褐藻色素蛋白复合物的分离及特性研究

【作者】 刘洪艳；

【导师】 侯和胜； 王广策；

【作者基本信息】 辽宁师范大学 ， 植物学， 2002， 硕士

【摘要】 以褐藻裙带菜(Undaria pinnatifida)为实验材料，建立一套能够分离出有光活性色素蛋白复合物的技术路线。通过对裙带菜色素蛋白复合物分离技术及其影响因素的研究，确立了褐藻的SDGC分离方法。采用蔗糖密度梯度超速离心的方法，以去污剂SDS为增溶剂(SDS：Chl=20：1，4℃增溶20分钟)蔗糖密度梯度为60％、50％、40％、30％、20％、15％和10％，从裙带菜中成功地分离出5条含色素的蛋白质复合物带。 测定裙带菜各色素蛋白复合物的吸收光谱、荧光发射光谱和荧光激发光谱，并进行了DCIP的光还原测定和化学法的氧化还原差示光谱测定。确定了10％和15％层带的色素蛋白复合物为捕光色素复合物；20％和30％层带的复合物为PSⅡ颗粒，具有DCIP的光还原活性；40％层带复合物为去除了捕光色素的PSⅠ复合物。对于有DCIP光还原活性的20％和30％层带的复合物，进一步DEAE离子交换层析纯化。150mM NaCl洗脱纯化后的样品经过荧光激发光谱测定发现，已经去除了叶绿素c和墨角藻黄素，并且仍然具有DCIP的光活性，分析是PSⅡ核心复合物。 在裙带菜纯化的类囊体膜和PSⅠ复合物的荧光光谱中，均未发现属于PSⅠ的730 nm的荧光发射峰，指出730 nm荧光峰不能作为PSⅠ的表征。更多还原

【Abstract】 The separating technique of the light active pigment-protein complexes from brown algae Undaria pinnatifida was developped in this paper. The results of SDGC separation were observed and the separating technique of brown algae was established. Five pigment-protein complexes were isolated by centrifugation on discontinuous sucrose density gradient of the SDS-treated thylakoid membrane. The best separation was obtained with detergent SDS (SDS:Chl=20:l, solubilized 20 min at 4’C), sucrose density gradient (10,15,20,30,40,50,60% w/w).Absorption, fluorescence emission and excitation spectra of five pigment-protein complexes were determined. Photosynthetic electron transfer was measured from the DCIP photoreduction. P700 concentration was assayed from the ferricyanide -oxdised minus ascorbate-reduced difference spectrum. 10% and 15% sucrose layers were identified as light harvesting complexes; 20% and 30% sucrose layers were identified as composed of active PS II particles; 40% sucrose layer represented rather pure PS I complex. The PS II native fractions(20% and 30%) were loaded onto a DEAE column.The fraction eluted with 150 mM NaCl was presented DCIP reduction activity and was highly depleted in Chi c and xanthophylls, and as such could be considered a PS II core complex.The fluorescence spectra of brown algae thylakoid membrane and PS I complex varied from higher plants, they had no fluorescence emission maximum at 730 nm. we concluded that 730 nm fluorescence was no more a symbol of PS I .更多还原