褐藻裙带菜色素蛋白复合物的性质*

用去污剂DMG增溶褐藻裙带菜(Undaria pinnatifida)的类囊体膜,通过PAGE分离色素-蛋白复合物并分析其性质,结果表明:CPⅠa和CPⅠ都含有66kDa的多肽,低温荧光发射光谱中有715nm的长波荧光峰,激发光谱测定结果表明CPⅠa是含有墨角藻黄素的叶绿素a/c-蛋白复合物,CPⅠ是只含有叶绿素a的色素-蛋白复合物。CPa含有51、37、34和20kDa四种多肽,低温荧光发射峰位于683nm,激发光谱表明它含有叶绿素a、c和少量墨角藻黄素,是裙带菜的PSⅠ复合物。其余5条为捕光色素-蛋白复合物,它们都是由20kDa的多肽组成,其中LHC₁和LHC₃有相似的光谱特性,是墨角藻黄素-叶绿素a/c-蛋白复合物,LHC₂、LHC₄和LHC₅的光谱特性相似,是叶绿素a/c-蛋白复合物。     

绿藻假根羽藻色素-蛋白复合物的分离及其特性研究

采用聚丙烯酰胺凝胶电泳(PAGE)和蔗糖密度梯度超速离心方法分离了假根羽藻(Bryopsis corticulans)的色素-蛋白复合物,并对其特性进行分析。结果表明:采用PAGE分离得到7条色素-蛋白复合物带,分别是CPⅠa1、CPⅠa2、CPⅠ、LHCP₁、LHCP₂、CPa、LHCP₃₊₃,和2条游离色素(free pigment,FP)FCa、FC。用改进的不连续蔗糖密度梯度离心法分离到五条带。区带Ⅰ是FP;区带Ⅱ主要是小分子量的PSⅡ捕光复合物LHCP₃₊₃;区带Ⅲ以PSⅡ捕光复合物的聚集体LHCP1为主,区带Ⅱ和Ⅲ的吸收光谱中除了Chla外,还含有大量的Chlb和管藻黄素,是管藻黄素-Chla/b-蛋白质复合物;区带Ⅳ在PAGE中只显示一条带,光谱中有Chlb吸收肩峰,含有66和56kDa两种多肽,是较小的PSⅠ复合物CPⅠa。     

三种不同制剂来源的叶绿素蛋白质复合物CPa的光谱特性及其组分的比较研究

用 SDS-PAGE 方法分离了菠菜叶绿体制剂、放氧光系统Ⅱ制剂和放氧光系统Ⅱ反应中心核心复合物的色素蛋白质复合物,对它们的 CPa 带进行的光谱特性的对比研究表明,在前两种制剂中 CPa 带不仅含有 Chl a 的蛋白质复合物带,它还含有少量 Chl b。且叶绿体制剂的 CPa 带中的 Chl b 含量高于放氧光系统Ⅱ制剂中的含量。此外,根据光系统Ⅱ反应中心核心复合物只有一条叶绿素蛋白质复合物带(CPa)的实验结果,我们认为光系统Ⅱ反应中心叶绿素蛋白质复合物即在 CPa 带中。但在叶绿体制剂和放氧光系统Ⅱ制剂的情况下,CPa 带还含有其它组分。     

低氮对金色奥杜藻色素-蛋白复合物特性及全细胞蛋白表达的影响

采用SDS-PAGE圆盘电泳分离技术, 研究两种氮浓度下17.6 mmol•L1 (高氮组)、5.87 mmol•L1 (低氮组)金色奥杜藻色素蛋白复合物的变化规律。结果表明, 从两种氮浓度所培养的金色奥杜藻细胞光合膜上, 均可分离得到6种色素蛋白复合物条带, 按照迁移速率从小到大依次为CPⅠa、CPⅠ、CPa1、CPa2、LHCP1、LHCP2。与高氮组相比, 低氮下电泳分离的复合物条带颜色明显变浅, 尤其CPⅠ、CPa1、CPa2三条带仅隐约可见。低温荧光发射光谱显示CPa1的荧光发射主峰也较高氮组蓝移10 nm左右,说明低氮主要影响光系统Ⅱ。双向电泳和质谱分析结果表明, 氮浓度降低后藻细胞中有6种蛋白表达下调, 3种蛋白表达上调, 这些差异蛋主要白涉及光合作用、氮素吸收、碳同化、能量转换等生理过程, 大多与维持叶绿体功能有关。     

小分子肽的Tricine-SDS-PAGE分离方法

在蛋白质的生化分析和基因表达产物的分离纯化中 ,经常要把某种或某些蛋白质成分分离开来。聚丙烯酰胺凝胶电泳 ( SDS- PAGE)是分离蛋白质的常用生化方法 ,样品的蛋白质分子热变性解聚后与 SDS结合形成带负电的蛋白质 - SDS复合物 ,复合物在电泳中的迁移率取决于蛋白质的分子大小 ,使用均匀浓度的 SDS-PAGE来分析分子量在 15～ 2 0 0 k D的蛋白质时 ,电泳迁移率与分子量的对数成线性关系。但常规定 Tris-甘氨酸 -盐酸系统中电泳分离分子量小于 10 k D的多肽效果差。作者在分离小分子肽的实验中改进了一套较简便的分离小分子肽的 T…     

褐藻裙带菜色素—蛋白质复合物的分离与命名

以非离子去污剂癸基－Ｎ－甲基葡萄糖胺为增溶剂,采用聚丙烯酰胺凝胶电泳技术从褐藻裙带菜（Ｕｎｄａｒｉａ ｐｉｎｎａｔｉｆｉｄａ Ｈａｒｖ．）的类囊体膜上分离到８种色素－蛋白质复合物。根据其表观分子量、光谱特性和多肽分析结果,并以高等植物菠菜（Ｓｐｉｎａｃｉａ ｏｌｅｒａｃｅａ Ｌ．）为对照,按照Ａｎｄｅｒｓｏｎ命名系统,８种色素－蛋白质复合物分别命名为ＣＰⅠ ａ、ＣＰⅠ、ＣＰａ、ＬＨＣ１、ＬＨＣ２、     

叶绿体膜的结构与功能 Ⅶ小麦囊状体膜上光系统Ⅱ反应中心复合体的探讨

小麦叶绿体膜用SDS短时间增溶后,用不连续的SDS—聚丙烯酰胺凝胶电泳分离出八条叶绿素带,我们依其迁移率的增加及参考文献上的定名称为CPI(P700—叶绿素a—蛋白质)、LHCP~1(捕光叶绿素a/b—蛋白质)、LHCP~2、LHCP~3,CPa(光系统Ⅱ反应中心)、LHCP~4和FC(游离色素—SDS复合物)。值得注意的是,在LHCP~4和FC之间观察到一条新的复合体,我们命名为CPa_1。 CPa_1的吸收光谱与CPa的吸收光谱相似,他们在红区的吸收峰分别在669nm和670nm,在蓝区的吸收峰为435nm,清楚地表明这些吸收光谱与文献中报导的复合体Ⅳ—系统Ⅱ反应中心复合物相似(Hayden等1977)。CPa和CPa_1具有相似的荧光发射光谱,最强的发射带分别在681nm和682nm。二者的荧光激发光谱亦是彼此相似的。CPa的分子量约为39.5KD,CPa_1的分子量约为19.6KD。因此,我们推测CPa可能是二聚体,而CPa_1可能是它的单体。     

从超声波破碎的蓝藻类囊体膜中分离的叶绿素蛋白复合物

当蓝藻的类囊体膜用超声波进行破碎,并在4℃下用聚丙烯酰胺凝胶电泳进行分离,有6条叶绿素带被分离出来,它们分别是 CPIa,CPIb,CP1,CPa1 CPa2,FC。CP1 在红区和蓝区的吸收峰分别位于674和435 nm 处。在液氮甲该组分在725和680 nm 处有两个荧光发射带。CPa1和 CPa2的吸收光谱相似,其红峰和蓝峰的位置分别位于667和431.5nm 处。它们在77 K 的荧光发射峰都位于684 nm 处。用超声破碎法分离的叶绿素蛋白复合物的光谱特性,除 CPa1和 CPa2在红峰和蓝峰的吸收位置蓝移了3—5 nm 之外,其余与用 SDS 增溶法分离的相应复合物相似。属于光系统Ⅰ的 CPIa-CPI 的叶绿素含量占总叶绿素的40.93%,而属于光系统Ⅱ的 CPa1和 CPa2的叶绿素则占总叶绿素的38.78%,二者之差仅有2.15%。     

柱孢鱼腥藻(Anabaena cylindkica)营养细胞和异形胞叶绿素蛋白复合体的比较研究

比较了柱孢鱼腥藻(Anabaena cylindrica)营养细胞和异形胞类囊体膜叶绿素蛋白复合体的种类和性质。以SDS增溶营养细胞类囊体膜和不连续聚丙烯酰胺电泳分离得到4个P700叶绿素a蛋白复合体,分别为GPIa、CPIb、CPIc和CPI;和1个系统Ⅱ叶绿素蛋白复合体CPa。相对迁移率小的4个复合体含有P700,呼收光谱红区吸收峰为675nm,液氮低温荧光发射光谱有728nm荧光发射峰。CPIa和CPI的分量子分别为205 和105千道尔顿。未见诸文献的CPIb和CPIc复合体的分子量介于CPIa和CPI之间。相对迁移率较大的CPa有着吸收光谱红区672nm吸收峰,液氮低温荧光发射光谱有687nm荧光发射峰,分子量为56千道尔顿。同时化学氧化还原差示光谱不表现P700吸收降低。柱孢鱼腥藻异形胞类囊体膜经SDS增溶和电泳分离得到2个系统Ⅰ叶绿素蛋白复合体,它们的吸收光谱特性和分子量大小相近于营养细胞分离的CPIa和CPI复合体。异形胞类囊体膜缺少系统Ⅱ叶绿索蛋白复合体。     

裙带菜PSⅠ复合物的荧光特异性

以褐藻裙带菜（Undaria pinnatifida）为实验材料,采用蔗糖密度梯度超速离心的方法,去污剂SDS为增溶剂（SDS:Chl＝20:1,4℃增溶20 min）,蔗糖密度梯度为60%、50%、40%、30%、20%、15%和10%,分离制备光系统Ⅰ(PSⅠ)复合物。结果表明, 40% 蔗糖层带所含色素蛋白复合物是PSⅠ复合物。利用红藻作参照对比,光谱结果表明从裙带菜中得到的PSⅠ复合物没有730 nm的荧光峰。分析认为这是所有褐藻包括裙带菜PSⅠ复合物的荧光特异性。     

Isolation and Nomenclature of Pigment-Protein Complexes from the Brown Alga (Undaria pinnatifida Harv.)

Eight kinds of pigment-protein complexes were resolved from the thylakoid membrane of the brown alga （Undaria pinnatifida Harv.) by using non-ionic detergent decanoyl-N-methylglucamide and PAGE technique. According to the apparent molecular weights, spectra characteristics, polypeptide compositions and referring to the higher plant spinach, eight pigment-protein complexes were named under Anderson′s terminology system as CPⅠa, CPⅠ, CPa, LHC1, LHC2, LHC3, LHC4, LHC5.     

The Chlorophyll-Protein Complexes Resolved from Ultrasonically Broken Thylakoid Memb-ranes of Blue-Green Algae

When the thylakoid membranes of blue-green algae were broken by ultrasonic vibrations and subjected to polyacrylamide gel electrophoresis at 4℃, six green zones were resolved. They were designated as CPIa, CPlb, CPI; CPal, CPa2, and FC. The absorption spectrum of CPI had a red maximum at 674 nm and a peak in the blue at 435 nm. It was identified as PS chlorophyll a-protein Complex, but was contaminated with minor PSⅡ which was implied by the appearance of fluorescence emission peak at 680 nm besides the main one at 725 nm at 77 K. The spectral properties of CPIa and CPlb were similar to that of CPl. The absorption spectra of CPa1 and CPa2 were similar, both having red maxima at 667 nm and peaks in the blue at 431.5 nm. Their fluorescence emission had the same peaks at 684 nm at 77 K indicating that they belonged to PSⅡ. It was recognized that CPal of 47 kD is the reaction center complex of photosystem Ⅱ and CPa2 of 40 kD is the internal antenna complex of photosystem Ⅱ. The spectral characteristics of the chlorophyll-protein complexes resolved by ultrasonic method were similar to those of the same complexes resolved by SDS solubilization, except the absorbance positions of CPa1 and CPa2 in the blue peak and the red one which shifted to blue about 3–5 nm. It was calculated that in thylakoid membranes of blue-green algae 40.93% chlorophyll was in PSⅠ, while 38.78% of chlorophyll in PSⅡ. The difference of chlorophyll contents between PSⅠ and PSⅡ was only 2.15%. Concerning the fact that minor PSⅡ compound remained in the part of PSⅠ zones, it might be concluded that the distribution of chlorophyll between PSⅠ and PSⅡ in blue-green algae was equal. This result was in agreement with the hypothesis that PSⅠ and PSⅡ operates in series in photosynthetic electron transport.     

Chlorophyll-Protein Complexes of Blue-Green Algae Isolated by a New Gel Electrophoresis System with High Resolving Power

At least 13 chlorophyll bands from the thylakoid membranes of blue-green algae could be clearly resolved by SDS-PAGE employing a new improved procedure. They were designated as CPIa, CPIb, CPIc, CPId, CPIe, CPIf, CPIg, CHIh, CPal, CPa2, CPa3, CPa4 and FC. 8 chlorophyll-protein complexes, CPIa-CPIh, had the same absorption spectrum at 676 nm in the red and 436 nm in the blue region. They belonged to the chlorophyll-protein complexes of PS Ⅰ. 4 chlorophyll-protein complexes, CPal-CPa4, had a red absorption peak at 670­672 nm and a blue one at 436 nm. Their fluorescence emission peak at 77K was at 685 nm. They were chlorophyll-protein complexes of PS Ⅱ.     

Functional and structural organization of chlorophyll in the developing photosynthetic membranes of Euglena gracilis Z. V-separation and characterization of pigment-protein complexes of the differentiated thylakoids

Low temperature sodium dodecyl sulfate polyacrylamide gel electrophoresis following mild solubilization of Euglena thylakoid components allowed to resolve, in addition to the main CP1, CPa and LHCP chlorophyll-protein complexes, the additional CP1a and LHCP green bands. A carotenoid enriched band CPc can be separated from CPa using high acrylamide concentration. Pigment and polypeptide composition of these complexes were analyzed by absorption and fluorescence measurements and two dimensional gel electrophoresis. Spectral properties of CP1 and CP1a indicate an heterogenous organization of chlorophyll and the presence of significant amount of chlorophyll b in these complexes. They both contain a major 68 kilodalton polypeptide associated with three minor low molecular weight polypeptides in CP1a. CPa and CPc exhibit a characteristic fluorescence emission at 687 nm and they each contain one polypeptide of 54 and 41 Kda respectively. LHCP and LHCP are less abundant than in higher plant thylakoids and they contain a lower proportion of chl b (chl a: chl b=3). They include two polypeptides of 26 and 29 Kda.Abbreviations chl chlorophyll - SDS Sodium Dodecyl Sulfate - EDTA Ethylene Diamine Tetraacetic Acid - DTT Dithiothreitol     

Life time deficiency of photosynthetic pigment-protein complexes CP1, A1, AB1, and AB2 in two cecidomyiid galls derived from <Emphasis Type="Italic">Machilus thunbergii</Emphasis> leaves

Two kinds of cecidomyiid galls induced by Daphnephila on Machilus thunbergii Sieb. & Zucc. leaves at various developmental stages, i.e., young, growing, and mature, were analyzed for their biochemical composition of photosynthetic pigment-protein complexes located in thylakoid membranes using the Thornber and MARS electrophoretic fractionation systems. Both kinds of galls were totally deficient in the pigment-protein complexes CP1, and A1, AB1, and AB2 through the whole period of gall formation. Immunoblotting of antibody against light-harvesting complex 2b (LHC2b) apoprotein confirmed this deficiency in gall’s lifetime, which never recovered under any condition. Electron microscopy demonstrated that already at the early developmental stage the gall chloroplasts had thylakoid morphology like that in a normal leaf.     

The action of lipases on chloroplast membranes. III. The effect of lipid hydrolysis on chlorophyll-protein complexes in thylakoid membranes

Bean thylakoid membranes treated with various lipolytic enzymes (bean galactolipase, phospholipases A₂, C, D) showed marked changes in their acyl lipid composition. As a consequence of acyl lipids hydrolysis, destruction of some chlorophyll a-protein complexes (CP1a, CP1, CPa) or monomerization of the oligomeric of light harvesting chlorophyll a/b protein complex (LHCP) was observed. It is concluded that galactolipids and phosphatidylcholine are responsible for the stability of CP1a, CP1 and CPa, respectively. Phosphatidylglycerol and to some extent monogalactosyldiacylglycerol are essential for the stabilization of oligomeric structures of light harvesting chlorophyll a/b protein complex.Abbreviations chl chlorophyll - CP1a, CP1 chl a-protein complexes, of PSI - CPa chl a-protein complex of PSII - DGDG diagalactosyldiacylglycerol - FC free chl - GL galactolipase - LHCP^1–3 light harvesting chl a/b protein complex - MGDG monogalactosyldiacylglycerol - PAGE polyacrylamide gel electrophoresis - PC phosphatidylcholine - PG phosphatidylglycerol - PLA₂ phospholipase A₂ - PL phospholipase C - PLD phospholipase D - PSI photosystem I - PSII photosystem II - SDS sodium dodecyl sulphate - SQDG sulfoquinovosyl-diacylglycerol - TCA trichloroacetic acid - Tricine N-tris-(hydroxymethyl)-methylglycine - Tris Tris-(hydroxymethyl)-aminomethan     

Reinvestigation of the chlorophyll distribution among the chlorophyll-proteins and chlorophyll-protein complexes of Hordeum vulgare L.

Solubilization of barley (Hordeum vulgare L.) thylakoid membranes with sodium dodecylsulphate plus sodium deoxycholate with or without Triton X-100 and subsequent fractionation in the polyacrylamide gel electrophoresis system described in this paper resulted: (1) in the resolution of the chlorophyll-proteins and chlorophyll-protein complexes commonly known as CP1a, CP1, LHCP¹, LHCP², CPa and LHCP³; (2) in the highly increased stability of CP1 and CP1a, as judged by their chlorophyll content, (3) at the expense of the free pigment concentration (4) which could be reduced to a negligible amount. Some 40% of the total chlorophyll contained in the mature higher plant thylakoid membrane is associated with CP1 and CP1 a and as already suggested before [19] no significant amount of free chlorophyll occurs in vivo.Abbreviations chl chlorophyll - CP1 P₇₀₀-chla-protein - CPa P680-chla-protein - DOC sodium deoxychlolate - FC free chlorophyll - LHCP light-harvesting chlorophyll a/b-protein - PAGE(S) polyacrylamide gel electrophoresis (system) - SDS sodium dodecylsulphate - TX-100 Triton X-100     

The P-700-chlorophyl a-protein complex and two major light-harvesting complexes of Acrocarpia paniculata and other brown seaweeds

Acrocarpia paniculata thylakoids were fragmented with Triton X-100 and the pigment-protein complexes so released were isolated by sucrose density gradient centrifugation. Three main chlorophyll-carotenoid-protein complexes with distinct pigment compositions were isolated.

1. (1) A P-700-chlorophyll a-protein complex, with a ratio of 1 P-700: 38 chlorophyll a: 4 ta-carotene molecules, had similar absorption and fluorescence characteristics to the chlorophyll-protein complex 1 isolated with Triton X-100 from higher plants, green algae and Ecklonia radiata.

2. (2) An orange-brown complex had a chlorophyll a : c₂ : fucoxanthin molar ratio of 2 : 1 : 2. This complex had no chlorophyll c₁ and contained most of the fucoxanthin present in the chloroplasts. This pigment complex is postulated to be the main light-harvesting complex of brown seaweeds.

3. (3) A green complex had a chlorophyll a : c₁ : c₂ : violaxanthin molar ratio of 8 : 1 : 1 : 1. This also is a light-harvesting complex.

The absorption and fluorescence spectral characteristics and other physical properties were consistent with the pigments of these three major complexes being bound to protein. Differential extraction of brown algal thylakoids with Triton X-100 showed that a chlorophyll c₂-fucoxanthin-protein complex was a minor pigment complex of these thylakoids.