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Structural Fluctuation of PhotosyntheticAntenna Complexes Studied by Single-Molecule Spectroscopy

单分子光谱研究光合天线复合物的结构波动

基本信息

批准号：
16340121
负责人：
MATSUSHITA Michio
金额：
$ 10.56万
依托单位：
Tokyo Institute of Technology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2006
项目状态：
已结题

项目摘要

A protein has numerous meta-stable structures. In the physiological conditions. The structural fluctuation was pointed out for long years to be important, because the structural fluctuation plays essential role in regulating function of proteins. However, there have been no methods to elucidate the structural dynamics. To break through, we are aiming at getting structural information of single proteins by performing optical spectroscopy of individual proteins. Because measuring a fluorescence spectrum takes at least several milli-seconds, troscopic measurements are performed at temperatures low enough for the structural dynamics to be followed.In this study we have succeeded to follow one pigment-protein complex to know the temperature dependence of structural dynamics. The experimental data, which are the very first data on the globe show clearly that substantial part of the structural movement is temperature-independent tunneling motion [Oikawa, et. al. J. Am. Chem. Soc. 130(20008)48501.].Spectroscopy at low temperature of single proteins has been limited to the near infrared region. The method was successfully applied to photosynthetic antenna complexes of photosynthetic bacteria. Nevertheless it is worth making an effort to extend the wavelength of the setup to cover the visible region, since many enzymes absorb or emit in the visible. In order to reduce the chromatic aberration of the objective, which is immersed into liquid helium together with a sample, we have developed a reflecting objective [Fujiyoshi, et. al. Appl. Phys. Lett. 91 (2007) 051125.]. With the reflecting objective we have made two-photon excitation and measured fluorescence spectrum of single GFPs. The spectra were taken in the background-free condition, and series of spectra of the same protein revealed structural dynamics of the protein [Fujiyoshi, et. al. Phys. Rev. Lett. 100 (2008) 168101.].

蛋白质具有许多亚稳定结构。在生理条件下。蛋白质的结构涨落对蛋白质的功能调节起着至关重要的作用，因此结构涨落的重要性一直被人们所重视。然而，一直没有方法来阐明结构动力学。为了取得突破，我们的目标是通过对单个蛋白质进行光谱分析来获得单个蛋白质的结构信息。因为测量荧光光谱至少需要几毫秒，所以在足够低的温度下进行troscopic测量，以便跟踪结构动力学。在这项研究中，我们成功地跟踪了一种色素-蛋白质复合物，以了解结构动力学的温度依赖性。实验数据是地球仪上的第一批数据，它清楚地表明，结构运动的实质部分是与温度无关的隧道运动[Oikawa，et. J. Am. 130（20008）48501.]。单一蛋白质在低温下的光谱仅限于近红外区域。该方法已成功应用于光合细菌光合天线复合体的研究。然而，值得努力扩展设置的波长以覆盖可见光区域，因为许多酶在可见光中吸收或发射。为了减少物镜的色差，这是浸没在液氦与样品一起，我们已经开发了一种反射物镜[Fujiyoshi，等。等.应用物理学通讯. 91（2007）051125.]。利用反射物镜进行双光子激发，测量了单个玻璃纤维蛋白的荧光光谱。光谱是在无背景条件下拍摄的，并且同一蛋白质的一系列光谱揭示了蛋白质的结构动力学[Fujiyoshi，et.等，Phys. Rev. Lett. 100（2008）168101.]。