假根羽藻Cytb_6 f蛋白复合体的分离与纯化  

ISOLATION AND PURIFICATION OF CYT b_6 fCOMPLEX FROM BRYOPSIS CORTICULANS

作　　者：李宾兴[1] 毛大璋[1] 高振泮[2] 李良璧[1] 匡廷云[1] 

机构地区：[1]中国科学院植物研究所光合作用与环境分子生理学重点实验室,北京100093 [2]中国科学院海洋研究所实验海洋生物学重点实验室,青岛266071

出　　处：《海洋与湖沼》2007年第1期69-76,共8页Oceanologia Et Limnologia Sinica

基　　金：国家自然科学基金资助项目;39890390号和30370347号

摘　　要：提要改进了以往分离纯化Cytb6f的方法,进行了海洋绿藻——假根羽藻(Bryopsiscorticu-lans)Cytb6f蛋白复合体分离纯的研究。结果表明,该Cytb6f制剂由Cytf、Cytb6、Rieske[Fe-s]蛋白和亚基IV四种主要的多肽亚基以及少量的Chl·a和类胡萝卜素组成。4个多肽亚基的表观分子量分别为34·8、24·0、18·7及16·7kD,该Cytb6f制剂的Cytb6/f比值接近2·0,其纯度值为9·9nmolCytf/mg,其催化电子传递的活性(C10-PQH2→PC)为73e/s。上述分析表明,假根羽藻Cytb6f在组成、纯度和催化活性方面均与已报道的高等植物、淡水绿藻和光合细菌的Cytb6f制剂相似。As a plastoquinol-plastocyanin oxidoreductase, cytochrome b6f complex plays an important role in electron transfer and energy transduction in photosynthesis. It mediates the linear electron flow between PSII and PSI, catalyzes the cyclic electron flow around PSI, and sets up a transmembrane proton electrochemical potential to support energy to form ATP. In addition, cytochrome b6f complex is also involved in the regulation of balanced light excitation energy distribution between photosystems since its redox states governs the activation of LHCII kinase. It consists of four major subunits（ Cyt f, Cyt b, Rieske iron-sulfur protein, and subunit IV） and four small subunits（pet G, L, M, and N）. Besides, about one chlorophyll a molecule and one or less than one carotenoid molecule have been found in each cytochrome b6f monomer. The cytochrome b6f complex has been purified from higher plants, freshwater green algae and cyanobacteria. However, no case has been reported of this complex from marine green alga. In this work, the cytochrome b6f complex was purified from a marine green alga Bryopsis corticulans successfully by an improved method, its composition and activity were studied in this paper. According to the purification of spinach b6f complex, Cyt b6f was not isolated from Bryopsis corticulans thylakoid. It was shown that most of the b6f complex has gone before last ammonium sulfate fractionations. To solve this problem, ammonium sulfate concentration was adjusted. Although this modification was proved to be useful, some 40-60kD extrinsic proteins still existed. To effectively remove these extrinsic proteins, 2mol/L NaBr membrane washing was repeated after that the extrinsic proteins were removed successfully shown by SDS-PAGE. This modified purification procedures for Bryopsis corticulans b6f complex are：Chloroplasts were isolated from 2kg pre-chilled fresh Bryopsis corticulans as regular way. After osmotically broken, the chloroplasts were resuspended in 2mol/L NaBr, 10mmol/L Tricine-NaOH（pH 8.0�

关 键 词：海洋绿藻 假根羽藻 CYT b6f 分离 纯化 

分 类 号：Q5[生物学—生物化学]