CAREER: High-resolution Structural Studies of Paramagnetic Proteins by Multidimensional Solid-state NMR Spectroscopy

职业：通过多维固态核磁共振波谱对顺磁性蛋白质进行高分辨率结构研究

• 批准号: 0745754
• 负责人: Christopher Jaroniec
• 金额: $ 69.82万
• 依托单位: Ohio State University Research Foundation -DO NOT USE
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-03-15 至 2014-02-28
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0745754&HistoricalAwards=false
• 关键词: CAREER; resolution; Structural; Studies; Paramagnetic

Multidimensional magic-angle spinning solid-state nuclear magnetic resonance (SSNMR) methods can now be applied to uniformly 13C,15N enriched proteins, enabling detailed atomic-level studies and even the de novo determination of three-dimensional molecular structure. However, one of the outstanding problems in the field, which impedes these efforts, is related to the paucity of long range (i.e., greater than 5 A) distance restraints that can be obtained using the existing SSNMR techniques. In this CAREER project, proteins modified with covalently-bound nitroxide spin labels and paramagnetic metal ions, will be investigated using solid-state NMR, with the main goal of deriving structural restraints on length scales inaccessible to traditional SSNMR methods (up to ~20 A). The specific research objectives are: (i) to explore in detail the paramagnetic relaxation enhancement (PRE) of nuclear spins in the solid phase caused by the dipole-dipole coupling to unpaired electrons, and (ii) to establish novel PRE-based SSNMR methods and apply them to long-range distance measurements, studies of protein-protein interactions, and systematic simplification of complex NMR spectra in challenging biological solids. The new PRE-based SSNMR methods can be adapted to a wide variety of problems involving molecular structure and interactions in non-crystalline biological systems.The main objective of the education-outreach activities is to directly impact students in the early stage of their educational experience, particularly students from groups traditionally underrepresented in science, technology, engineering and mathematics (STEM) by introducing them to current, interdisciplinary science at the interface of chemistry, biology, and physics and encouraging them to think of science in a broad, discovery-based manner, with the ultimate goal of increasing student participation, retention and graduation rates in the STEM fields. The specific education-outreach activities include: (i) a summer research internship program with a team of undergraduate and local high-school students, (ii) an undergraduate laboratory module, which will introduce modern NMR spectroscopy to second-year undergraduates and will interface with other research-oriented laboratory modules, and (iii) a seminar course for a diverse audience of first-year undergraduate students directly related to the PI's research interests in protein structure and function. This project is jointly supported by the Molecular Biophysics Program in the Division of Molecular and Cellular Biosciences in the Directorate for Biological Sciences and the Experimental Physical Chemistry Program in the Division of Chemistry in the Mathematical and Physical Sciences Directorate.

多维魔角旋转固态核磁共振（SSNMR）方法现在可以应用于均匀富集13 C，15 N的蛋白质，从而能够进行详细的原子水平研究，甚至从头确定三维分子结构。然而，阻碍这些努力的该领域的突出问题之一与远距离（即，大于5A）的距离约束，其可以使用现有的SSNMR技术获得。在这个CAREER项目中，将使用固态NMR研究用共价结合的氮氧自旋标记和顺磁性金属离子修饰的蛋白质，主要目标是推导传统SSNMR方法无法达到的长度尺度上的结构限制（高达~20 A）。具体的研究目标是：（i）详细探索由偶极-偶极耦合到未配对电子引起的固相核自旋的顺磁弛豫增强（PRE），以及（ii）建立新的基于PRE的SSNMR方法，并将其应用于长距离测量，蛋白质-蛋白质相互作用的研究，以及具有挑战性的生物固体中复杂NMR谱的系统简化。新的基于PRE的SSNMR方法可以适用于涉及非晶体生物系统中分子结构和相互作用的各种问题。教育推广活动的主要目标是直接影响学生在教育经历的早期阶段，特别是来自传统上在科学，技术，工程和数学（STEM）方面代表性不足的群体的学生，通过向他们介绍当前，在化学，生物学和物理学的接口跨学科科学，并鼓励他们认为科学在广泛的，发现为基础的方式，以提高学生的参与，保留和毕业率在STEM领域的最终目标。具体的教育推广活动包括：（i）一个由本科生和当地高中生组成的暑期研究实习计划，（ii）一个本科生实验室模块，将向二年级本科生介绍现代核磁共振光谱学，并将与其他研究型实验室模块相结合，及（iii）为不同的本科一年级学生举办的研讨会课程，与研究主任对蛋白质结构和功能的研究兴趣直接相关。该项目由生物科学理事会分子和细胞生物科学部的分子生物物理学计划和数学和物理科学理事会化学部的实验物理化学计划共同支持。