Modeling Green Fluorescent Protein and GFP-like Proteins

绿色荧光蛋白和 GFP 样蛋白建模

• 批准号: 6503161
• 负责人: MARC ZIMMER
• 金额: $ 12.18万
• 依托单位: CONNECTICUT COLLEGE
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-04-01 至 2005-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6503161
• 关键词: bioluminescence; chromophore; computer simulation; conformation; fluorescence; fluorescence resonance energy transfer; green fluorescent proteins; hydrogen bond; molecular biology information system; molecular dynamics; mutant; point mutation; protein folding; protein structure; protein structure function; thermodynamics

DESCRIPTION (provided by applicant): In the last ten years, green fluorescent protein (GFP) has changed from a nearly unknown protein to a frequently used tool in molecular biology, medicine and cell biology. The most common uses of GFP are as a fusion tag to visualize dynamic cellular events and to monitor protein localization, and as a report gene to monitor gene expression. GFP is commonly used in medicinal research. For example, GFP has been used as a marker for tumor cells, to illuminate tumor progression and allow for detection of metastases down to the single cell level.Computational methods will be used to investigate the following phenomena: * Yellow Fluorescent Proteins are GFP mutants that have lower fluorescent quantum yields than GFP's. The conformational freedom of the chromophore might be responsible for the lower quantum yields. * Mature GFP; i.e. fully fluorescent GFP, is most efficiently formed at temperatures well below 37xC. Computational methods will be used to search for mutants that mature more efficiently at higher temperatures and to establish the mechanism of their action. * GFP has an 11-sheet beta-barrel structure. Water and halide ions can enter the barrel. The pathways into the barrel can be found with activated molecular dynamics. The resultant information could be used to design more specific halide ion sensors. * DsRed is a red-emitting fluorescent protein from discosoma that is commercially available and has been studied in some detail. Two of the largest disadvantages of DsRed are its slow chromophore formation and the fact that DsRed is an obligate tetramer. These drawbacks could be overcome by computationally designing DsRed mutants that do not form multimeric structures, or by designing GFP mutants that can form chromophores with extended conjugation such as that found in DsRed. * The autocatalytic chromophore forming reaction of the purple GFP-like protein from Anemonia sulcata is significantly different to that observed in GFP. Computational methods will be used to investigate structural aspects of the chromophore formation in the purple GFP-Iike protein.

描述（由申请人提供）：在过去十年中，绿色荧光蛋白（GFP）已从一种几乎未知的蛋白质转变为分子生物学、医学和细胞生物学中常用的工具。GFP最常见的用途是作为融合标签来可视化动态细胞事件并监测蛋白质定位，以及作为报告基因来监测基因表达。GFP通常用于医学研究。例如，GFP作为肿瘤细胞的标记物，可以在单个细胞的水平上观察肿瘤的进展，并检测转移灶。计算方法将用于研究以下现象：* 黄色荧光蛋白是荧光量子产率低于GFP的GFP突变体。发色团的构象自由度可能是导致较低量子产率的原因。 * 成熟GFP（即完全荧光GFP）在远低于37 ° C的温度下最有效地形成。计算方法将用于寻找在较高温度下更有效地成熟的突变体，并建立其作用机制。* GFP具有11片β桶结构。水和卤素离子可以进入桶。进入枪管的路径可以用活化的分子动力学来找到。所得信息可用于设计更特异的卤素离子传感器。 * DsRed是一种来自discosoma的发红光的荧光蛋白，其可商购获得，并且已经进行了一些详细的研究。DsRed的两个最大缺点是其缓慢的发色团形成和DsRed是专性四聚体的事实。这些缺点可以通过计算设计不形成多聚体结构的DsRed突变体，或者通过设计可以形成具有延长缀合的发色团的GFP突变体（例如在DsRed中发现的）来克服。 * 从Anemonia sulcata的紫色GFP样蛋白的自催化发色团形成反应是显着不同的，在GFP中观察到的。计算方法将被用来研究紫色GFP样蛋白质中发色团形成的结构方面。