Development of 13C Nuclear Magnetic Resonance Methods for Studying theStructural Dynamics of Nucleic Acids in Solution.

Nuclear Magnetic Resonance;Nucleic Acid;Relaxation;HIV;Structural Dynamics;Chemical Exchange;Chemistry;Science;Chemistry

Hansen, Alexandar LouisRamamoorthy, Ayyalusamy ;

University of Michigan

关键词: Nuclear Magnetic Resonance;    Nucleic Acid;    Relaxation;    HIV;    Structural Dynamics;    Chemical Exchange;    Chemistry;    Science;    Chemistry;   

Others  :  https://deepblue.lib.umich.edu/bitstream/handle/2027.42/62347/viochem_1.pdf?sequence=1&isAllowed=y

瑞士|英语

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

【 摘 要 】

An understanding of both structure and dynamics is essential to the full characterizationof any biomolecule, but is especially relevant with respect to RNA for which dynamics isused in myriad ways to achieve functional complexity that would otherwise beinaccessible based on its rigid framework composed of only four chemically similarnucleotides. Due to experimental difficulties in resolving the plethora of motional modesthat exist in RNA, their dynamical properties remain poorly understood. Solutionnuclear magnetic resonance (NMR) is one of the most powerful tools for thecharacterization of structural dynamics, as it provides atomic level detail on a variety oftimescales, from picoseconds to seconds. Spin relaxation measurements can in principleprovide information at sub‐nanosecond timescales, providing that internal motions arenot correlated to overall molecular tumbling. Residual dipolar couplings and residualchemical shift anisotropies (RCSAs) report on the average global RNA structure andprovide insight into sub‐millisecond motions. Finally, chemical exchange measurements can provide quantitative kinetic information on the micro‐to‐millisecond timescale.Unfortunately, many of the techniques commonly used for studies of RNA are limited tonitrogen resonances, which are not frequently observable in functionally relevant, noncanonicalregions of RNA. In addition, target RNAs are relatively small, typically lessthan 30 nucleotides or 10,000 molecular weight. In this thesis, I develop the muchneeded NMR methods which can target the carbon nuclei of RNA and in systems up to150 nucleotides. A combination of new spin relaxation, RCSA, and chemical exchangetechniques are developed to probe site specific motions over the picosecond tomillisecond time regime and provide important insight into some of the fundamentalproperties of RNA. Spin relaxation revealed a surprisingly complex dynamicallandscape for the relatively simple transactivation response element from HIV‐1 RNAwhere intriguing entropy compensation occurs upon ligand binding in the bulge region,with order parameters of 0.2‐0.3, as global domain motions are suppressed. New,selective R1ρ dispersion experiments detected previously unobservable chemicalexchange in functionally important regions of the bacterial ribosomal A‐site RNA, with atimescale of 320 μs, and the modified base in a 1,N6‐ethenoadenine ‐ damaged DNA.