RUI: Catalytic regulation of ribosome processing factors: Investigation of peripheral domain effects on the enzymatic capabilities of the DEAD-box protein Rok1p.

RUI：核糖体加工因子的催化调节：研究外周结构域对 DEAD-box 蛋白 Rok1p 酶促能力的影响。

• 批准号: 1515153
• 负责人: Ivelitza Garcia
• 金额: $ 17.4万
• 依托单位: Allegheny College
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2020-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1515153&HistoricalAwards=false
• 关键词: RUI; Catalytic; regulation; ribosome; processing

Title: RUI: Catalytic regulation of ribosome processing factors: Investigation of peripheral domain effects on the enzymatic capabilities of the DEAD-box protein Rok1p.Ribonucleic acids, RNAs, play a critical role in a variety of cellular processes. In order for RNAS to perform their specific functions in cells they often need to be processed by protein factors. Protein processing is generally associated with structural changes that are very important to the processing protein function. This research project will develop chemical-based methodologies to identify and characterize the protein structural elements that can regulate the activity and initial structural state of essential RNA processing proteins. The second objective of this research is to provide an effective training environment for students interested in RNA-protein research. Undergraduate students will perform all investigations, including the synthesis and purification of all RNAs and proteins utilized in this study. To cultivate a rich learning environment, comprehensive multi-approach mentoring and Just-in-Time teaching of experimental research will be used. This mentoring infrastructure is designed to advance problem solving and critical thinking skills in experimental design and analysis, and it has been successful in the past. Thus, this project will not only further the understanding of protein regulation, but also provide a strong foundation for students in the sciences. DEAD-box proteins constitute a large sub-class of putative unwindases essential in many RNA metabolic pathways. For example, Ribosomal RNA (rRNA) processing and assembly are highly regulated pathways requiring several DEAD-box proteins. This family of proteins couples cellular function to protein structural changes. In a subset of DEAD-box proteins, activity is mediated through additional peripheral elements/domains. The goal of this research is to understand the structural and thermodynamic nature of peripheral domain effects on the activity and conformational state of a model protein. Specifically, a DEAD-box protein model will be biochemically characterized and comparatively analyzed to evaluate the catalytic consequence of domain deletion variants. Equilibrium and kinetic methodologies will be developed to study the effects of initial conformations on nucleotide binding. Lastly, solvent-accessible and dynamic regions will be identified for the model system. Since the deregulation of the model protein and its human homolog has critical cellular consequences, the proposed research will further the understanding of (1) how DEAD-box proteins can bind early during processing and function in later events, and (2) how DEAD-box proteins and other RNA folding and processing accessory factors can utilize various conformations to regulate activity. Thus, the characterization of these states is important for any protein that utilizes ATP hydrolysis within a dynamic multi-factor complex.

职务名称：- 我知道核糖体加工因子的催化调节：周边结构域对DEAD盒蛋白Rok 1 p酶促能力影响的研究核糖核酸（RNA）在多种细胞过程中起着关键作用。为了使RNAS在细胞中发挥其特定功能，它们通常需要被蛋白质因子加工。 蛋白质加工通常与对加工蛋白质功能非常重要的结构变化相关。 该研究项目将开发基于化学的方法来识别和表征可以调节必需RNA加工蛋白的活性和初始结构状态的蛋白质结构元件。本研究的第二个目的是为对RNA-蛋白质研究感兴趣的学生提供一个有效的训练环境。 本科生将进行所有研究，包括本研究中使用的所有RNA和蛋白质的合成和纯化。为了培养一个丰富的学习环境，综合多方法指导和即时教学的实验研究将被使用。这种指导基础设施旨在提高实验设计和分析中的问题解决能力和批判性思维能力，并且在过去取得了成功。因此，该项目不仅将进一步了解蛋白质调控，而且还为学生提供了坚实的基础。死亡盒蛋白质构成了许多RNA代谢途径中必不可少的推定解旋酶的一大亚类。例如，核糖体RNA（rRNA）加工和组装是高度调节的途径，需要几种DEAD盒蛋白。该蛋白质家族将细胞功能与蛋白质结构变化偶联。在DEAD盒蛋白的一个子集中，活性通过额外的外周元件/结构域介导。本研究的目的是了解外周结构域对模型蛋白质的活性和构象状态的影响的结构和热力学性质。具体而言，将对DEAD盒蛋白模型进行生物化学表征和比较分析，以评估结构域缺失变体的催化后果。将开发平衡和动力学方法来研究初始构象对核苷酸结合的影响。最后，将确定模型系统的溶剂可及和动态区域。由于模型蛋白及其人类同源物的失调具有关键的细胞后果，因此拟议的研究将进一步了解（1）DEAD-box蛋白如何在加工过程中早期结合并在后期事件中发挥作用，以及（2）DEAD-box蛋白和其他RNA折叠和加工辅助因子如何利用各种构象来调节活性。因此，这些状态的表征对于在动态多因子复合物内利用ATP水解的任何蛋白质都是重要的。