Study of the regulation mechanism of ligand affinity of hemoglobin by protein engineering
蛋白质工程研究血红蛋白配体亲和力调控机制
基本信息
- 批准号:05670043
- 负责人:
- 金额:$ 1.28万
- 依托单位:
- 依托单位国家:日本
- 项目类别:Grant-in-Aid for General Scientific Research (C)
- 财政年份:1993
- 资助国家:日本
- 起止时间:1993 至 1994
- 项目状态:已结题
- 来源:
- 关键词:
项目摘要
The oxygen affinity of hemoglobin is regulated in a range of 25,000-folds depending on the living environment of animal species. To know by what mechanism the regulation is realized from amino acid sequence differences in protein moiety, we synthesized eight artificial mutants of human hemoglobin with particular amino acids replaced by others by means of protein engineering based on site-directed mutagenesis and elucidated their characteristics such as oxygen binding properties, giving the following results.1. Proximal His mutants : Six mutants, in which the heme iron-bound invariable His-87alpha residue is replaced by either Leu, Val, Ile, Ala or Phe, or His-92beta is replaced by Tyr, showed such oxygen equilibrium properties as 36-fold variation of oxygen affinity, various decreases in the effect of IHP(inositol hexaphosphate), the Bohr effect and cooperativity, and increase in autooxidation of heme iron.2. Thr-38alpha mutants : Two mutants in which Thr-38alpha, participating in hydrogen bond formation between the alpha1-beta2 subunits, is replaced by Ser or Val were synthesized. The former showed almost normal oxygen binding properties whereas the latter showed somewhat altered properties (2.8-fold increase in oxygen affinity). The analysis by means of electronic, vibration and proton NMR Spectroscopy indicated that their high order structure is normal.3. The present experimental results indicate that the residues at the proximal side extensively participate in oxygen affinity regualtion whereas Thr-38alpha does not so much. They also indicate that a 36-fold regulation of oxygen affinity can be realized by the proximal residue. However, examination with other investigators' data shows that the affinity reguation by replacements of only the residues surrounding the heme group is far from the 25,000-fold variation, suggesting that some other mechanism must be invoked.
根据动物物种的生活环境,血红蛋白的氧亲和力被调节在25,000倍的范围内。为了了解蛋白质部分氨基酸序列差异是通过什么机制实现这种调节的,我们利用基于定点突变的蛋白质工程方法合成了8个特定氨基酸取代的人血红蛋白人工突变体,并阐明了它们的氧结合性质等特征,给出了以下结果。His近端突变体:用Leu、Val、Ile、Ala或Phe或His-92β替换为Leu、Val、Ile、Ala或Phe,或用Tyr替换His-92β的6个突变体,表现出氧亲和力变化36倍、IHP(六磷酸肌醇)效应、玻尔效应和协作性降低,以及血红素铁的自氧化增加等氧平衡特性。Thr-38pha突变体:合成了两个突变体,其中参与α1-β2亚基之间氢键形成的Thr-38pha被Ser或Val取代。前者表现出几乎正常的氧结合性质,而后者表现出一些改变的性质(氧亲和力增加2.8倍)。电子光谱、振动光谱和质子核磁共振波谱分析表明,它们的高阶结构正常。目前的实验结果表明,近端残基广泛参与氧亲和力调节,而Thr-38α不参与。他们还表明,近端残基可以实现36倍的氧亲和力调节。然而,与其他研究人员的数据检验表明,仅通过取代血红素基团周围残基的亲和力调节远远不是25,000倍的变化,这表明必须调用其他机制。
项目成果
期刊论文数量(14)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Imai, K.: "Molecular mechanism of hemoglobin function (in Japanese)" Proteins, Nucleic Acids and Enzymes. 39 (7). 1102-1110 (1994)
Imai, K.:“血红蛋白功能的分子机制(日语)”蛋白质、核酸和酶。
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- 影响因子:0
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- 通讯作者:
Imai, K.: "Adair fitting to oxygen equilibrium curves of hemoglobin" Methods Enzymol.232. 559-576 (1994)
Imai, K.:“Adair 拟合血红蛋白的氧平衡曲线”Methods Enzymol.232。
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- 影响因子:0
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Hashimoto,M.et al.: "Site-directed mutagenesis in hemoglobin:Functional and structural study of the intersubunit hydrogen bond of threonine-38(C3)α at the α1-β2 interface in human hemoglobin" Biochemistry. 32. 13688-13695 (1993)
Hashimoto, M. 等人:“血红蛋白定点诱变:人血红蛋白 α1-β2 界面处苏氨酸-38(C3)α 亚基间氢键的功能和结构研究”生物化学 32。13688-13695。 (1993)
- DOI:
- 发表时间:
- 期刊:
- 影响因子:0
- 作者:
- 通讯作者:
Hashimoto, M.et al: "Site-directed mutagenesis in hemoglobin : Functional and structural study of the intersubunit hydrogen bond of threonine-38 (C3) alpha at the alpha1-beta2 interface in human hemoglobin." Biochemsitry. 32 (49). 13688-13695 (1993)
Hashimoto, M.等人:“血红蛋白定点诱变:人血红蛋白 alpha1-beta2 界面上苏氨酸 38 (C3) α 亚基间氢键的功能和结构研究。”
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- 影响因子:0
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今井 清博: "ヘモグロビン機能の分子論" 蛋白質核酸酵素. 39. 1102-1110 (1994)
Kiyohiro Imai:“血红蛋白功能的分子理论”蛋白质核酸酶。39。1102-1110(1994)
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- 影响因子:0
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IMAI Kiyohiro其他文献
IMAI Kiyohiro的其他文献
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{{ truncateString('IMAI Kiyohiro', 18)}}的其他基金
Clarification of hemoglobin evolution process by reverse molecular evolution
通过逆向分子进化阐明血红蛋白进化过程
- 批准号:
22570217 - 财政年份:2010
- 资助金额:
$ 1.28万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Experimental verification of acquisition of hemoprotein high-order functions by reverse molecular evolution
逆向分子进化获得血红素蛋白高阶功能的实验验证
- 批准号:
19570222 - 财政年份:2007
- 资助金额:
$ 1.28万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Structure, function and evolution of cyclostomata hemoglobin and myoglobin
环口血红蛋白和肌红蛋白的结构、功能和进化
- 批准号:
13680740 - 财政年份:2001
- 资助金额:
$ 1.28万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Molecular manipulation of oxygen binding proteins and new development of precise structure-function studies on them
氧结合蛋白的分子操控及其精确结构功能研究新进展
- 批准号:
07308071 - 财政年份:1995
- 资助金额:
$ 1.28万 - 项目类别:
Grant-in-Aid for Scientific Research (A)
Study on the differentiation of physiological function of hemoglobin using artificial mutants
人工突变体对血红蛋白生理功能分化的研究
- 批准号:
03670037 - 财政年份:1991
- 资助金额:
$ 1.28万 - 项目类别:
Grant-in-Aid for General Scientific Research (C)
Studies on the Mechanism of Physiological Function of Hemoglobin by Using Artificial Mutants
利用人工突变体研究血红蛋白生理功能的机制
- 批准号:
01570045 - 财政年份:1989
- 资助金额:
$ 1.28万 - 项目类别:
Grant-in-Aid for General Scientific Research (C)
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