Single Molecular Biochemistry of G protein signaling system in photoreceptor cell
感光细胞G蛋白信号系统的单分子生物化学
基本信息
- 批准号:15201027
- 负责人:
- 金额:$ 28.7万
- 依托单位:
- 依托单位国家:日本
- 项目类别:Grant-in-Aid for Scientific Research (A)
- 财政年份:2003
- 资助国家:日本
- 起止时间:2003 至 2005
- 项目状态:已结题
- 来源:
- 关键词:
项目摘要
We have tried to construct a foundation of single molecular biochemistry of G protein mediated signal transduction system in vertebrate rod photoreceptor cells. By using a single fluorescent molecule imaging technique, we have succeeded to observe single molecular behavior of three key components of vertebrate phototransduction, i.e. a typical GPCR rhodopsin, G protein transducin, and its target enzyme cGMP-PDE. We have employed near far-red fluorescent-labeled antibody Fab' fragment or direct conjugation of these dyes for labeling these proteins. Rhodopsin was proved with Cy7-labeled Fab' fragment of monoclonal antibody (1D4) against the C-terminus of rhodopsin. The single molecular behavior of rhodopsin on the disc membrane was observed at video rate and 70 nm/pixel in spatial resolution. It was found that the rhodopsin does not form static paracrystalline as reported recently on the basis of AFM observation. Rhodopsin undergoes anomalous diffusion changing diffusion rate frequently. FRET experiments revealed that rhodopsins form dimer or a cluster of short lifetime. Artificially dimerized rhodopsin by the use of IgG also shows slow diffusion rate. The formation of cluster of low diffusion rate is apparently disadvantageous for rapid amplification in the phototransduction. Thus, it was suggested paradoxically that the cluster formation has an important meaning for photo-transduction. In addition, I succeeded in labeling transducin by Cy7 maintaining its activity to bind rhodopsin and PDE. Single molecular behavior of transducin was totally different from that of rhodopsin. Single molecular biochemistry of phototransduction has started to evolve.
本研究试图建立G蛋白介导的视杆细胞信号转导系统的单分子生物学基础。利用单荧光分子成像技术,我们成功地观察了脊椎动物光转导的三个关键组分,即典型的GPCR视紫红质、G蛋白转导素及其靶酶cGMP-PDE的单分子行为。我们采用近远红荧光标记抗体Fab'片段或直接偶联这些染料来标记这些蛋白。用Cy 7标记的抗视紫红质C端的单克隆抗体(1D 4)的Fab'片段证明了视紫红质。在视频速率和70 nm/像素的空间分辨率下观察了视紫红质在盘膜上的单分子行为。通过AFM观察发现,视紫红质不像最近报道的那样形成静态的次晶。视紫红质经历异常扩散,经常改变扩散速率。FRET实验显示视紫红质形成二聚体或短寿命的簇。通过使用IgG的牺牲性二聚视紫红质也显示出缓慢的扩散速率。低扩散速率团簇的形成明显不利于光转导过程中的快速放大。因此,这是矛盾的建议,集团的形成有一个重要的意义,光转导。此外,我成功地标记转导蛋白的Cy 7保持其活性,结合视紫红质和PDE。转导素的单分子行为与视紫红质完全不同。光传导的单分子生物化学已经开始发展。
项目成果
期刊论文数量(4)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Ahamed, A.: "Isolation of chromosaponin I-specific antibody by affinity chromatography"Biochem. Biophys. Res. Commun.. 302. 587-592 (2003)
Ahamed,A.:“通过亲和层析分离铬皂苷 I 特异性抗体”Biochem。
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- 影响因子:0
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- 通讯作者:
Liu, H.: "Active Transducin a Subunit Carries PDE6 to Detergent-Resistant Membranes in Rod Photoreceptor Outer Segments"Biochem.Biophys.Res.Commun.. 303. 19-23 (2003)
Liu, H.:“活性转导蛋白亚基将 PDE6 携带到视杆光感受器外节中的去垢剂抗性膜”Biochem.Biophys.Res.Commun.. 303. 19-23 (2003)
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HAYASHI Fumio其他文献
越南北部齿蛉及鱼蛉种类纪要(广翅目:齿蛉科)(英文)
- DOI:
- 发表时间:
2012 - 期刊:
- 影响因子:0
- 作者:
LETARDI Agostino;HAYASHI Fumio;刘星月;LETARDI Agostino 1,HAYASHI Fumio 2,LIU Xingyue 3① - 通讯作者:
LETARDI Agostino 1,HAYASHI Fumio 2,LIU Xingyue 3①
HAYASHI Fumio的其他文献
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{{ truncateString('HAYASHI Fumio', 18)}}的其他基金
Elucidation of the mechanism of thickening and fibrosis of ligamenta flava
阐明黄韧带增厚和纤维化的机制
- 批准号:
18K16663 - 财政年份:2018
- 资助金额:
$ 28.7万 - 项目类别:
Grant-in-Aid for Early-Career Scientists
Evolution of male insect genitalia: Sperm removal in Euphaeidae damselflies
雄性昆虫生殖器的进化:Euphaeidae豆娘的精子去除
- 批准号:
16K07486 - 财政年份:2016
- 资助金额:
$ 28.7万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Examination of the function of insect male genitalia using a fine fluorescent beads
使用细荧光珠检查昆虫雄性生殖器的功能
- 批准号:
24657063 - 财政年份:2012
- 资助金额:
$ 28.7万 - 项目类别:
Grant-in-Aid for Challenging Exploratory Research
In vitro reconstitution of Type III secretion system - basic and applied research -
III型分泌系统的体外重建-基础与应用研究-
- 批准号:
23790468 - 财政年份:2011
- 资助金额:
$ 28.7万 - 项目类别:
Grant-in-Aid for Young Scientists (B)
A Dynamic General Equilibrium Analysis of the Prewar Japanese Economy
战前日本经济的动态一般均衡分析
- 批准号:
22530334 - 财政年份:2010
- 资助金额:
$ 28.7万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Asymmetric male genitalia in calopterygid damselflies : their function and evolutionary patterns
豆娘的不对称雄性生殖器:它们的功能和进化模式
- 批准号:
21570099 - 财政年份:2009
- 资助金额:
$ 28.7万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Ecological mechanisms to promote or suppress interspecific hybridization
促进或抑制种间杂交的生态机制
- 批准号:
19570090 - 财政年份:2007
- 资助金额:
$ 28.7万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Single Molecular Biochemistry of The Operating Mechanism of Photoreoeptor G protein and its Trafficking Mechanism.
光感受器G蛋白运作机制及其运输机制的单分子生物化学。
- 批准号:
18310088 - 财政年份:2006
- 资助金额:
$ 28.7万 - 项目类别:
Grant-in-Aid for Scientific Research (B)
Mechanisms of sperm competition revealed by sperm viability assessment using live/dead dual fluorescence
使用活/死双荧光评估精子活力揭示精子竞争机制
- 批准号:
15570019 - 财政年份:2003
- 资助金额:
$ 28.7万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
Morphologically divergent sperm in Odonata: phylogenetic patterns and relations with sperm competition
蜻蜓目精子形态分化:系统发育模式及其与精子竞争的关系
- 批准号:
13640632 - 财政年份:2001
- 资助金额:
$ 28.7万 - 项目类别:
Grant-in-Aid for Scientific Research (C)
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