Determination of Population-Weighted Dynamic Ensembles of RNA Using NMR Residual Dipolar Couplings.

Ensemble;Population-weighted;NMR;RDC;Chemistry;Science;Chemistry

Yang, ShanWalter, Nils G. ;

University of Michigan

关键词: Ensemble;    Population-weighted;    NMR;    RDC;    Chemistry;    Science;    Chemistry;   

Others  :  https://deepblue.lib.umich.edu/bitstream/handle/2027.42/108776/shanyang_1.pdf?sequence=1&isAllowed=y

瑞士|英语

来源: The Illinois Digital Environment for Access to Learning and Scholarship

【 摘 要 】

RNA undergoes large-scale conformational transitions in response to cellular cues, including changes in physiological conditions such as temperature and pH, recognition of proteins and ligand molecules, and RNA synthesis itself to perform a wide range of regulatory functions. A predictive understanding of how RNAs carry out their functions requires studies that go beyond static structure determination toward characterization of dynamic ensembles representing the broad RNA structure landscape. This thesis describes the development and application of Nuclear Magnetic Resonance techniques that rely on measurements of residual dipolar couplings (RDCs) for partially oriented RNAs in determination of dynamic ensembles. The ability to assess methods for ensemble determination hinges on the ability to compare the similarity between two ensembles. We have developed a new method that successfully captures both population overlap and structural similarity that relies on measuring population overlap as a function of the bin size used to bin ensemble distributions. Using this new method, we showed fundamental limitations in conventional approaches for measuring ensemble similarity and also find unexpected similarities between ensembles determined for HIV-1 TAR RNA with the use of NMR RDCs and computational molecular dynamics simulations. Using the new method for measuring ensemble similarity, we examined the accuracy with which ensembles can be determined with the use of RDCs under ideal conditions involving two domains and five perfectly orthogonal datasets. We found that the two factors resulting in uncertainty in determined dynamic ensembles of RNA are the experimental uncertainty of measured RDC as well as the ensemble size used to construct the ensemble. We developed an approach that takes into account these sources of uncertainty and applied it in the determination of ensembles for the bulge containing HIV-1 TAR in free state and ligand-bound states, for a TAR mutant with distinct dynamics, and for the HIV-1 ESS3 RNA containing an AC wobble base pair.