Photodynamics and Maturation of Coral Fluorescent Proteins
珊瑚荧光蛋白的光动力学和成熟
基本信息
- 批准号:0417290
- 负责人:
- 金额:$ 52.5万
- 依托单位:
- 依托单位国家:美国
- 项目类别:Continuing Grant
- 财政年份:2004
- 资助国家:美国
- 起止时间:2004-08-01 至 2007-07-31
- 项目状态:已结题
- 来源:
- 关键词:
项目摘要
Time resolved crystallographic and kinetic studies will be used to investigate the unusual photocycle of amFP595, the "kindling fluorescent protein", a GFP-like chromoprotein isolated from reef organisms. The reversible light-induced transition from the dark nonfluorescent state to the fluorescent state and its subsequent slow relaxation in the dark will be analyzed at very high resolution using optical pumping and synchrotron radiation in various temperature regimes. The research will determine the nature of the structural transitions of the chromophore within the protein matrix, the nature of intermediate states and determine how the protein matrix controls both the ground state conformation and the excited state decay pathways of the chromophore. Mutagenesis combined with Arrhenius plot analysis will be used to study the energetic contributions of selected side chains to conformational stability, dark relaxation rates and reversibility of the fluorescence transition. These studies will be combined with limited crystallographic analysis and mutagenesis of two very closely related fluorescent proteins, blue fluorescent amFP486 and yellow fluorescent zFP538, in order to elucidate the structural basis for the differences in maturation behavior. It is postulated that the same chemical intermediates are formed in each protein, but suffer different final fates, due to the subtle effects of a small number of protein side chains close to the chromophore. While the research is directly relevant to the photodynamic processes involved in human vision, a potential practical outcome of the project is the development of a transient fluorescent tag for use in studies of material transport within living cells.Broader Impacts: The results of the proposed studies are expected to have broad impact as they will be relevant to the photophysics of many biological molecules, such as those involved in vision and photosynthesis. The results may also provide insight into the transient light/dark switching observed for small molecule and protein fluorophores used by many researchers. In the long term, the understanding gained from this work can be used to improve the properties of fluorescent proteins for use as tools for the study of the development and dynamic behavior of living creatures on both the subcellular and whole organism levels. The methodology will include a great variety of techniques drawn from physical chemistry and molecular biology and will thus provide excellent multidisciplinary training for graduate students and postdoctoral researchers. Due to their visual appeal, ease of production and isolation combined with exceptional physical stability, fluorescent proteins form an outstanding teaching resource for laboratory courses at the high school and university levels.
时间分辨结晶学和动力学研究将被用来研究amFP595的异常光周期,amFP595是从珊瑚礁生物中分离出来的一种类似GFP的色素蛋白,是一种“点燃荧光蛋白”。利用光泵浦和同步辐射,在不同的温度范围内,我们将以很高的分辨率分析由暗非荧光态到荧光态的可逆光诱导跃迁及其在黑暗中的缓慢弛豫。这项研究将确定蛋白质基质中发色团结构转变的性质,中间态的性质,并确定蛋白质基质如何控制发色团的基态构象和激发态衰变途径。突变结合Arrhenius图分析将被用来研究所选侧链对构象稳定性、暗松弛速率和荧光跃迁的可逆性的能量贡献。这些研究将结合有限的结晶学分析和两种非常相似的荧光蛋白-蓝色荧光amFP486和黄色荧光zFP538的突变,以阐明成熟行为差异的结构基础。据推测,每种蛋白质中都形成了相同的化学中间体,但由于靠近发色团的少量蛋白质侧链的微妙影响,最终的命运不同。虽然这项研究与人类视觉中涉及的光动力学过程直接相关,但该项目的一个潜在实用成果是开发出一种用于研究活细胞内物质运输的瞬时荧光标签。广泛影响:拟议的研究结果预计将产生广泛的影响,因为它们将与许多生物分子的光物理相关,例如涉及视觉和光合作用的分子。这些结果也可能为许多研究人员观察到的小分子和蛋白质荧光团的瞬时明/暗切换提供洞察力。从长远来看,从这项工作中获得的理解可以用来改进荧光蛋白的性质,用于在亚细胞和整个生物体水平上研究生物的发育和动态行为。该方法将包括从物理化学和分子生物学中提取的各种技术,因此将为研究生和博士后研究人员提供出色的多学科培训。由于它们的视觉吸引力、易于生产和分离以及特殊的物理稳定性,荧光蛋白成为高中和大学水平实验室课程的优秀教学资源。
项目成果
期刊论文数量(0)
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Stephen Remington其他文献
A new vision for employment support
- DOI:
10.1016/j.ics.2005.05.117 - 发表时间:
2005-09-01 - 期刊:
- 影响因子:
- 作者:
Stephen Remington - 通讯作者:
Stephen Remington
Stephen Remington的其他文献
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{{ truncateString('Stephen Remington', 18)}}的其他基金
Excited State Proton Transfer in Fluorescent Proteins
荧光蛋白中的激发态质子转移
- 批准号:
1021374 - 财政年份:2010
- 资助金额:
$ 52.5万 - 项目类别:
Continuing Grant
Structure-Function in Red and Yellow Fluorescent Proteins
红色和黄色荧光蛋白的结构-功能
- 批准号:
0111053 - 财政年份:2001
- 资助金额:
$ 52.5万 - 项目类别:
Continuing Grant
Enzyme Structure-Function Relationships
酶结构与功能的关系
- 批准号:
9728162 - 财政年份:1998
- 资助金额:
$ 52.5万 - 项目类别:
Continuing Grant
Enzyme Structure-Function Relationships
酶结构与功能的关系
- 批准号:
9418479 - 财政年份:1995
- 资助金额:
$ 52.5万 - 项目类别:
Continuing Grant
Enzyme Structure-Function Relationships
酶结构与功能的关系
- 批准号:
9118302 - 财政年份:1992
- 资助金额:
$ 52.5万 - 项目类别:
Continuing Grant
Enzyme Structure-Function Relationships
酶结构与功能的关系
- 批准号:
8817438 - 财政年份:1989
- 资助金额:
$ 52.5万 - 项目类别:
Continuing Grant
X-ray Structure of Macromolecular Complexes
大分子配合物的X射线结构
- 批准号:
8517785 - 财政年份:1986
- 资助金额:
$ 52.5万 - 项目类别:
Continuing Grant
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