Structure of cytochrome b6/f deduced from its function

从细胞色素 b6/f 的功能推导出其结构

• 批准号: 06640848
• 负责人: KOIKE Hiroyuki
• 金额: $ 1.28万
• 依托单位: Himeji Institute of Technology
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1995
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-06640848/
• 关键词: Cytochrome b6; f complex; thermophilic cyanobacterium; thylakoid membranes; heptylthioglucoside; duroquinone; plastoquinone

Cytochrome b6/f complex accepts electrons from plastoquinol which was reduced by photosystem II.The complex transfers electrons to photosystem I and forms H^+ gradient across the thylakoid membranes to form ATP simultaneously. In this research project, we have purified the complex from thylakoid membranes of a cyanobacterium focussing on development of simple purification procedure and its high yield. We also have investigated the substrate (quinone) specificity of the complex and estimated the structure of active site of the quinone binding pocket.The complex was purified from a thermophilic cyanobacterium, Synechococcus vulcanus. We have improved on the existing procedure by the following two points : 1) thylakoid membranes were sonicated simultaneously during NaBr treatment to remove all the extrinsic proteins and 2) deoxycholate was added together with heptylthioglucoside to solubilize the complex. By this improvement, the complex was effectively precipitated in a narrow range of ammonium sulfate concentration and could be purified further.Tetramethyl-p-benzoquinone is normally used for activity measurement. Trimethyl-p-benzoquinone was found to be effective as well. Dimethyl-p-benzoquinones or dichloro-p-benzoquinones (DCBQ) were, however, not avtive as electron donor to the complex. Judging from the fact that DCBQ is the most effective electron acceptor for photosystem II,the structure of quinone binding pocket of b6/f complex is rather different from that of Q_B site of photosystem II.

细胞色素b6/f复合物接受光系统II还原的质体醌醇的电子，将电子传递给光系统I，同时形成H^+梯度穿过类囊体膜形成ATP。在本研究中，我们从蓝藻类囊体膜中纯化了该复合物，重点是开发简单的纯化程序和高产量。我们还研究了复合物的底物（醌）特异性，并估算了醌结合口袋活性位点的结构。我们对现有的方法进行了以下两点改进：1）在NaBr处理的同时超声处理类囊体膜以去除所有的外源蛋白; 2）加入脱氧胆酸盐和庚基硫代葡萄糖苷以溶解复合物。在较窄的硫酸铵浓度范围内，络合物得到有效沉淀，并可进一步纯化。发现三甲基对苯醌也是有效的。而二甲基对苯醌和二氯对苯醌（DCBQ）则不能作为配合物的电子给体。从DCBQ是光系统II最有效的电子受体这一事实判断，b6/f复合物的醌结合口袋结构与光系统II的Q_B位点结构有很大不同。