偏光赤外分光を用いたロドプシンの動的構造解析

使用偏振红外光谱分析视紫红质的动态结构

• 批准号: 11480193
• 负责人: KANDORI Hideki
• 金额: $ 9.79万
• 依托单位: KYOTO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B).
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-11480193/
• 关键词: polarized IR; infrared spectroscopy; bacteriorhodopsin; rhodpsin; hydrogen-bond; isotope-effect; mutant; structure changes

This research project aims at revealing the correlation between structure and function in rhodopsins by means of polarized infrared spectroscopy. The target molecules are bacteriorhodopsin (bR) as a light-driven proton pump, halorhodopsin (hR) as a light-driven chloride pump, rhodopsin (Rh) as a light-sensor in twilight vision, and color visual pigments (Chicken-Red and Chicken-Green) as light-sensors in color vision. Infrared spectroscopy of these proteins as well as their mutants and isotope-labeled molecules provided the following results.(1) Bridged water stretching vibrations inside bR under strong hydrogen bond were directly observed by highly refined polarized infrared spectroscopy. Rotation of a water molecule was observed accompanying retinal isomerization.(2) Polarized infrared spectroscopy of bR provides information on the structural changes of protein side upon photoisomerization (K intermediate). Investigation of isotope-labeled sample on threonine side chain and mutants r … More evealed that 3 of the threonines of a total of 18 change their hydrogen bonding. The structural change of Thr17, which is located > 11 Å from the retinal chromophore, implicates a specific perturbation channel in the protein that accompanies the retinal motion.(3) Polarized infrared spectroscopy of bR provides information on the structural changes of protein side before and after the primary proton transfer (L and M intermediates). It was found that hydrogen bond of Thr89 is strengthened upon photoisomerization, and the strong hydrogen bond is persistent even after the "switch" event for proton pump. The results implicate that the Thr89-Asp85 region is not involved in the switch, suggesting that the switch is indeed local machinery.(4) Protein structural changes of hR, Chicken-Red, Chicken-Green, and photoactive yellow protein were extensively studied by low-temperature infrared spectroscopy.(5) Excited-state dynamics of bovine rhodopsin was studied by femtosecond fluorescence spectroscopy. Less

该研究项目旨在通过偏光红外光谱揭示杜鹃花素的结构与功能之间的相关性。靶分子是细菌后紫红蛋白（BR），作为轻驱动的质子泵，甲霍顿蛋白（HR）作为轻驱动的氯化物泵，Rhodopsin（RH），作为暮光视觉中的轻度传感器，以及彩色视觉颜料（鸡肉红色和鸡肉），作为彩色视觉。这些蛋白质的红外光谱以及它们的突变体和同位素标记的分子提供了以下结果。（1）高度精制的极化感染光谱直接观察到了强氢键在强氢键下BR内部桥接的水拉伸振动。观察到水分子的旋转被观察到参与的残留异构化。（2）BR的极化感染光谱提供了有关光异构化后蛋白质侧的结构变化的信息（K中间）。在苏氨酸侧链和突变体上对同位素标记的样品进行了研究，更忽略了总共18种的三种三种苯胺改变了它们的氢键。 THR17的结构变化位于视网膜发色团中>11Å，它在蛋白质中实现了涉及视网膜运动的蛋白质中的特定扰动通道。（3）BR的偏振红外光谱法提供了有关主要质子转移之前和之后蛋白质方面的结构变化的信息（L和M InterMideDiates）。发现THR89的氢键在光异构化时得到增强，即使在质子泵的“开关”事件之后，强氢键也持续。结果暗示，THR89-ASP85区域不参与转换，这表明转换确实是本地机械。（4）HR，鸡肉红，鸡绿和光活性黄色蛋白的蛋白质结构变化被低温的红外光谱进行了广泛研究。较少的

项目成果

Kandori, H., Yamazaki, Y., Shichida, Y., Raap, J., Lugtenburg, J., Belenky, M., and Herzfeld, J.: "Tight Asp85-Thr89 Association during the Pump Switch of Bacteriorhodopsin"Proc.Natl.Acad.Sci.USA. 98. 1571-1576 (2001)

Kandori, H.、Yamazaki, Y.、Shichida, Y.、Raap, J.、Lugtenburg, J.、Belenky, M. 和 Herzfeld, J.：“细菌视紫红质泵开关期间的紧密 Asp85-Thr89 关联”Proc

Yamazaki Y.: "Fourier Transform Infrared Studies on Conformation Changes of bd-type Ubiguinol Oxidase from Escherichia coli upon Photoreduction of the Redox Metal Centers"J. Biochem.. 125. 1131-1136 (1999)

Yamazaki Y.：“基于氧化还原金属中心光还原作用的大肠杆菌 bd 型泛甘醇氧化酶构象变化的傅立叶变换红外研究”J.

Kandori, H., Kinoshita, N., Yamazaki, Y., and Shichida, Y.: "FTIR Spectroscopy Reveals Water Structural Changes of Bacteriorhodopsin upon Photoisomerization."in Fourier Transform Spectroscopy (K.Itoh and M.Tasumi, Eds), Waseda University Press. 455-456 (1

Kandori, H.、Kinoshita, N.、Yamazaki, Y. 和 Shichida, Y.：“FTIR 光谱揭示光异构化时细菌视紫红质的水结构变化。”傅里叶变换光谱（K.Itoh 和 M.Tasumi，编辑），

Imamoto, Y., Hirano, T., Imai, H., Kandori, K., Maeda, A., Yoshizawa, T., Groesbeek, M., Lugtenburg, J., and Shichida, Y.: "Anion Binding Effect to Iodopsin Studied by Low-Temperature FTIR Spectroscopy."Biochemistry. 38. 11749-11754 (1999)

Imamoto, Y.、Hirano, T.、Imai, H.、Kandori, K.、Maeda, A.、Yoshizawa, T.、Groesbeek, M.、Lugtenburg, J. 和 Shichida, Y.：“阴离子结合效应

Yamazaki, Y., Kandori, H., and Mogi, T.: "Effects of Amino-Acid Substitutions on Redox-induced Conformational Changes of the Escherichia coli bo-type Ubiquinol Oxidase Revealed by Fourier-transform Infrared Spectroscopy."J.Biochem.. 126. 194-199 (1999)

Yamazaki, Y.、Kandori, H. 和 Mogi, T.：“傅里叶变换红外光谱揭示氨基酸取代对氧化还原诱导的大肠杆菌 bo 型泛醇氧化酶构象变化的影响。”J.Biochem