Investigation of the functional mechanisms of artificial photosynthetic pigment-protein supra-molecular complexes

三种不同的色素-蛋白质复合物（两个称为LH 2和LH 1的触角以及反应中心（RC））深深参与了紫色光合细菌的初级过程。在本研究项目中，为了更深入地了解这些色素-蛋白质复合物的功能机制，广泛研究了人工色素-蛋白质复合物，其中类胡萝卜素色素的结构通过利用有机合成进行了修饰。1.实现了类胡萝卜素类似物的大规模全合成：实现了类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidoxanthin、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidoxanthin、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidoxanthin、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroidene、类胡萝卜素类似物spheroid类胡萝卜素的类似物类胡萝卜素被整合到光合细菌的无类胡萝卜素反应中心（RC）中， ...更多信息 球形红杆菌R26.1菌株。将制备的人工RC结晶，并通过X射线晶体学测定其晶体结构。这些结构分析揭示了类胡萝卜素分子被掺入RC时的单向插入。PheM162是门控氨基酸残基，而TrpM75是当类胡萝卜素分子重新并入R26.1 RC时的锁定氨基酸残基。3.天然LH 2天线复合物上的电吸收光谱：双锁相电吸收光谱在检测中被认为是最有前途的方法之一，以检测色素和载脂蛋白之间的静电相互作用，色素蛋白复合物建立了精确的实验方案和分析方法，并将其应用于光合细菌Rb，sphaeroides菌株GIC的天然LH2复合物。4.人工反应中心的电吸收光谱，其中类似物类胡萝卜素已重新纳入：电吸收光谱应用于Rb的无类胡萝卜素反应中心（RC）。1以及其中已掺入类似物类胡萝卜素的相同RC。我们已经，第一次，定量地确定了由周围的细菌叶绿素分子绑定到RC的类胡萝卜素分子的存在或不存在诱导的静电环境的差异。5.子20 fs的时间分辨吸收光谱的类胡萝卜素：时间分辨吸收光谱与前所未有的时间分辨率子20 fs的应用到F3-胡萝卜素和番茄红素。首次在真实的时域中对介于单光子允许态1^1B_u ^+（S_2）和单光子禁止态2^1Ag ^-（S_1）之间的中间单重激发态（Sx态）进行了时间分辨，证明了该中间激发态参与了光激发到1^1B_u ^+态的弛豫过程.本研究扩展到具有不同多烯链长的系列胡萝卜素分子。在真实的时域中，除了具有5个共轭C = C键的胡萝卜素分子外，所有的胡萝卜素分子在从S_2到S_1的弛豫过程中都具有S_x态。红及其电吸收光谱：来自红的B880色素蛋白复合物的无类胡萝卜素单体亚基。红色菌株G9+与纯化的全反式油菜黄素重组，形成人工捕光1（LH 1）复合物。将电吸收光谱应用于如此重构的LH1，以评估复合物中色素和脱辅基蛋白之间的静电相互作用。少

Three different kinds of pigment-protein complexes (two antennae called LH2 and LH1, and reaction center (RC)) are deeply involved in the primary process of purple photosynthetic bacteria. In this research project, for the deeper understanding of the functional mechanisms of these pigment-protein complexes, artificial pigment-protein complexes in which the structure of the carotenoid pigments were modified by utilizing organic synthesis were extensively studied. The following remarkable features have been achieved.1.Achievement of the large-scale total synthesis of analogue carotenoids : Larges-scale total syntheses of a series of analogue carotenoids of spheroidene, spirilloxanthin, and lycopene which have different extent of conjugation were achieved.2.X-ray crystallography of the reaction centers in which the analogue carotenoids have been incorporated : The analogue carotenoids of spheroidene were incorporated into the carotenoidless reaction center (RC) from a photosynthetic bacte … More rium Rhodobacter sphaeroides strain R26.1. Thus prepared artificial RCs were crystallized and their crystal structures were determined by X-ray crystallography. These structure analyses revealed the unidirectional insertion of the carotenoid molecule when it is incorporated into the RC. PheM162 is a gate-keeper amino-acid residue, while TrpM75 is a locking amino-acid residue when the carotenoid molecule is re-incorporated into the R26.1 RC.3.Electroabsorption spectroscopy on the native LH2 antenna complex : Electroabsorption spectroscopy using dual-phase lock-in detection was proposed to be one of the most promising methods to detect the electrostatic interaction between the pigments and apoproteins in the pigment-protein complexes. Exact experimental protocols together with the analysis methods were constructed, and they were applied to the native LH2 complex from a photosynthetic bacterium Rb, sphaeroides strain GIC.4.Electroabsorption spectroscopy of the artificial reaction centre in which an analogue carotenoid has been re-incorporated : Electroabsorption spectroscopy was applied to the carotenoidless reaction centre (RC) from Rb. sphaeroides strain R26.1 as well as the same RC in which an analogue carotenoid has been incorporated. We have, for the first time, quantitatively determined the difference of the electrostatic environment induced by the presence or absence of a carotenoid molecule around the bacteriochlorophyll molecules bound to the RC.5.Sub-20 fs time-resolved absorption spectroscopy of carotenoids : Time-resolved absorption spectroscopy with unprecedented temporal resolution of sub-20 fs was applied to f3-carotene and lycopene. The intermediate singlet excited-state (Sx state) in-between the one-photon allowed 1^1B_u^+(S_2) and forbidden 2^1Ag^-(S_1) states was time-resolved for the first time in the real time-domain, and it was evidenced that this intermediate excited state was involved in the relaxation process following the photoexcitation up to the 1^1B_u^+ state. This study was extended to the series of carotene molecules which have different polyene chain-lengths. Except the case of the carotene molecule having 5 conjugated C=C bonds, all the carotene molecules were evidenced in real time-domain to have the S_x state in the relaxation process from S_2 to S_1.6.Reorganization of the LHI antenna complex of Rs. rubrum and its electroabsorption spectroscopy : Carotenoid-free monomeric subunits of B880 pigment-protein complex from Rs. rubrum strain G9+ was reorganized with purified all-trans-spirilloxanthin to form the artificial light-harvesting 1(LH1) complex. Electroabsorption spectroscopy was applied to thus reconstituted LH1 in order to evaluate electrostatic interaction between pigments and apo-proteins in the complex. Less