一种海生超微型黄藻的色素蛋白复合体

从海生超微型黄藻品系PP983中分离得到一种水溶性色素蛋白复合物,其光学特性与普遍存在于甲藻叶绿体中的多甲藻素-叶绿素α-蛋白复合物（Peridinin-Chlorophyllα-Protein Complexes,PCP)类似,初步测得该色素蛋白复合体的分子量约为154,200,分子中多甲藻素,叶绿素α,蛋白的摩尔比为16：2：5。研究结果显示,PP983中的PCP是作为聚光色素复合体存在于藻细胞的叶绿体中的。     

一种分析叶绿体类囊体膜色素蛋白复合物的蓝绿温和胶电泳系统

采用蓝绿温和胶电泳系统可以非常有效地分离叶绿体蛋白质复合物,包括PSⅠ, PSⅡ, ATP合酶,细胞色素b₆f复合物,捕光色素复合物和1,5-二磷酸核酮糖羧化酶.还结合SDS-聚丙烯酰胺凝胶电泳将叶绿体多亚基复合物的50多种蛋白质分开,利用免疫印迹对蛋白质复合物进行了初步鉴定,同时还应用蓝色温和胶电泳分析基质、基粒类囊体复合物的组成.     

两种电泳系统分离裙带菜色素-蛋白复合物的比较

裙带菜的类囊体膜经过去污剂癸基-N-甲基匍萄糖胺增溶,采用SDS-PAGE分离技术,在Tris-Gly和Tris-硼酸两种电泳系统中分离其色素-蛋白质复合物,并比较其复合物的光谱特性。结果表明:采用Tris-Gly电泳分离系统从裙带菜中分离到8种色食-蛋白质复合物,分别是CP Ia、CPI、LHC₁、CPa、LHC₂、LHC₃、LHC₄和LHC₅。在Tris-硼酸电泳分离系统中共分离到5种色素-蛋白质复合物,分别是CPI、CPa、LHC₁、LHC₂、LHC₃。吸收光谱和荧光光谱的测定结果表明,两种电泳系统中分离的相对应条带的光谱特性基本相近。     

氮素水平对小麦幼苗叶绿体色素蛋白复合体含量的影响

在水培条件下,研究了不同氮素水平对小麦幼苗叶绿体色素、色素蛋白复合体含量及其光谱特征的影响。结果显示:(1)氮素水平较低时PSⅡ捕光色素蛋白复合体LHCⅡ在24~30 kD范围内的蛋白含量降低,不供氮时,色素蛋白复合体含量最低,而高分子量区域的蛋白组分相对较为稳定,说明氮素水平影响PSⅡ的多肽组分,而对PSⅠ多肽组分的影响相对较小。(2)室温吸收光谱分析表明,氮素水平较低时结合态色素的含量及比例发生改变,影响植物对光的吸收能力;荧光激发及发射光谱的峰值均随氮素浓度的升高而升高,说明增加施氮量时,叶绿体类囊体中受激发的色素分子数目增加,荧光强度也随之增大;叶绿体蛋白含量在16.86 mg.L-1氮素浓度时最大。     

蓝藻叶绿素蛋白复合体的分离研究

蓝藻类囊体膜用声波超时处理,然后在4℃下用低浓度的LDS增溶,并经改进的SDS-聚丙烯酰胺凝胶电泳后被分离成15条绿色的带. 其中CPa1～CPa6有着相似的吸收光谱. 这6个组分的低温荧光光谱也很相似,其荧光发射光谱的发射峰都位于685 nm处,表明它们都属于光系统Ⅱ叶绿素a蛋白复合体. 该系统对光系统Ⅱ的分离能力是传统电泳的3倍.     

一个高分辨率的凝胶电泳系统及其从蓝藻中分离的叶绿素蛋白复合体

用一高分辨率的凝胶电泳系统从蓝藻类囊体膜中分离出至少１３ 个清晰的叶绿素带,它们是ＣＰＩａ、ＣＰＩｂ、ＣＰＩｃ、ＣＰＩｄ、ＣＰＩｅ、ＣＰＩｆ、ＣＰＩｇ、ＣＰＩｈ、ＣＰａ１、ＣＰａ２、ＣＰａ３、ＣＰａ４ 和ＦＣ,其分辨率较传统方法高出１ 倍多。ＣＰＩａ—ＣＰＩｈ ８ 种组分有相同的吸收光谱,其红峰和蓝峰的位置分别位于６７６ ｎｍ 和４３６ ｎｍ 处。它们都属于光系统Ｉ叶绿素蛋白复合体。ＣＰａ１—ＣＰａ４ ４ 种组分的光谱性质亦基本相同,其吸收峰的位置分别位于６７０—６７２ ｎｍ 和４３６ ｎｍ 处,而低温荧光发射峰的位置都位于６８５ ｎｍ 处。它们都属于光系统Ⅱ叶绿素蛋白复合体     

茶叶片阶段性返白过程中色素蛋白复合体的变化

用ＳＤＳ－ＰＡＧＥ盘状电泳从温敏型阶段性返白的安吉白茶完全复绿叶中分离出２条叶绿素ａ蛋白复合物（ＣＰⅠ）、２条叶绿素ａ／ｂ捕光蛋白复合物（ＣＰⅡ：ＬＨＣＰ１、ＬＨＣＰ２）和１条游离色素带,而全白期叶片中则未发现ＣＰⅠ和ＣＰⅡ,但有很少量ＣＰⅡ的脱辅基蛋白和游离色素。此类叶片开始得绿时,ＣＰⅠ、ＬＨＣＰ１和ＬＨＣＰ２同时出现,但寡聚形式（ＣＰⅠａ）的恢复稍迟些,各种色素蛋白复合体的含量均随绿逐步上升     

固氮红细菌(Rhodobacter azotoforman)色素蛋白复合体的分离纯化与特性

【目的】为揭示不产氧光合细菌产氢菌株色素蛋白复合体(PPC)色素组成和含量与光合放氢的关系奠定基础。【方法】以PPC特征光谱为检测指标,采用硫酸铵分级分离、DEAE-纤维素层析、吸收光谱和SDS-PAGE等方法进行了固氮红细菌(Rhodobacter azotoformans,R.azotoformans)R7产氢菌株PPC的分离纯化、纯度分析和鉴定;采用表面增强激光解吸电离离子飞行时间质谱、HPLC-MS和荧光光谱法对其中一种PPC进行了组成分析和能量传递活性测定。【结果】从R7菌株获得了3种纯化的PPC,1种为反应中心与中心捕光色素蛋白复合体(RC-LH1),2种为外周捕光色素蛋白复合体(LH2),其中一种LH2的吸收光谱具有异常的423nm强吸收峰,其蛋白的两种亚基的分子量分别为5356.8Da和5697.8Da,类胡萝卜素属球形烯系,分子量为562Da,激发光能够从类胡萝卜素向细菌叶绿素以及细菌叶绿素向细菌叶绿素传递。【结论】固氮红细菌产氢菌株含有2种不同光谱特性的LH2,其中一种具有新光谱特性。     

蛋白质复合体非变性凝胶电泳技术及其应用新进展

非变性凝胶电泳技术是研究蛋白质复合体的强有力工具。重点介绍了蓝绿温和非变性凝胶电泳(BN-PAGE)技术的原理和特点,比较了由BN-PAGE衍生的蓝色温和琼脂糖凝胶电泳、清澈温和非变性凝胶电泳(CN-PAGE)和高分辨清澈温和非变性凝胶电泳(HrCN-PAGE)技术的差异和适用范围,并概括地介绍了这些技术在植物蛋白质复合体研究中应用的新进展。     

一个适用于分离光系统Ⅱ的凝胶电泳系统

用一高分辨率的凝胶电泳系统从延长破碎时间的蓝藻类囊体膜增溶物中分离出１４条绿色的带。按照电泳迁移率的增加顺序,自上而下分别是ＣＰＩａ,ＣＰＩｂ,ＣＰＩｃ,ＣＰＩｄ,ＣＰＩｅ,ＣＰＩｆ,ＣＰＩｇ,ＣＰＩｈ,ＣＰａ１,ＣＰａ２,ＣＰａ３,ＣＰａ４,ＣＰａ５和ＦＣ。ＣＰａ１,ＣＰａ２,ＣＰａ３,ＣＰａ４和ＣＰａ５５种叶绿素蛋白复合体的吸收光谱相似,它们在蓝区的吸收峰位子４３６ｎｍ,而红区的吸收峰则位于６７０—６７３ｎｍ附近。它们的低温荧光发射光谱亦很相似,其荧光发射峰都位于６８５ｎｍ处。因此它们都属于光系统Ⅱ叶绿素ａ蛋白复合体．跟传统电泳相比,该系统对光系统Ⅱ的分离能力提高了１．５倍。     

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.     

一种丙烯酰胺凝胶的新型光聚合法

以亚甲基蓝为引发剂,甲苯亚磺酸钠为还原剂,二苯氯化碘为氧化剂的丙烯酰胺凝胶光聚合方法,具有试剂配制简单, 适用范围广,方法稳定性和重现性好,且聚合速度快,操作易于控制等特点, 是一种不同于最为广泛使用的过硫酸铵/四甲基乙二胺和核黄素/四甲基乙二胺法的新型光聚合法. 用这一方法做了SDS和在酸性缓冲体系中的尿素变性梯度胶电泳,初步试用表明这一新型光聚合法具有很好的聚合效果.     

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.     

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、     

突变体大麦Mb1832C的叶绿素蛋白复合体

用低浓度ＳＤＳ对突变体大麦Ｍｂ１８３２Ｃ的类囊体膜进行增溶,再经不连续的ＳＤＳ－聚丙烯酰胺凝胺电泳后,按电泳迁移率的增加顺序,分离出ＣＰＩ,ＣＰａｌ,ＣＰａ２和ＦＣ４条蓝绿色的带。ＣＰＩ在红区和蓝区的吸收峰分别位于６６７ｎｍ和４３８ｎｍ处。低温荧光发射光谱表明ＣＰＩ为含有少量光系统Ⅱ的光系统Ⅰ反应中心复合体。     

Chlorophyll-protein complex composition and photochemical activity in developing chloroplasts from greening barley seedlings

The time course for the observation of intact chlorophyll-protein (CP) complexes during barley chloroplast development was measured by mild sodium dodecyl sulfate polyacrylamide gel electrophoresis. The procedure required extraction of thylakoid membranes with sodium bromide to remove extrinsic proteins. During the early stages of greening, the proteins extracted with sodium bromide included polypeptides from the cell nucleus that associate with developing thylakoid membranes during isolation and interfere with the separation of CP complexes by electrophoresis. Photosystem I CP complexes were observed before the photosystem II and light-harvesting CP complexes during the initial stages of barley chloroplast development. Photosystem I activity was observed before the photosystem I CP complex was detected whereas photosystem II activity coincided with the appearance of the CP complex associated with photosystem II. Throughout chloroplast development, the percentage of the total chlorophyll associated with photosystem I remained constant whereas the amount of chlorophyll associated with photosystem II and the light-harvesting complex increased. The CP composition of thylakoid membranes from the early stages of greening was difficult to quantitate because a large amount of chlorophyll was released from the CP complexes during detergent extraction. As chloroplast development proceeded, a decrease was observed in the amount of chlorophyll released from the CP complexes by detergent action. The decrease suggested that the CP complexes were stabilized during the later stages of development.Abbreviations Chl chlorophyll - CP chlorophyll-protein - CPI P700 chlorophyll-a protein complex of photosystem I - CPa electrophoretic band that contains the photosystem II reaction center complexes and a variable amount of the photosystem I light-harvesting complex - CP A/B the major light-harvesting complex associated with photosystem II - DCIP 2,6-dichlorophenolindophenol - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DPC diphenyl carbazide - MV methyl viologen - PAR photosynthetically active radiation - PSI photosystem I - PSII photosystem II - SDS sodium dodecyl sulfate - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis - TEMED N,N,N,N-tetramethylethylenediamine - TMPD N,N,N,N-tetramethyl-p-phenylenediamine Cooperative investigations of the United States Department of Agriculture, Agricultural Research Service, and the North Carolina Agricultural Research Service, Raleigh, NC 27695-7601. Paper No. 9949 of the Journal Series of the North Carolina Agricultural Research Service, Raleight, NC 27695-7601.     

Agarose Gel Electrophoresis for the Separation of DNA Fragments

Agarose gel electrophoresis is the most effective way of separating DNA fragments of varying sizes ranging from 100 bp to 25 kb¹. Agarose is isolated from the seaweed genera Gelidium and Gracilaria, and consists of repeated agarobiose (L- and D-galactose) subunits². During gelation, agarose polymers associate non-covalently and form a network of bundles whose pore sizes determine a gel''s molecular sieving properties. The use of agarose gel electrophoresis revolutionized the separation of DNA. Prior to the adoption of agarose gels, DNA was primarily separated using sucrose density gradient centrifugation, which only provided an approximation of size. To separate DNA using agarose gel electrophoresis, the DNA is loaded into pre-cast wells in the gel and a current applied. The phosphate backbone of the DNA (and RNA) molecule is negatively charged, therefore when placed in an electric field, DNA fragments will migrate to the positively charged anode. Because DNA has a uniform mass/charge ratio, DNA molecules are separated by size within an agarose gel in a pattern such that the distance traveled is inversely proportional to the log of its molecular weight³. The leading model for DNA movement through an agarose gel is "biased reptation", whereby the leading edge moves forward and pulls the rest of the molecule along⁴. The rate of migration of a DNA molecule through a gel is determined by the following: 1) size of DNA molecule; 2) agarose concentration; 3) DNA conformation⁵; 4) voltage applied, 5) presence of ethidium bromide, 6) type of agarose and 7) electrophoresis buffer. After separation, the DNA molecules can be visualized under uv light after staining with an appropriate dye. By following this protocol, students should be able to: 1. Understand the mechanism by which DNA fragments are separated within a gel matrix 2. Understand how conformation of the DNA molecule will determine its mobility through a gel matrix 3. Identify an agarose solution of appropriate concentration for their needs 4. Prepare an agarose gel for electrophoresis of DNA samples 5. Set up the gel electrophoresis apparatus and power supply 6. Select an appropriate voltage for the separation of DNA fragments 7. Understand the mechanism by which ethidium bromide allows for the visualization of DNA bands 8. Determine the sizes of separated DNA fragments       

Effects of Aluminium on Pigments and Pigment-Protein Complexes of Soybean

The effect of aluminium (0.5 –1.0 mM) on contents of phosphorus, pigments, and pigment-protein complexes was studied in soybean (Glycine max Merril.) grown in different nutrient medium with and without P. Increased Al concentrations led to the decrease in the contents of chlorophylls (Chl) a and b, and carotenoids (Car) in soybean leaves, but Chl a/b ratio did not vary significantly. In long-term experiments, P ameliorates the negative effects of Al.