Folding of a Multidomain Ribosomal Protein

罗利9600866阐明氨基酸序列如何决定结构，即蛋白质折叠问题，是现代结构生物学的中心问题。该建议描述了一个综合的方法来研究蛋白质L9，一个多结构域核糖体蛋白的折叠。这种蛋白质形成了一个有趣的双叶结构，具有一个紧凑的N-末端结构域，通过一个长的溶剂暴露的α-螺旋连接到一个紧凑的C-末端结构域。N-末端结构域是一个稳定的折叠单位，代表了一类重要的层状片状螺旋结构的简单例子之一。这种类型的结构存在于其他核糖体蛋白和许多小RNA结合结构域中。相对而言，人们对稳定这种重要结构的相互作用或它如何折叠知之甚少。L9蛋白还提供了一个很好的模型系统来研究多结构域蛋白质的折叠。近年来，人们致力于阐明单结构域球状蛋白的折叠途径，但对多结构域球状蛋白的折叠知之甚少。 该蛋白质的折叠途径将使用动力学测量进行探索。将研究分离的结构域和完整的L9蛋白的平衡和动力学折叠转变。这些实验将表明结构域之间的相互作用是否影响蛋白质的折叠或稳定性。展开状态将通过核磁共振和圆二色性表征。肽片段的研究将用于解决稳定N-末端结构域的局部相互作用的作用。这些研究还将确定任何潜在的折叠起始位点。片段互补实验将用于探测三级相互作用在稳定新生二级结构中的作用。总之，这些实验将提供一个详细的描述的折叠途径和相互作用，稳定这个有趣的蛋白质。 在一所大的大学里教学的主要挑战之一是平衡本科教育和研究。许多学生感到他们“在洗牌中迷失了方向”，或者本科教育远远落后于研究和研究生教育。另一方面，现有的研究基础设施有可能为本科生提供独特的机会。该职业奖的教育方面包括一些旨在将本科生融入部门研究生活的举措。正在编制一门新的课程“生物化学的当前趋势”。本课程旨在向本科生介绍现代生物化学的研究领域，并围绕生物化学系系列研讨会展开。一个非正式的研讨会系列旨在向学生介绍化学家和生物化学家的许多职业选择也正在开发中。物理化学实验课作为本科生的核心课程之一，正在进行广泛的修订和重新定义。新的，更相关的实验已经推出，更多的互动教学方法正在使用。在研究生阶段，正在设计一个新的生物化学课程。这包括一个新的研讨会系列，这也构成了课程的基础“生物化学的当前趋势”：在化学系建立一个新的学位选择，设计一个新的蛋白质结构课程和编写跨部门的培训补助金。 阐明氨基酸序列如何决定结构，即蛋白质折叠问题，是现代结构生物学的中心问题。该建议描述了一个综合的方法来研究多结构域核糖体蛋白L9的折叠。人们对单结构域球状蛋白的折叠途径进行了大量的研究，但对多结构域球状蛋白的折叠却知之甚少。该分子的分离结构域也是感兴趣的。N-末端结构域是一个稳定的折叠单位，代表了核糖体蛋白和小RNA结合结构域中发现的重要结构类型的简单例子之一。 在大型大学教学的主要挑战之一是平衡本科教育与研究。许多学生感到他们“在洗牌中迷失了方向”，本科教育远远落后于研究生和研究生教育。该职业奖的教育方面包括一些旨在将本科生融入部门研究生活的举措。其中包括新课程的开发和一个新的本科生系列研讨会。在研究生阶段，正在设计一个新的生物化学课程。这包括在化学系建立一个新的学位选择。 ***

Raleigh 9600866 Elucidating how the amino acid sequence determines structure, the protein folding problem, is a central issue in modern structural biology. This proposal describes an integrated approach to studying the folding of the protein L9, a multidomain ribosomal protein. This protein forms an interesting bilobal structure with a compact N-terminal domain connected by a long solvent exposed a-helix to a compact C-terminal domain. The N-terminal domain is a stable folding unit and represents one of the simpler examples of an important class of layered sheet-helix structures. This type of structure is found in other ribosomal proteins and in many small RNA binding domains. Relatively little is known about the interactions which stabilize this important structure or how it folds. The L9 protein also offers a excellent model system to study the folding of multi-domain proteins. In recent years, considerable effort has been devoted to elucidating the folding pathways of single domain globular proteins but much less is known about the folding of multidomain proteins. The folding pathway of this protein will be explored using kinetic measurements. The equilibrium and kinetic folding transitions of the isolated domain and of the full L9 protein will be studied. These experiments will indicate if interactions between domains affect the folding or stability of the protein. The unfolded state will be characterized by Nuclear Magnetic Resonance and Circular Dichroism. Studies of peptide fragments will be used to address the role of local interactions in stabilizing the N-terminal domain. These studies will also identify any potential folding initiation sites. Fragment complementation experiments will be used to probe the role of tertiary interactions in stabilizing nascent secondary structure. Taken together, these experiments will provide a detailed description of the folding pathway and of the interactions which stabilize this interesting protein. One of the major challenges of teaching at a lar ge university is balancing undergraduate education with research. Many of the students feel that they are "lost in the shuffle" or that undergraduate education is ranked a distant third behind research and graduate education. On the other hand, the available research infrastructure has the potential to allow undergraduates unique opportunities. The educational aspects of this Career Award include a number of initiatives designed to integrate undergraduate students into the research life of the department. A new course "Current Trends in Biological Chemistry" is being developed. This course is designed to introduce undergraduates to research areas in modern biological chemistry and is built around the departmental seminar series in biological chemistry. An informal seminar series designed to introduce students to the many career options available to chemists and biochemists is also being developed. One of the core undergraduate courses, physical chemistry laboratory is being extensively revised and redefined. New, more relevant experiments have been introduced and more interactive methods of teaching are being used. At the graduate level, a new program in biological chemistry is being designed. This includes a new seminar series, which also forms the basis of the course "Current Trends in Biological Chemistry': the establishment of a new degree option in the Department of Chemistry, the design of a new course on Protein Structure and the writing of an interdepartmental training grant. %%% Elucidating how the amino acid sequence determines structure, the protein folding problem, is a central issue in modern structural biology. This proposal describes an integrated approach to studying the folding of the multidomain ribosomal protein L9. Considerable effort has been devoted to elucidating the folding pathways of single domain globular proteins but much less is known about the folding of multidomain proteins. The isolated domains of this molecule are also of interest. The N-terminal domain is a st able folding unit and represents one of the simpler examples of an important type of structure found in ribosomal proteins and in small RNA binding domains. One of the major challenges of teaching at a large university is balancing undergraduate education with research. Many of the students feel that they are "lost in the shuffle" and that undergraduate education is ranked a distant third behind research and graduate education. The educational aspects of this Career Award include a number of initiatives designed to integrate undergraduate students into the research life of the department. These include the development of new courses and a new undergraduate seminar series. At the graduate level, a new program in biological chemistry is being designed. This includes the establishment of a new degree option in the Department of Chemistry. ***