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1.
Rhodosorus mairinus中藻红蛋白的纯化及其性质的研究   总被引:4,自引:0,他引:4  
从Rhodosorus marinus中提取了藻红蛋白,通过改进纯化方法,得到了三种电泳纯的藻红蛋白:B-型藻红蛋白1,B-型藻红蛋白2和b-型藻红蛋白(以下简称B-PE1,B-PE2和b-PE)。分别测定这三种藻红蛋白聚合体和亚单位的分子量。测定了它们的可见光的吸收光谱和荧光光谱。两种B-PE的可见光的吸收光谱比b-PE的多一个498nm的吸收峰,三种藻红蛋白的氨基酸组成以酸性氨基酸和疏水性氨基  相似文献   

2.
条斑紫菜中R-藻红蛋白的生化特性   总被引:5,自引:0,他引:5  
条斑紫菜的R-藻红蛋白(R-PE),在CM-52柱上用含8mol/L脲的0.02mol/L乙酸铵缓冲液(pH=5.05)洗脱,观察到3条色带,经吸收光谱测定表明,它们分别是α、β、γ亚基。用SDS-PAGE测定的α、β和γ亚基分子量分别是17.0kd,18.0kd和31.7kd。R-PE中亚基的摩尔比是6α:6β:1γ。条斑紫菜的R-PE最稳定的聚集态分子量是229kd。各亚基的发色团含量:α亚基含2个藻红胆素(PEB),β亚基含1个PEB和0.5个藻尿胆素(PUB),γ亚基含2个PEB和3个PUB。结合R-PE和各亚基的氨基酸组成分析,条斑紫菜的R-PE亚基组成是(αβ)6γ。  相似文献   

3.
兔阑尾中一种新的21kD的钙结合蛋白的纯化与鉴定   总被引:3,自引:0,他引:3  
纯化与鉴定了B淋巴细胞中一种新的分子量为21kD的钙结合蛋白(CaBP21)。兔阑尾淋巴细胞匀浆经热变性,Phenyl-Sepharose与DEAE-Sepharose柱层析,自每1kg细胞沉积物中获得SDS-PAGE均一的CaBP215.3mg。HCl水解后的酸性氨基酸(Asp+Glu)含量为26%。如同大多数钙结合蛋白一样,N末端封闭阻止其进行Edman降解。CaBP21中疏水性氨基酸(计Gly,不计Trp)约占46%,碱性氨基酸10%,酸性氨基酸与极性氨基酸约44%。CaBP21有较高的Ser、Tyr含量。肽谱分析等确证CaBP21为2个相同或相似亚基二聚体。以ArsenazoⅢ作Ca2+结合分析表明每分子CaBP21可结合4分子Ca2+,对Ca2+的结合常数约为10-5mol/L。各种性质表明CaBP21是一种不同于其他已知钙结合蛋白的新钙结合蛋白。  相似文献   

4.
几种海洋红藻中的R—藻红蛋白对胰鸟素抗体的免疫反…   总被引:1,自引:0,他引:1  
本文利用点免疫结合测定法研究了条斑紫菜,红毛菜,多管藻,海萝和江蓠中两种不同分子量的R-和r-藻红蛋白(PE)对胰岛素抗体的免疫反应性。这5种红藻中的R-藻红蛋白都可用蔗糖密度线性梯度超速离心法,得到分子量不同的两种R-PE。它们与胰岛素抗体都发生免疫结合反应,但用反射扫描色谱测定,在抗原量相同的比较实验中,反映它们免疫结合反应的棕色斑点的扫描峰面积有所不同。即它们与胰岛素抗体的结合率不同,其中海  相似文献   

5.
研究了优雅粘囊藻藻胆体(PBS)的特性。从聚丙烯酰胺凝胶电泳(PAGE)、吸收光谱和二阶导数图谱可证明,它的PBS含有一种红色藻胆蛋白,两种C-藻蓝蛋白和三种别藻蓝蛋白。但是,用PAGE和羟基磷灰石柱层析分离和纯化所得藻胆蛋白,一般仅得到一种C-PC和一种亚基组成特殊的别藻蓝蛋白APC。这种APC吸收光谱和荧光发射相同于已报告的AFC660nm。但是它的亚基组成是(αα′ββ′)2而不是(α′α2β2β′)Lc10或(αβ)3。  相似文献   

6.
光动力复合藻胆蛋白及其分子内能量传递现象   总被引:2,自引:0,他引:2  
通过杂双官能团偶联试剂,3-(2-吡啶联疏基)丙酸N-羟基琥珀酰亚胺酯(SPD),我们合成了R-藻红蛋白(R-PE)与变藻蓝蛋白(APC)的共轭复合物。这种光动力复合藻胆蛋白具有一些特别的光物理性质,如很大的stokes位移(约170um,490nm激发,665nm发出荧光)、高效的分子内能量传递效率等。复合物中R—PE与APC的摩尔比为14,且R—PE和APC在复合物中保持了它们各自的光谱性质。通过荧光发射光谱,我们观察到了这种复合藻胆蛋白的分子内能量传递现象,计算表明从R—PE到APC的分子内能量传递效率为65%。二硫苏糖醇(DTT)对复合物中联结R—PE与APC间的脂族二硫桥键的还原导致分子内能量传递的阻断这一现象进一步证实了复合物中存在分子内能量传递现象。根据Forster能量转移机理,计算得出给体与受体发色团间距离为72A,这一距离与两种蛋白的大小是基本相符的。  相似文献   

7.
研究了优雅粘囊藻藻胆体的特性。从聚丙烯酰胺凝胶电泳、吸收光谱和二阶导数图谱可证明,它的PBS含有一种红色藻胆蛋,两种C-藻蓝蛋白和三种别藻蓝蛋白。但是,用PAGE和羟基磷灰石柱层析分离和纯化所得藻胆蛋白,一般仅得到一种C-PC和一种亚基组成特殊的别藻蓝蛋白APC。这种APC吸收光谱和荧光发射相同于已报告的APC660nm。但是它的亚基组成是(αα'ββ')2而不是(α'α2β2β')Lc^10或α  相似文献   

8.
将人肝金属硫蛋白(MT)突变体β基因插入到中间载体pRL-439上的强启动子PpsbA 下游,利用载体pRL-β上的PpsbA 和β基因与phasm id pTZ18-8上整合平台PsbB,构建整合表达载体pTZ-β.整合平台PsbB与集胞藻(Synechocystissp.PCC6803)染色体DNA 上psbB基因下游片段为同源序列.为了发生单交换同源重组,将外源基因β插入到整合平台PsbB下游的克隆位点.利用自然转化方法将表达载体pTZ-β整合到Synechocystitsp.PCC6803的染色体上.经氨苄青霉素筛选得到遗传性状稳定的转基因蓝藻.Southern blotting 证明β基因已整合到Synechocystis sp.PCC6803的染色体上;Western blotting 表明β基因已在蓝藻中表达.ELISA 测定在Zn2+ 浓度为150 μm l/L时表达量最高,为590 μg/g 鲜藻;原子吸收表明转β基因的藻对Zn2+ 的富集能力约为野生型的2倍.  相似文献   

9.
鸡减蛋综合征病毒(EDSV—76)基因组E1区结构特点分析   总被引:1,自引:0,他引:1  
金奇  李茂祥 《病毒学报》1998,14(3):253-256
EDSV-76病毒中国株AA-2经常规方法提取其病毒DNA后,建立了限制性内切酶PstI水解片段的全基因文库。对其中PstI-G片段和PstI-A片段的正反链进行序列测定,获得EDSV E1区(0-8.8m.u)的核苷酸序列。经分析,EDSV E1区具有与其他腺病毒E1区类似的结构。以大于60个氨基酸残基为标准,EDSV E1区共有7个开放读码框架(ORF),其中R1、R2、ElbsT和E1b1T  相似文献   

10.
抗真菌蛋白Rs—AFPs基因在大肠杆菌中的表达   总被引:3,自引:0,他引:3  
将抗真菌蛋白Rs-AFP1和Rs-AFP2全长cDNA插入表达质粒pET-22b/NcoI+SacI位点,构建成融合蛋白表达载体pRAF1和pRAF2.将不含信号肽编码序列的Rs-AFP1和Rs-AFP2cDNA分别插入pET-22b/Ncol+Sacl和pET-22b/Ndel+SacI位点,构建成不含信号肽序列的融合蛋白表达载体pRAF3、pRAF4和非融合蛋白表达载体pRAF5和pRAF6.将构建的上述各种表达载体转化E.coliBL21,挑菌落培养,IPTG诱导,使Rs-AFPs基因得到表达,并用体外抑菌试验检测表达产物的活性,结果表明,各种表达载体的表达产物均具有不同程度的抑菌活性,其中,pRAF3和pRAF4表达产物的抑菌活性较明显.  相似文献   

11.
Two isoproteins of the "native" B-phycoerythrin of the red alga, Rhodella violacea, were purified from crude extracts by preparative polyacrylamide gel electrophoresis and subsequently characterized. The slower moving pigment in gel electrophoresis was designated B-PE I, the faster as B-PE II. Both were found to occur in about equal amounts. B-PE I has a molecular weight of about 280000 and an IEP at 4.39, B-PE II a molecular weight of nearly 265000 and an IEP at 4.23. B-PE I and II are characterized by absorption maxima at 568 and 542 nm and a shoulder at 500 nm in the visible part of the absorption spectra. Their absorption coefficients at 542 nm differ with values of 5.54 and 5.63, respectively. The fluorescence emission spectra show a single maximum at 575 for B-PE I and at 578 nm for B-PE II. Both spectra have a shoulder at 630 nm. The fluorescence yield of B-PE II is lower by 25%. In calibrated SDS gel electrophoresis of the purified pigments B-PE I and II show two subunits of molecular weights of 18900 and 29200 and 18500 and 29900, respectively. Quantitative amino acid analyses indicated, that the isoproteins are very similar. B-PE II, however, has a significantly higher content of acidic amino acids and a lower percentage of basic residues, which is in keeping with its lower isoelectric point. Functional aspects of the occurrence of two isoproteins of B-phycoerythrin are discussed.  相似文献   

12.

Background

Photosynthetic organisms have developed multiple protective mechanisms to prevent photodamage in vivo under high-light conditions. Cyanobacteria and red algae use phycobilisomes (PBsomes) as their major light-harvesting antennae complexes. The orange carotenoid protein in some cyanobacteria has been demonstrated to play roles in the photoprotective mechanism. The PBsome-itself-related energy dissipation mechanism is still unclear.

Methodology/Principal Findings

Here, single-molecule spectroscopy is applied for the first time on the PBsomes of red alga Porphyridium cruentum, to detect the fluorescence emissions of phycoerythrins (PE) and PBsome core complex simultaneously, and the real-time detection could greatly characterize the fluorescence dynamics of individual PBsomes in response to intense light.

Conclusions/Significance

Our data revealed that strong green-light can induce the fluorescence decrease of PBsome, as well as the fluorescence increase of PE at the first stage of photobleaching. It strongly indicated an energetic decoupling occurring between PE and its neighbor. The fluorescence of PE was subsequently observed to be decreased, showing that PE was photobleached when energy transfer in the PBsomes was disrupted. In contrast, the energetic decoupling was not observed in either the PBsomes fixed with glutaraldehyde, or the mutant PBsomes lacking B-PE and remaining b-PE. It was concluded that the energetic decoupling of the PBsomes occurs at the specific association between B-PE and b-PE within the PBsome rod. Assuming that the same process occurs also at the much lower physiological light intensities, such a decoupling process is proposed to be a strategy corresponding to PBsomes to prevent photodamage of the photosynthetic reaction centers. Finally, a novel photoprotective role of γ-subunit-containing PE in red algae was discussed.  相似文献   

13.
A simple general procedure is described for the purification of high molecular weight phycoerythrin from red algae. Protein of purity adequate for precise spectroscopic characterization was obtained from as little as 0.2 g wet wt of fresh algal tissue. The absorption, excitation and fluorescence emission spectra of over a hundred phycoerythrins from representatives of all of the orders of the Bangiophyceae and Florideophyceae were determined. On the basis of visible absorption spectra, the phycoerythrins were subdivided into five groups: B-phycoerythrin Type I [542 > 567 > 502(s)], B-phycoerythrin Type II [566 ? 528 > 500(s)], R-phycoerythrin Type I (565 > 543 > 497), R-phycoerythrin Type II [566 > 551(s) > 496], and R-phycoerythrin Type III (567 > 539 < 496), where the numbers in parentheses specify the absorption maxima in nm and (s) denotes shoulder. Phycoerythrins do not appear to be useful at familial, ordinal and class levels in taxonomic studies. However, they do appear to be of limited value in discriminating taxonomic groupings at the generic and specific level. Audouinella (Acrochaetiales) can be separated into two groups of species, B-PE and R-PE types, but this is not correlated with cytological, morphological or life-history patterns. Rhodophysema can be removed from the Cryptonemiales and placed in the Acrochaetiales on the basis of its B-PE content, morphology and life history.  相似文献   

14.
The discovery that the phycocyanobilin group attached to Cys-155 of the beta subunit of C-phycocyanin is D-ring linked (Bishop, J. E., Lagarias, J. C., Nagy, J. O., Schoenleber, R. W., Rapoport, H., Klotz, A. V., and Glazer, A. N. (1986) J. Biol. Chem. 261, 6790-6796) prompted examination of the linkage mode for phycoerythrobilin (PEB) groups attached at the corresponding position in other biliproteins. Appropriate small peptides were obtained by exhaustive enzymatic digestion of Porphyridium cruentum R-phycocyanin (peptide R-PC beta-2TP PEB) and B-phycoerythrin (peptide B-PE beta-2TP PEB). These peptides had the following structures R-PC beta-2TP PEB Gly-Asp-Cys(PEB)-Ser-Ser B-PE beta-2TP PEB Cys(PEB)-Thr-Ser. The spectroscopic and chemical properties of these peptides were compared with those of P. cruentum B-phycoerythrin peptide alpha-1 PEB, Cys(PEB)-Tyr-Arg, in which the bilin is A-ring linked (Schoenleber, R. W., Leung, S.-L., Lundell, D. J., Glazer, A. N., and Rapoport, H. (1983) J. Am. Chem. Soc. 105, 4072-4076). The PEB groups in peptides R-PC beta-2TP PEB and B-PE beta-2TP PEB were shown to be D-ring linked on the basis of the following criteria. Secondary ion mass spectrometry showed the bilins in these peptides and in alpha-1 PEB to have the same mass. The 18'-CH3, 18'-H, and 15-H resonances in the 1H NMR spectra of R-PC beta-2TP PEB and B-PE beta-2TP PEB appear significantly upfield from the corresponding thioether-linked ring A resonances seen in the spectrum of peptide alpha-1 PEB. The CD spectra of the two former peptides showed a strong positive Cotton effect at 300 nm. Such a Cotton effect is absent from the CD spectrum of peptide alpha-1 PEB and those of other A-ring-linked PEB peptides. Refluxing in methanol led to a near-quantitative release of PEB from alpha-1 PEB but no release from R-PC beta-2TP PEB and less than 20% release from B-PE beta-2TP PEB. In conjunction with earlier studies, these results show that distinctive amino acid sequences are found about the attachment sites for A-ring-linked, D-ring-linked, and dilinked (A- and D-ring-linked) bilins on the alpha and beta subunits of cyanobacterial and red algal phycobiliproteins and that the mode of linkage can be correctly predicted from inspection of the amino acid sequence.  相似文献   

15.
对龙须菜 (GracilarialemaneiformisGreville)及其色素突变体藻胆蛋白吸收光谱进行了比较研究 ,结果显示不同藻株藻红蛋白的吸收光谱有显著的变化 ,而藻蓝蛋白和别藻蓝蛋白的基本相同。我们克隆了龙须菜及其色素突变体的藻红蛋白亚基的部分基因序列 ,用该基因序列推导出的氨基酸序列进行分析以揭示这一变化的分子机理 ,结果显示除几个氨基酸残基的替换外 ,几株藻间的藻红蛋白的氨基酸序列十分相似 ,一些氨基酸的替换发生在决定藻红蛋白二级结构及亚基间相互作用的区域 ,可能会影响藻胆蛋白的构型及相互作用 ,导致光谱性质的变化。  相似文献   

16.
对龙须菜(Gracilaria lemaneiformis Greville)及其色素突变体藻胆蛋白吸收光谱进行了比较研究,结果显示不同藻株藻红蛋白的吸收光谱有显著的变化,而藻蓝蛋白和别藻蓝蛋白的基本相同.我们克隆了龙须菜及其色素突变体的藻红蛋白亚基的部分基因序列,用该基因序列推导出的氨基酸序列进行分析以揭示这一变化的分子机理,结果显示除几个氨基酸残基的替换外,几株藻间的藻红蛋白的氨基酸序列十分相似,一些氨基酸的替换发生在决定藻红蛋白二级结构及亚基间相互作用的区域,可能会影响藻胆蛋白的构型及相互作用,导致光谱性质的变化.  相似文献   

17.
Two isoproteins of the native B-phycoerythrin of the red alga, Rhodella violacea, were purified from crude extracts by preparative polyacrylamide gel electrophoresis and subsequently characterized. The slower moving pigment in gel electrophoresis was designated B-PE I, the faster as B-PE II. Both were found to occur in about equal amounts.B-PE I has a molecular weight of about 280000 and an IEP at 4.39, B-PE II a molecular weight of nearly 265000 and an IEP at 4.23. B-PE I and II are characterized by absorption maxima at 568 and 542 nm and a shoulder at 500 nm in the visible part of the absorption spectra. Their absorption coefficients at 542 nm differ with values of 5.54 and 5.63, respectively. The fluorescence emission spectra show a single maximum at 575 for B-PE I and at 578 nm for B-PE II. Both spectra have a shoulder at 630 nm. The fluorescence yield of B-PE II is lower by 25%.In calibrated SDS gel electrophoresis of the purified pigments B-PE I and II show two subunits of molecular weights of 18 900 and 29 200 and 18 500 and 29 900, respectively.Quantitative amino acid analyses indicated, that the isoproteins are very similar. B-PE II, however, has a significantly higher content of acidic amino acids and a lower percentage of basic residues, which is in keeping with its lower isoelectric point.Functional aspects of the occurrence of two isoproteins of B-phycoerythrin are discussed.Abbreviations PE phycoerythrin - MG molecular weight - IEP isoelectric point - SDS sodium dodecyl sulphate - AA acrylamide - BIS N,N-methylenebisacrylamide - Tris Tris(hydroxymethyl)aminomethan  相似文献   

18.
Summary Porphyridium cruentum extract was treated with rivanol for the precipitation and elimination of the polysaccharide typical for this alga, while all phycobiliproteins remained solubilized. After their precipitation with ammonium sulphate, B-phycoerythrin was differentially separated from the other phycobiliproteins, and rivanol was removed by Sephadex G-25 gel filtration. The purity of B-phycoerythrin was proved.Abbreviations B-PE B-phycoerythrin - b-PE b-phycoerythrin - R-PC R-phycocyanin - APC allophycocyanin - PBP phycobiliproteins  相似文献   

19.
The genes for the alpha and beta subunits of a novel six bilin-bearing (class II) phycoerythrin were cloned from Synechococcus sp. WH8020 and sequenced. The cloned genes (mpeA and mpeB) were detected by homology with the genes for C-phycoerythrin from Pseudanabaena sp. PCC7409. The mpe locus occurs once in the genome and is arranged similarly to that of many other phycobiliproteins, with mpeA shortly 3' of mpeB. Sequence comparison suggests that this phycoerythrin (and perhaps all class II phycoerythrins) occupy a branch of the phycoerythrin family separate from five-chromophore per alpha beta (class I) phycoerythrins, C-phycoerythrin, and B-phycoerythrin. The position of the sixth chromophore of the class II phycoerythrin of WH8020 was determined by comparison of the amino acid sequence of the chromopeptides (Ong, L. J., and Glazer, A. N. (1991) J. Biol Chem. 266, 9515-9527) with the sequence deduced from the gene. This located the chromophore at residue 75 of the alpha subunit, very close to the alpha-83 chromophore in the primary structure and, presumably, in the three-dimensional structure.  相似文献   

20.
Marine Synechococcus strains WH8103, WH8020, and WH7803 each possess two different phycoerythrins, PE(II) and PE(I), in a weight ratio of 2-4:1. PE(II) and PE(I) differ in amino acid sequence and in bilin composition and content. Studies with strain WH7803 indicated that both PE(II) and PE(I) were present in the same phycobilisome rod substructures and that energy absorbed by PE(II) was transferred to PE(I). Strain WH8103 and WH8020 PE(I)s carried five bilin chromophores thioether-linked to cysteine residues in sequences homologous to those previously characterized in C-, B-, and R-PEs. In contrast, six bilins were attached to strain WH8103 and WH8020 PE(II)s. Five of these were at positions homologous to bilin attachment sites in other phycoerythrins. The additional bilin attachment site was on the alpha subunit. The locations and bilin types in these PE(s) and in the marine Synechocystis strain WH8501 PE(I) (Swanson, R. V., Ong, L. J., Wilbanks, S. M., and Glazer, A. N. (1991) J. Biol. Chem. 266, 9528-9534) are: (table; see text) Since phycourobilin (PUB) (lambda max approximately 495 nm) transfers energy to phycoerythrobilin (PEB) (lambda max approximately 550 nm), inspection of these data shows that the invariant PEB group at beta-82 is the terminal energy acceptor in phycoerythrins. The adaptations to blue-green light, high PUB content and the presence of an additional bilin on the alpha subunit, increase the efficiency of light absorption by PE(II)s at approximately 500 nm.  相似文献   

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