been cloned from the filamentous cyanobacterium Spirulina platensis. The two genes, located very closely on a 4.6 kbp DNA fragment, appear to be expressed although to a different extent in minicells of Escherichia coli. The amount of large subunit produced in the bacterial host represents at least 10% of the total protein.     

Characterization of the str operon genes from Spirulina platensis and their evolutionary relationship to those of other prokaryotes

Summary A 5.3 kb DNA segment containing the str operon (ca. 4.5 kb) of the cyanobacterium Spirulina platensis has been sequenced. The str operon includes the structural genes rpsL (ribosomal protein S12), rpsG (ribosomal protein S7), fus (translation elngation factor EF-G) and tuf (translation elongation factor EF-Tu). From the nucleotide sequence of this operon, the primary structures of the four gene products have been derived and compared with the available corresponding structures from eubacteria, archaebacteria and chloroplasts. Extensive homologies were found in almost all cases and in the order S12>EF-Tu>EF-G>S7; the largest homologies were generally found between the cyanobacterial proteins and the corresponding chloroplast gene products. Overall codon usage in S. platensis was found to be rather unbiased.     

Energy transfer processes in Gloeobacter violaceus PCC 7421 that possesses phycobilisomes with a unique morphology

We examined energy transfer dynamics in phycobilisomes (PBSs) of cyanobacteria in relation to the morphology and pigment compositions of PBSs. We used Gloeobacter violaceus PCC 7421 and measured time-resolved fluorescence spectra in three types of samples, i.e., intact cells, PBSs, and rod assemblies separated from cores. Fremyella diplosiphon, a cyanobacterial species well known for its complementary chromatic adaptation, was used for comparison after growing under red or green light. Spectral data were analyzed by the fluorescence decay-associated spectra with components common in lifetimes with a time resolution of 3 ps/channel and a spectral resolution of 2 nm/channel. This ensured a higher resolution of the energy transfer kinetics than those obtained by global analysis with fewer sampling intervals. We resolved four spectral components in phycoerythrin (PE), three in phycocyanin (PC), two in allophycocyanin, and two in photosystem II. The bundle-like PBSs of G. violaceus showed multiple energy transfer pathways; fast (≈ 10 ps) and slow (≈ 100 ps and ≈ 500 ps) pathways were found in rods consisting of PE and PC. Energy transfer time from PE to PC was two times slower in G. violaceus than in F. diplosiphon grown under green light.     

低温胁迫对转新疆雪莲sikRbcs2基因烟草光合作用的影响

研究分布于新疆的雪莲(Saussurea involucrata)中的sikRbcs2基因在低温条件下对植物光合作用的影响,以16、10、6、4℃温度梯度处理非转基因型和转sikRbcs2基因型烟草,每个温度处理72 h,比较研究其叶绿素荧光特性和光合特性。实验分析结果:低温胁迫下,转基因型烟草叶片叶绿素a(Chla)、叶绿素b(Chlb)、叶绿素a+b(Chla+Chlb)、类胡萝卜素(Car)含量都显著高于非转基因型烟草。叶绿素荧光参数分析:低温胁迫下,转基因烟草PSⅡ最大光化学效率(Fv/Fm)、q P(光化学猝灭系数)、ETR(电子传递效率)都显著或极显著高于非转基因型,光合参数测定:随着低温胁迫程度的加剧,转基因烟草和非转基因型烟草净光合速率(Pn)、气孔导度(Gs)、蒸腾速率(Tr)、胞间CO_2浓度(Ci)总体都呈下降趋势,但转基因烟草的净光合速率有一个明显的动态变化,先急剧下降后有一个稳定上升的趋势。通过对生长指标的测定发现:在低温处理后和恢复后转基因烟草生物量的积累都显著高于非转基因型烟草。研究结果表明:转新疆雪莲sikRbcs2基因的烟草在低温条件下具有较高的Fv/Fm、q P、ETR,对光合机构的损伤小,降低了低温胁迫效应,提高了烟草在低温胁迫条件下的耐受性。     

CO2浓度倍增对小麦叶绿体超微结构、超分子结构及光谱特性的影响

超薄切片及冰冻撕裂电镜观察、吸收光谱及77 K低温荧光发射光谱的测定结果表明:CO2浓度倍增对小麦( Triticum aestivum L.)叶绿体的超微、超分子结构及光谱特性的影响均为正效应.具体反映在:(1)小麦叶绿体中除了比对照积累有较多的淀粉粒外,其基粒和基质类囊体膜发育较好;(2)叶绿体的光合膜系,无论是垛叠和非垛叠膜区,其镶嵌于内质膜撕裂面(EFs和EFu)及原生质膜撕裂面(PFs和PFu)的功能蛋白粒均比其对照的发育良好,尤其PFs 与EFs面较为突出,即它们除了所含蛋白粒的密度较大外,在EFs面上有时还呈现出密集有序的阵列结构;(3)叶绿体整个吸收谱带,尤其红区和蓝区的主峰均较其对照有较大的光吸收,表明对光能的捕获能力明显高于对照;(4)无论是以436 nm还是以480nm波长激发的,其叶绿体的F684/F733 (PSⅡ/PSⅠ)的比值均较对照的高,表明CO2浓度倍增条件下生长的小麦叶片叶绿体的PSⅡ相对荧光强度有所增强,这与叶绿体的超微、超分子结构及吸收光谱的测定结果相一致.以上结果可为小麦在高CO2浓度下增产提供理论依据.     

CO2浓度倍增对小麦叶绿体超微结构、超分子结构及光谱特性的影响

超薄切片及冰冻撕裂电镜观察、吸收光谱及 77K低温荧光发射光谱的测定结果表明 :CO2 浓度倍增对小麦(TriticumaestivumL .)叶绿体的超微、超分子结构及光谱特性的影响均为正效应。具体反映在 :(1)小麦叶绿体中除了比对照积累有较多的淀粉粒外 ,其基粒和基质类囊体膜发育较好 ;(2 )叶绿体的光合膜系 ,无论是垛叠和非垛叠膜区 ,其镶嵌于内质膜撕裂面 (EFs和EFu)及原生质膜撕裂面 (PFs和PFu)的功能蛋白粒均比其对照的发育良好 ,尤其PFs与EFs面较为突出 ,即它们除了所含蛋白粒的密度较大外 ,在EFs面上有时还呈现出密集有序的阵列结构 ;(3)叶绿体整个吸收谱带 ,尤其红区和蓝区的主峰均较其对照有较大的光吸收 ,表明对光能的捕获能力明显高于对照 ;(4)无论是以 4 36nm还是以 4 80nm波长激发的 ,其叶绿体的F684/F73 3 (PSⅡ /PSⅠ )的比值均较对照的高 ,表明CO2 浓度倍增条件下生长的小麦叶片叶绿体的PSⅡ相对荧光强度有所增强 ,这与叶绿体的超微、超分子结构及吸收光谱的测定结果相一致。以上结果可为小麦在高CO2 浓度下增产提供理论依据。     

Correlation of the photosystem I and II reaction center chlorophyll-protein complexes, CP-a1 and CP-aII with photosystem activity and low temperature fluorescence emission properties in mutants of Scenedesmus

Abstract Comparative studies on the low temperature fluorescence emission of whole cells, purified chlorophyll-protein (CP) complexes and on patterns noted in sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) for chlorophyll-protein complexes and chloroplast membrane polypeptides of Scenedesmus obliquus with several distinct mutant classes has allowed further correlation between the fluorescence emission bands seen at 77K and the known chlorophyll-protein complexes. In mutants deficient in photosystem II (PS-II; total loss of the reducing side) the fluorescence emission spectra showed only two peaks, i.e., 686 and 718 nm, but in the wild type, in mutants lacking the oxidizing side of PS-II and in phenotypes missing the CP-a₁ complex (and P-700 activity) all three emission bands at 686, 696 and 718 nm were present. In a mutant lacking the light-harvesting CP-a/b complex the emission peak at 686 nm was strongly reduced and the longer wavelength emissions predominated. Gel electrophoresis studies showed that the PS-II (reducing side) mutants lacked the polypeptides of apparent molecular weight 54 and 51 kilodaltons and the chlorophyll-protein complex, CP-a_II, of apparent molecular weight 32 kilodaltons. Contrarily, the loss of the oxidizing side of PS-II did not result in any alteration of these components. Genetic deletion of CP-a₁ did not alter significantly the long wavelength emission even though the isolated CP-a₁ shows the low temperature-dependent long wavelength emission comparable to that seen in the whole cell. It was deduced that remaining PS-I antennae chlorophylls must account for the emission seen at 718 nm. The absence of the CP-a/b complex and the strong simultaneous decrease of the 686 nm emission strengthens the concept that this complex is the primary emitter of fluorescence at room temperature. Its absence facilitated the detection of the CP-a_II complex in SDS-PAGE and enhanced the in vivo fluorescence by the two photosystems. Parallel experiments with two mutants which green and develop, one to the wild-type and the other to the CP-a/b deficient phenotype, provided additional evidence for the source of the low temperature emission bands.     

Short term effect of ultraviolet-B radiation on photosystem 2 photochemistry in the cyanobacteriumSynechococcus 6301

Exposure of intact cells of theSynechococcus 6301 to UV-B radiation induced a loss in photosystem 2 (PS 2) electron transport activity prior to the alteration in pigment complexes. Thus the degradation of PS 2 was not directly related to pigment alteration. Supported by Young Scientist grant No. SRJSYIB-05192 from the Department of Science and Technology, Goverment of India to SDSM.     

The role of ApcD and ApcF in energy transfer from phycobilisomes to PS I and PS II in a cyanobacterium

The role of the phycobilisome core components, ApcD and ApcF, in transferring energy from the phycobilisome to PS I and PS II in the cyanobacterium Synechocystis sp. PCC6803 has been investigated. The genes encoding these proteins have been disrupted in the genomes of wild type Synechocystis sp. PCC6803 and a PS II deficient mutant, PsbD1CD2^-, by inserting antibiotic resistance genes into their coding regions. Data from fluorescence emission spectra and pigment content analysis for these inactivation mutants is presented. These data suggest that both ApcD and ApcF are involved in the energy transfer route to PS II and PS I. In both cases, the energy transfer may to the reaction centres may be via the chromophore of ApcE (the L cm) or anchor polypeptide). The major route of energy transfer to both kinds of reaction centre appears to involve ApcF rather than ApcD. When both ApcF and ApcD are absent, the phycobilisomes are unable to transfer energy to either reaction centre. We suggest a model for the pathways of energy transfer from the phycobilisomes to PS I and PS II.