Direct observation of molecular assembly of photosynthetic membrane proteins in solid-supported lipid bilayers

直接观察固相脂质双层中光合膜蛋白的分子组装

• 批准号: 18550150
• 负责人: DEWA Takehisa
• 金额: $ 2.64万
• 依托单位: Nagoya Institute of Technology
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2006
• 资助国家: 日本
• 起止时间: 2006 至 2007
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18550150/
• 关键词: photosynthesis; membrane protein; supported lipid bilaver; direct observation; organization; molecular assembly

This research has aimed to establish experimental platforms using solid-supported lipid bilayer in order to reveal the relation between the assembling structure and function of photosynthetic membrane proteins, light-harvesting complex 2 (LH2) and light-harvesting mm complex (LH1-RC). Recently, the array structure of LH2 and LH1-RC in a bacterial photosynthetic membrane has been revealed by atomic force microscopy (AFM). In the membrane, hexameric LH1-RCs were surrounded by LH2 molecules, forming 'protein nay' structure. We attempted to construct assembly of LH1-RC and LH2 in a supported lipid bilayer membrane.LH1, reaction center(RC), and LH1-RC complexes were successfully assembled onto an amino-modified ITO electrode. Enhanced photocurrent generation by RC was observed when the RC was assembled with LHl, indicating that light harvesting and energy transfer by LH1 could work on the electrode.We have established a novel method for incorporation of LH2 into a phew-separated lipid domai … More n in the membrane. In brief, LH2 incorporated into a cationic vesicle cargo (proteoliposome) is selectively arrayed on the fluid domain of the anionic planar membrane via vesicle fusion and subsequent lateral diffusion. The prceess of arraying has been observed by total internal reflection fluorescence (TIRF) microscopy An LH2/LLH1-RC coexistent array in a supported lipid bilayer was successfully constructed by consecutive deposition of vesicle cargos incorporating LH2 and LH1-RC onto an amino-modified coverslip.LH1, reaction center (RC), and LH1-RC complexes were successfully assembled onto an amino-modified ITO electrode. Enhanced photocurrent generation by RC was observed when the RC was assembled with LH1, indicating that light harvesting and energy transfer by LH1 could work on the electrode.A planar lipid bilayer tethered by an avidin-immobilized substrate via avidin-biotin interaction was successfully constructed. The architecture provides thin water layer between the planar membrane and the substrate. The water layer allows the LH1-RC possessing large protrudent hydrophilic domain to diffuse laterally within the membrane. As the experimental platform, this architecture makes it possible to investigate dynamic assembling of LH1-RC and LH2 in the planar membrane Less

本研究旨在利用固体支撑脂质双层膜建立实验平台，以揭示光合膜蛋白捕光复合物2（LH 2）和捕光复合物mm（LH 1-RC）的组装结构与功能之间的关系。最近，原子力显微镜（AFM）揭示了细菌光合膜中LH 2和LH 1-RC的阵列结构。在细胞膜中，六聚体LH 1-RC被LH 2分子包围，形成“蛋白质奈”结构。我们尝试构建LH 1-RC和LH 2在支撑脂质双层膜中的组装体，成功地将LH 1、反应中心（RC）和LH 1-RC复合物组装到氨基修饰的ITO电极上。当RC与LH 1组装时，观察到RC的增强的光电流产生，表明LH 1可以在电极上进行光捕获和能量传递。 ...更多信息 膜中的N。简而言之，LH 2掺入到阳离子囊泡货物（蛋白脂质体）选择性地排列在阴离子平面膜的流体域通过囊泡融合和随后的横向扩散。利用全内反射荧光（TIRF）显微镜观察了阵列的形成过程。通过将含有LH 2和LH 1-RC的囊泡货物连续沉积在氨基修饰的盖片上，成功构建了负载脂双层中LH 2/LLH 1-RC共存的阵列。LH 1、反应中心（RC）和LH 1-RC复合物成功组装到氨基修饰的ITO电极上。结果表明，LH 1能够有效地捕获光并传递能量，从而使RC的光电流产生增强，并成功构建了一个由亲和素固定的基底通过亲和素-生物素相互作用连接的平面脂双层膜。该结构在平面膜和基底之间提供薄水层。水层允许LH 1-RC具有大的亲水性结构域，以横向扩散内的膜。作为实验平台，该结构使得研究LH 1-RC和LH 2在平面膜中的动态组装成为可能。

项目成果

光合成でのアンテナ系モデルタンパク質/色素複合体の金基板上での電子伝達挙動

光合作用过程中金基底上基于天线的模型蛋白质/色素复合物的电子转移行为

• 发表时间: 2008
• 作者: 大坂 伸一郎;他
• 通讯作者: 他

Reconstitution of antenna LH1 complex from purple photosynthetic bacterium and its functional analysis of carotenoids

紫色光合细菌天线LH1复合体的重建及其类胡萝卜素功能分析

• 发表时间: 2008
• 作者: K. Nakagawa;et. al.
• 通讯作者: et. al.

Organization of Photosynthetic Membrane Proteins in Supported Lipid Bilayers (invited)

支持脂质双层中光合膜蛋白的组织（特邀）

• 发表时间: 2007
• 作者: 中尾亮太;堤由佳;東元祐一郎;野口正人;安永卓生;坂本寛;T. Dewa
• 通讯作者: T. Dewa

光合成細菌由来のアンテナ系タンパク質複合体の基板上への組織化と機能評価

源自光合细菌的天线蛋白复合物在基质上的组织和功能评估

• 发表时间: 2007
• 作者: 中尾亮太;堤由佳;東元祐一郎;野口正人;安永卓生;坂本寛;T. Dewa;堤 由佳;T.Dewa (invited);中島 正太;佐藤 秀明;出羽毅久;杉島 正一;T. Dewa;中尾 亮太;竹内稔和;大村 昇;坂本寛;中川勝統;八木達彦;水野愛弓;安永卓生;櫻井智彦;福井直美;竹内 稔和;大坂 伸一郎;中川勝統;中川勝統;廣 昭人;畑佐幹男
• 通讯作者: 畑佐幹男

固定化脂質二分子膜を用いた生体分子の組織化と動態観察

固定化脂质双层膜生物分子的组织和动态观察

• 发表时间: 2006
• 作者: M.Yoshiki;R.Hayashi;D.Sugiyama;S.Osada;I.Fujita;他;出羽毅久
• 通讯作者: 出羽毅久