Takabe T, Ishikawa H, Niwa S, Itoh S
J Biochem. 1983 Dec;94(6):1901-11. doi: 10.1093/oxfordjournals.jbchem.a134544.
Treatment of isolated spinach thylakoid fragments with Triton X-100 followed by repeated sucrose density gradient centrifugations and Sephacryl S-300 and DEAE-Sephacel chromatographies yielded a highly purified P700-chlorophyll a protein complex complex which consists of five polypeptides. The protein complex is virtually free of chlorophyll b (Ch1 alpha/Ch1 b greater than 10) with approximately 30 chlorophylls per P700, and contains iron-sulfur centers A, B, and X. At pH values higher than 6, divalent cations, but not monovalent or trivalent cations, efficiently accelerated the electron transfer from reduced spinach plastocyanin to the photooxidized P700 in the P700-chlorophyll alpha protein complex. At pH values lower than 6, the reaction rate drastically increased with decreasing pH with a maximum at about pH 4.3 without cations. Divalent salts as well as monovalent or trivalent salts decreased the P700 reduction rate at low pH, indicating the involvement of electrostatic interaction in those pH regions. The rate of electron transfer from plastocyanin to the photooxidized P700 in the reaction center protein, which consists of only the largest peptide subunit and no iron-sulfur centers, was reduced only 50% at pH 7.0 in the presence of MgCl2 as compared to the case of P700-chlorophyll alpha protein complex. Essentially similar effects of pH and metal ions on this electron transfer reaction were observed as in the case of P700-chlorophyll alpha protein complex. These results strongly suggest that plastocyanin donates electrons directly to the largest peptide of P700-chlorophyll alpha protein complex and the observed effects of pH and cations are mainly due to the interaction between the largest peptide of P700-chlorophyll alpha protein complex and plastocyanin. The four small subunits in the protein complex seemed to have only a minor role in the reaction with plastocyanin.
用Triton X-100处理分离的菠菜类囊体片段，随后进行反复的蔗糖密度梯度离心以及Sephacryl S-300和DEAE-Sephacel柱层析，得到了一种高度纯化的P700-叶绿素a蛋白复合物，该复合物由五条多肽组成。该蛋白复合物几乎不含叶绿素b（叶绿素α/叶绿素b大于10），每P700约有30个叶绿素，并且含有铁硫中心A、B和X。在pH值高于6时，二价阳离子而非一价或三价阳离子能有效地加速从还原的菠菜质体蓝素到P700-叶绿素α蛋白复合物中光氧化的P700的电子转移。在pH值低于6时，反应速率随着pH值降低而急剧增加，在约pH 4.3时达到最大值，此时无需阳离子。二价盐以及一价或三价盐在低pH值时会降低P700的还原速率，表明在这些pH区域存在静电相互作用。在反应中心蛋白中，仅由最大的肽亚基组成且不含铁硫中心，从质体蓝素到光氧化的P700的电子转移速率，在pH 7.0且存在MgCl2的情况下，与P700-叶绿素α蛋白复合物相比仅降低了50%。观察到pH值和金属离子对该电子转移反应的影响与P700-叶绿素α蛋白复合物的情况基本相似。这些结果强烈表明，质体蓝素直接向P700-叶绿素α蛋白复合物的最大肽提供电子，并且观察到的pH值和阳离子的影响主要是由于P700-叶绿素α蛋白复合物的最大肽与质体蓝素之间的相互作用。该蛋白复合物中的四个小亚基在与质体蓝素的反应中似乎只起次要作用。
