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化学学报
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化学学报 ›› 2011, Vol. 69 ›› Issue (13): 1559-1564. 上一篇    下一篇

研究论文

光诱导高铁肌红蛋白还原的光谱法研究

周华伟1, 曹洪玉1, 唐乾1, 安良梅1, 郑学仿*,1,2   

  1. (1大连大学生物工程学院 大连 116622)
    (2辽宁省生物有机化学重点实验室 大连大学 大连 116622)
  • 投稿日期:2010-08-18 修回日期:2011-01-21 发布日期:2011-03-03
  • 通讯作者: 郑学仿 E-mail:dlxfzheng@163.com
  • 基金资助:

    国家自然基金;辽宁省高校创新团队项目;大连市科技计划项目

Spectral Study on the Photoreduction of Metmyoglobin

Zhou Huawei1; Cao Hongyu1; Tang Qian1; An Liangmei1; Zheng Xuefang*,1,2   

  1. (1 College of Bioengineering, Dalian University, Dalian 116622)
    (2 Liaoning Key Laboratory of Bio-organic Chemistry, Dalian University, Dalian 116622)
  • Received:2010-08-18 Revised:2011-01-21 Published:2011-03-03
  • Contact: 仿 学郑 E-mail:dlxfzheng@163.com

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1493

被引次数

22

采用紫外-可见吸收、荧光及圆二色(CD)等光谱法为主要分析手段, 对光诱导高铁肌红蛋白(metmyoglobin, metMb)还原及其引起的蛋白结构变化进行了研究. 采用氙灯430 nm的单色光和紫外灯对高铁肌红蛋白溶液进行光照实验, 实验结果显示, 在metMb的Q吸收带545 nm和580 nm附近吸收峰强度显著增强. 通过验证性实验, 证明光照后metMb发生了还原反应. 验证性实验包括: metMb中加入还原剂连二亚硫酸钠实验|光照后样品加入氧化剂铁氰化钾的可逆性实验|光照的同时通入CO实验. 进一步讨论表明, 采用氙灯430 nm的单色光诱导metMb还原的机理为光诱导的分子内电子转移. 此外, 对光还原反应后的样品进行同步荧光和CD光谱分析, 发现蛋白质色氨酸残基微环境极性增加, α-螺旋的含量由 63%下降到57%, 而β-折叠的含量增加了7%.

关键词: 高铁肌红蛋白, 光还原, 激发态, 结构变化

The photoreduction of metmyoglobin (metMb) and its effects on the changes of protein structure were investigated by UV-visible absorbance, fluorescence and circular dichroism (CD) spectra. MetMb in Na2HPO4-NaH2PO4 buffer were irradiated with xenon lamp at 430 nm and ultraviolet lamp. The results indicated that the absorbance intensity at Q band (545 nm and 580 nm) increased evidently after illumination. Through experimental verification, metMb had been reduced after the illumination. The verifying experiment included: adding sodium dithionite to metMb|adding potassium ferricyanide to the sample after the illumination|irradiating meanwhile adding CO to the sample. Through further discussion, it can be seen that the photoreduction mechanism of metMb with xenon lamp at 430 nm was photoinduced intramolecular electron transfer. In addition, the results of synchronous fluorescence and CD spectra of the sample after irradiation suggested that the polarity of microenvironment of tryptophan residues increased and the secondary structure of protein (CD data) varied with α-helices reduction from 63% to 57%, and β-sheet increase by 7%.

Key words: metmyoglobin, photoreduction, excited state, structure changes