NMR Determination of Biomolecular Structural Dynamics Using Residual Dipolar Couplings Measured in Multiple Alignment Media

Johns Hopkins University, Baltimore MD

Investigators

Abstract

The goal of this project is the development of residual dipolar coupling (RDC)-based NMR methods for the determination of biomolecular structure and dynamics. This approach will be based on the measurement of multiple sets of RDCs under conditions of different anisotropic solvent environments. Novel methods for modulating the nature of the anisotropic solvent environment will be developed in order to improve the efficiency of data acquisition. In particular, these studies will focus on the exploitation of the interference observed between aligning forces for composite, mechanically ordered media such as bacteriophage Pf1 particles embedded within a strained polyacrylamide gel matrix. The DIDC approach to the interpretation of RDCs, demonstrated by the PI for backbone amide N-H bonds, will be extended to the determination of side-chain conformation and dynamics. Furthermore, these multi-alignment RDC-based approaches will be adapted for application to the protein Neisseria meningitidis heme oxygenase, for which dynamics have been implicated as necessary for its function. These studies will serve as a test case for the ability to characterize in detail the structural dynamics of a moderately sized protein (24 kDa) using RDC-based methods. It is anticipated that these studies will lay a foundation for future NMR investigations and provide novel insights into the relationship between protein structural dynamics and function. This research will provide powerful tools for the characterization of mobility and structure of biomolecular systems and thus will have a far reaching impact on the ability to characterize the mechanical aspects of biomolecular function at atomic resolution. This project will also broadly impact undergraduate teaching and research in biomolecular NMR spectroscopy, a technique often inaccessible to undergraduates due to the expense and geographical concentration of high field NMR instrumentation. In order to increase undergraduate research opportunities in biomolecular NMR, one undergraduate summer research position has been established, primarily intended for students outside of the Johns Hopkins community. Sustained efforts are being made to stimulate collaboration and facilitate training of local biophysical and biochemical researchers such that they can employ state of the art biomolecular NMR spectroscopic techniques to advantage