Determining Transient Nucleic Acid Structures by NMR
Current structure determination techniques for bio-macromolecules including X-ray crystallography, nuclear magnetic resonance spectroscopy (NMR) and cryo-electron microscopy, focus on obtaining snapshots of the most stable conformer in the free-energy lan
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Contents 1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 The R1ρ NMR Experiment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.1 Understanding Exchange Broadening and Suppression . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 Sample Preparation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4 Data Acquisition . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.1 Protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5 Data Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 Model Building . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6.1 Proving the Model . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7 Concluding Remarks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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Abstract Current structure determination techniques for bio-macromolecules including X-ray crystallography, nuclear magnetic resonance spectroscopy (NMR) and cryo-electron microscopy, focus on obtaining snapshots of the most stable conformer in the free-energy landscape. It is, however, imperative to fathom the conformations of other occupants of energy landscape as well that are transient in existence but are often the functionally active structures. NMR spectroscopy is a versatile technique that, in addition to providing static snapshots, has ability to measure various motional modes of a macromolecule. These motional modes connect distinct local minima in the free-energy landscape and help decipher additional conformational states that are accessible to a macromolecule. This book chapter gives a glimpse of the concept of existence of excited states for nucleic acids on free-energy landscape. Subsequently, this chapter focuses on using R1ρ relaxation dispersion NMR experiments to characterize the transient J. Chugh (*) Department of Chemistry, Indian Institute of Science Education and Research (IISER), Dr. Homi Bhabha Road, Pune 411008, Maharashtra, India e-mail: cjeet@iiserpu
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