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

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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.