Pigment quantitation and analysis by HPLC reverse phase chromatography: a characterization of antenna size in oxygen-evolving photosystem II preparations from cyanobacteria and plants.
Pigment quantitation and analysis by HPLC reverse phase chromatography: a characterization of antenna size in oxygen-evolving photosystem II preparations from cyanobacteria and plants.
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DOI：
10.1021/bi960056z
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发表时间：
1996-06
期刊：
影响因子：
2.9
通讯作者：
J. S. Patzlaff;B. Barry
中科院分区：
文献类型：
--
作者：
J. S. Patzlaff;B. Barry
光系统II，光合水氧化复合物，可以从植物和蓝藻中分离出来。已经开发了多种方法来纯化这种酶，它可以从几种功能和结构形式中分离出来。光系统II制剂的色素含量的知识是重要的精确光谱，生化和功能分析。我们已经确定了色素化学计量在植物和蓝藻进化的光系统II制剂。我们采用溶剂系统进行反相高效液相色谱柱的等密度洗脱，测定了相关颜料的消光系数。从四种光系统II制剂中提取色素。这些制备包括菠菜光系统II膜[Berthold， D. A., Babcock， G. T., & Yocum， C. F. (1981) FEBS Lett. 134, 231-234]，菠菜光系统II反应中心配合物[Ghanotakis， D. F., & Yocum， C. F. (1986) FEBS Lett. 197, 244-248]，菠菜光系统II配合物[MacDonald， G. M., & Barry， B. A. (1992) Biochemistry 31,9848 -9856]，以及从蓝藻，Synechocystis sp. PCC 6803中分离的光系统II颗粒[Noren， G. H., Boerner， R. J., & Barry，B. A.（1991）生物化学，30(3):344 - 349。色素化学计量学采用两种不同的数据分析方法确定，并基于每个光系统II反应中心有两个叶绿素a分子的假设。所获得的色素化学计量学与两种数据分析方法具有可比性，并且与先前的制剂的生物物理和生化表征一致。两种数据分析方法测定的平均色素化学计量量（叶绿素：质体醌-9 / 2叶绿素a）如下：光系统II膜，274:3.2；光系统II反应中心配合物，78:2.5；聚囊藻PS II颗粒，55:2.4；光系统II配合物，121:2.0。
Photosystem II, the photosynthetic water-oxidizing complex, can be isolated from both plants and cyanobacteria. A variety of methods have been developed for purification of this enzyme, which can be isolated in several functional and structural forms. Knowledge of the pigment content of photosystem II preparations is important for precise spectroscopic, biochemical, and functional analysis. We have determined pigment stoichiometries in oxygen-evolving photosystem II preparations from plants and cyanobacteria. We have employed a solvent system for the isocratic elution of a reverse phase HPLC column in which we have determined the extinction coefficients of the relevant pigments. Pigments were extracted from four photosystem II preparations. These preparations included spinach photosystem II membranes [Berthold, D. A., Babcock, G. T., & Yocum, C. F. (1981) FEBS Lett. 134, 231-234], spinach photosystem II reaction center complexes [Ghanotakis, D. F., & Yocum, C. F. (1986) FEBS Lett. 197, 244-248], spinach photosystem II complexes [MacDonald, G. M., & Barry, B. A. (1992) Biochemistry 31, 9848-9856], and photosystem II particles isolated from the cyanobacterium, Synechocystis sp. PCC 6803 [Noren, G. H., Boerner, R. J., & Barry, B. A. (1991) Biochemistry 30, 3943-3950]. Pigment stoichiometries were determined using two different methods of data analysis and were based on the assumption that there are two pheophytin a molecules per photosystem II reaction center. The pigment stoichiometries obtained were comparable for the two methods of data analysis and agreed with previous biophysical and biochemical characterizations of the preparations. The average pigment stoichiometries (chlorophyll:plastoquinone-9 per 2 pheophytin a) determined using the two data analysis methods were as follows: photosystem II membranes, 274:3.2; photosystem II reaction center complexes, 78:2.5; Synechocystis PS II particles, 55:2.4; photosystem II complexes, 121:2.0.