Elucidation of Reaction Mechanisms of Biological Nitric Oxide by Vibrational Spectroscopy

振动光谱法阐明生物一氧化氮的反应机制

• 批准号: 07454157
• 负责人: KITAGAWA Teizo
• 金额: $ 5.06万
• 依托单位: (Institute for Molecular Science) Okazaki National Research Institutes
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07454157/
• 关键词: Nitric Oxide; Guanylate Cyclase; Resonance Raman; Vibrational Spectroscopy; NO stretching mode; 赤外吸収; ダイオードレーザー; 振動分光法; 共鳴ラマン散乱

Soluble guanylate cyclase (sGC) was isolated from bovine lung and its resonance Raman (RR) spectra were investigated for the reduced, CO-bound (CO-sGC), NO-bound (NO-sGC), oxidized, and oxidized NO-bound forms in the presence and absence of GTP.The RR spectra of sGC and CO-sGC,including the Fe-His stretch at 204 cm^<-1> and Fe-CO stretch at 473 cm^<-1>, were unaltered by binding of GTP,but apparent RR spectra of NO-sGC in the presence of GTP changed with time and concentrations of GTP.In the absence of GTP,the RR bands of N-O stretch (nu_<NO>) and Fe-No stretch (nu_<Fe-NO>) were observed at 1679 and 521 cm^<-1>, respectively. In its presence, two types of RR spectra were obtained, but after all GTP was exhausted by the enzymatic reaction, the spectrum for the absence of GTP was restoed. In one type of RR spectra, which appeared prior to the other, no NO-isotope sensitive band was detected and is heme spectrum was of a five-coordinate ferric high-spin type. In the other type, two nu_<NO> bands were observed at 1700 and 1681 cm^<-1>, which exhibited ^<15>NO isotopic frequency shifts of 21 and 34 cm^<-1>, respectively. Structural implications of the two types of RR spectra arizing from reaction intermediates have been discussed.

从牛肺中分离出可溶性鸟苷酸环化酶（sGC），研究了其还原型、CO结合型（CO-sGC）、NO结合型（NO-sGC）、氧化型和氧化型NO结合型的共振拉曼（RR）光谱。sGC和CO-sGC的RR光谱，包括204 cm ~ 2处的Fe-His伸缩峰<-1>和473 cm ~ 2处的Fe-CO伸缩峰<-1>，不受GTP结合的影响。在无GTP存在时，在1679和521 cm ~ 2处分别观察到N-O伸缩（nu_<NO>）和Fe-No伸缩（nu_<Fe-NO>）的RR谱带<-1>。在有GTP存在时，得到两种类型的RR光谱，但在所有GTP被酶促反应耗尽后，恢复了不存在GTP时的光谱。在一种类型的RR光谱，出现在另一个之前，没有NO同位素敏感带被检测到，是血红素光谱是一个五坐标铁高自旋型。在另一种类型中，<NO>在1700和1681 cm^处观察到两个nu_带<-1>，其分别显示出<15>21和34 cm^的^ NO同位素频移<-1>。讨论了反应中间体的两类RR谱的结构意义。

项目成果

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T.Kohzuma：“来自环碎无色杆菌的 1 型铜蛋白假天青蛋白活性位点转变的光谱和电化学研究。”

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D.A.Proshlyakov：“同时测定酶反应中间体拉曼光谱和吸收光谱的微循环系统及其在细胞色素 C 氧化酶与过氧化氢反应中的应用。”

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S.Hirota：“观察非基本 Fe-O_2 和 Fe-CO 振动以及氧和碳单氧血红蛋白的潜在非谐性；支持 Fe-C-O 弯曲基本原理新分配的证据。”

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M.Nagai：“使用色氨酸 b37 突变体对血红蛋白四级结构进行紫外共振拉曼研究。”