Molecular Science for Elucidation of Proton Active Transport and Electron Transfers through Proteins.

通过蛋白质阐明质子主动传输和电子转移的分子科学。

• 批准号: 02453157
• 负责人: KITAGAWA Teizo
• 金额: $ 4.48万
• 依托单位: Okazaki National Research Institutes, Institute for Molecular Science
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1990
• 资助国家: 日本
• 起止时间: 1990 至 1991
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-02453157/
• 关键词: Hemoglobin; Cytochrome c Oxidase; Time-resolved resonance Raman; UV resonance Raman; Proton Transport; Electron Transfer; 時間分解ラマン; 共鳴ラマン分光; 酸素

Active transport of protons by proteins is presumably based on pKa changes of some proton-carrying residues upon conformation change. In order to examine protonation-dependent protein dynamics, time-resolved ultraviolet resonance Raman spectra were pursued for the quaternary structure change of hemoglobin (Hb) from R to T upon photodissociation of its carbonmonoxide adduct. Raman spectra were probed by laser pulses at 218nm with 10ns width, while CO was photodissociated by another laser pulses at 416nm with lons width. The delay time of the probe pulse from the pump pulse was varied from -200mus to 500mus. The relative intensity of a tryptophane band at 1011cm^<-1> to a phenylalanine band at 1003cm^<-1> changed with the quaternary structure change. The change took place at 5 - 20mus after photolysis at pH7.4 but it had been completed at lOmus after photolysis at pH5.8 where Bohr residues are protonated. We deduced the tryptophane residue responsible for the Raman spectral change is beta37-Trp and discussed a mechanism by which a structural change of heme in alpha subunit is communicated to the beta subunit.We also investigated reaction intermediates of cytochrome oxidase by using time-resolved resonance Raman spectroscopy and proposed likely structures of partially reduced oxygen. The results are published separately.

蛋白质对质子的主动转运可能是基于构象变化时一些携带质子的残基的 pKa 变化。为了检查质子化依赖性蛋白质动力学，我们利用时间分辨紫外共振拉曼光谱来研究血红蛋白 (Hb) 一氧化碳加合物光解离后从 R 到 T 的四级结构变化。拉曼光谱通过 218nm、10ns 宽度的激光脉冲探测，而 CO 通过另一个 416nm、lons 宽度的激光脉冲光解离。泵浦脉冲的探测脉冲延迟时间从-200mus 到500mus 不等。 1011cm^-1处的色氨酸带与1003cm^-1处的苯丙氨酸带的相对强度随着四级结构的变化而变化。该变化发生在pH7.4光解后5-20mus，但在pH5.8光解后10mus完成，其中玻尔残基被质子化。我们推断引起拉曼光谱变化的色氨酸残基是β37-Trp，并讨论了α亚基中血红素结构变化传递给β亚基的机制。我们还利用时间分辨共振拉曼光谱研究了细胞色素氧化酶的反应中间体，并提出了部分还原氧的可能结构。结果单独发表。

项目成果

北川 禎三: "ラマン分光,第4版 実験化学講座" 372-388 (1991)

Teizo Kitakawa：“拉曼光谱，第四版实验化学课程”372-388（1991）

S.Hashimoto,Y.Mizutani,Y.Tatsuno,T.Kitagawa: "Resonance Raman characterization of ferric-and ferrylporphyrin πcation radicals and the Fe^<IV>=0 stretching frequency" J.Am.Chem.Soc.113. 6542-6549 (1991)

S. Hashimoto、Y. Mizutani、Y. Tatsuno、T. Kitakawa：“三价铁和铁卟啉 π 阳离子自由基的共振拉曼表征以及 Fe^<IV>=0 伸缩频率”J.Am.Chem.Soc.113。 -6549 (1991)

T.Egawa,T.Ogura,R.Makino,Y.Ishimura,T.Kitagawa: "Observation of the 0-0 stretching Raman band for cytochrome P-450_<cam> under catalytic conditions" J.Biol.Chem.266. 10246-10248 (1991)

T.Ekawa、T.Ogura、R.Makino、Y.Ishimura、T.Kitakawa：“催化条件下细胞色素 P-450_<cam> 0-0 拉伸拉曼带的观察”J.Biol.Chem.266。

T.Ogura,T.Kitagawa,S.Takahashi,K.Shinzawa,S.Yoshikawa: "Observation of the Fe^<4+>+O stretching Raman band fof cytochrome oxidase compound B at ambient temperature." Journal of Biological Chemistry. 265. 14721-14723 (1990)

T.Ogura、T.Kitakawa、S.Takahashi、K.Shinzawa、S.Yoshikawa：“在环境温度下观察细胞色素氧化酶化合物 B 的 Fe^<4>O 拉伸拉曼带。”

N. Kitajima, T. Koda, S. Hashimoto, T. Kitagawa, Y. Moro-oka: "Synthesis and characterization of the dinuclear copper (II) complexes [Cu (HB (3, 5-Me_2pz)_3) ]_2 X (X=O^<2->, (OH)_2^<2->, CO_3^<2->, O_2^<2->)" J. Am. Chem. Soc.113. 5664-5671 (1991)

N. Kitajima、T. Koda、S. Hashimoto、T. Kitakawa、Y. Moro-oka：“双核铜 (II) 配合物 [Cu (HB (3, 5-Me_2pz)_3) ]_2 X 的合成和表征