CAREER: NMR Insights into the Influence of Dynamics on SH3 Domain Mediated Protein Interactions

职业：NMR 洞察动力学对 SH3 结构域介导的蛋白质相互作用的影响

• 批准号: 0347100
• 负责人: Ranajeet Ghose
• 金额: $ 70.36万
• 依托单位: CUNY City College
• 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: 2004-03-01 至 2009-02-28
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0347100&HistoricalAwards=false
• 关键词: CAREER; NMR; Insights; into; Influence

The objective of this project is to develop novel techniques to characterize Nuclear Magnetic Resonance (NMR) relaxation of proteins in the solution state. The development and implementation of these techniques will be facilitated by (i) advances in theoretical methods for experimental design and analysis, (ii) major improvements in commercially available instrumentation, and (iii) the development of cost-effective methods for the bacterial expression and purification of labeled proteins. These methods will be applied to study protein dynamics over a wide range of timescales, with special emphasis on the characterization of protein sidechain dynamics since sidechains play a central role in defining protein-ligand interactions and hence physiological function. These methods will alleviate many of the deficiencies of current experiments by (i) providing information on the details of local dynamics on the fast, ps-ns timescale, (ii) allowing complete characterization of slow, correlated motions on the microsecond-ms timescale, and (iii) being applicable to larger proteins and protein complexes. The PI will utilize molecular dynamics simulations in conjunction with theoretical modeling to identify specific dynamic modes that contribute to the measured relaxation rates. This combination of experimental and computational techniques will allow the identification of key functional modes and the changes thereof on protein interactions. The PI will apply this methodology to elucidate the dynamic determinants of the intra- and inter-molecular interactions involving two cytosolic components of the NADPH oxidase complex - p47phox and p67phox. A very important structural module, the SH3 domain plays a central role in defining protein interactions in these cases. Thus, information gleaned from these studies will provide a self-consistent, space-time view of SH3 domain-mediated protein interactions.The PI will place special emphasis on recruiting students from various backgrounds and at various levels of sophistication, undergraduate and graduate, into his research program. He will redesign undergraduate courses to provide students with the knowledge of cutting edge biophysical techniques and develop a new course on NMR spectroscopy targeted towards advanced undergraduates and graduate students. This course will introduce students to both the theoretical as well as the applied aspects of modern biomolecular NMR spectroscopy both through lectures as well as a series of "hands-on" workshops. The PI currently participates, and will continue to do so, in science outreach programs such as ACS-SEED. This program is designed to facilitate underprivileged high-school students to gain research experience over the summer. CUNY City College has a large minority population. Thus the research and teaching components of this project will encourage those sections of society underrepresented in the sciences to pursue scientific careers both in academia and in industry. Facilitating minority participation in the sciences through teaching, mentoring, career counseling and participating in outreach programs for disadvantaged students is a key goal of this CAREER project.

该项目的目的是开发新技术来表征溶液状态下蛋白质的核磁共振（NMR）松弛。这些技术的开发和实施将通过（i）实验设计和分析的理论方法的进步，（ii）商业上可用的仪器的重大改进，以及（iii）开发具有成本效益的细菌表达和纯化标记蛋白质的方法。这些方法将应用于在广泛的时间尺度上研究蛋白质动力学，特别强调了蛋白质侧链动力学的表征，因为SideChains在定义蛋白质 - 配体相互作用及其生理功能方面起着核心作用。这些方法将通过（i）提供有关快速，PS-NS时间尺度上局部动力学的详细信息的信息来减轻当前实验的许多缺陷，（ii）允许对微秒MS时尺度上缓慢的相关运动完全表征，以及（III）适用于较大的蛋白质和蛋白质复合物。 PI将利用分子动力学模拟与理论建模一起确定有助于测得的松弛速率的特定动力模式。实验技术和计算技术的这种组合将允许识别关键功能模式及其在蛋白质相互作用上的变化。 PI将应用这种方法来阐明涉及NADPH氧化酶复合物的两个胞质成分-P47Phox和P67Phox的胞质成分的动态决定因素。 SH3结构域是一个非常重要的结构模块，在这些情况下定义蛋白质相互作用方面起着核心作用。因此，从这些研究中收集的信息将为SH3域介导的蛋白质相互作用提供自洽的时空视图。PI将特别强调来自各种背景的招募学生，以及各种成熟，本科生和毕业生的级别。他将重新设计本科课程，以为学生提供尖端生物物理技术的知识，并开发针对高级本科生和研究生的NMR光谱学课程。本课程将通过讲座以及一系列“动手”研讨会介绍学生的理论和现代生物分子NMR光谱的应用方面。 PI目前参加，并将继续参加ACS-SEED等科学外展计划。该计划旨在促进贫困的高中生，以在夏季获得研究经验。 Cuny City College的少数人口。因此，该项目的研究和教学组成部分将鼓励科学中人数不足的社会部分从事学术界和工业的科学职业。通过教学，指导，职业咨询和参加弱势学生的外展计划，促进少数群体参与科学是这个职业项目的关键目标。