We propose to study how dynamics of proteins and nucleic acids contribute to molecular recognition by probing dynamic processes over a wide range of time scales by means of solution and solid state NMR spectroscopy.
We aim to integrate these two complementary experimental approaches with each other and exploit new computational methods to gain a quantitative understanding of the role that protein and nucleic acid dynamics play in molecular recognition by providing a description of the potential energy surfaces upon which motion occurs. Through our joint approach, it will be possible to cover the wide range of time scale and motional characteristics that occurs in biological molecules as they bind each other, and therefore provide a comprehensive description of the dynamics involved in binding. We will study native DNA and two protein-RNA complexes with outstanding biological importance where motion has been shown to play an essential functional role. We have defined questions and hypotheses concerning the role of motion in recognition that we are poised to test through the experimental program we propose.
We aim to fulfill this proposal by focusing on 2 specific aims using a common experimental approach developed in our previous studies. Specifically, we aim to use solution NMR and solid state NMR to study how motion affects DNA methylation.
This specific aim i s essentially a continuation of earlier solid state NMR studies to which we now add resolution NMR relaxation techniques.
As specific aim 2 we will compare two different examples of RNA recognition by RRM proteins. In one case, motion is quenched upon binding to allow very high specificity by human U1A; in the other case, motion is instead generated upon binding by CstF-64 to allow diffuse specificity and transient interactions to be established.

Agency
National Institute of Health (NIH)
Institute
National Institute of Biomedical Imaging and Bioengineering (NIBIB)
Type
Research Project (R01)
Project #
5R01EB003152-07
Application #
7096664
Study Section
Biophysical Chemistry Study Section (BBCB)
Program Officer
Mclaughlin, Alan Charles
Project Start
1999-02-01
Project End
2008-06-30
Budget Start
2006-07-01
Budget End
2007-06-30
Support Year
7
Fiscal Year
2006
Total Cost
$456,289
Indirect Cost
Name
University of Washington
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
605799469
City
Seattle
State
WA
Country
United States
Zip Code
98195
Huang, Wei; Bardaro Jr, Michael F; Varani, Gabriele et al. (2012) Preparation of RNA samples with narrow line widths for solid state NMR investigations. J Magn Reson 223:51-4
Emani, Prashant S; Olsen, Gregory L; Varani, Gabriele et al. (2012) Correction to ""Theory of nonrigid rotational motion applied to NMR relaxation in RNA"". J Phys Chem A 116:7253-60
Huang, Wei; Varani, Gabriele; Drobny, Gary P (2011) Interactions of protein side chains with RNA defined with REDOR solid state NMR. J Biomol NMR 51:347-56
Emani, Prashant S; Olsen, Gregory L; Varani, Gabriele et al. (2011) Theory of nonrigid rotational motion applied to NMR relaxation in RNA. J Phys Chem A 115:12055-69
Olsen, Greg L; Bardaro Jr, Michael F; Echodu, Dorothy C et al. (2010) Intermediate rate atomic trajectories of RNA by solid-state NMR spectroscopy. J Am Chem Soc 132:303-8
Lee, Mi-Kyung; Gal, Maayan; Frydman, Lucio et al. (2010) Real-time multidimensional NMR follows RNA folding with second resolution. Proc Natl Acad Sci U S A 107:9192-7
Emani, Prashant S; Olsen, Gregory L; Echodu, Dorothy C et al. (2010) Slow exchange model of nonrigid rotational motion in RNA for combined solid-state and solution NMR studies. J Phys Chem B 114:15991-6002
Huang, Wei; Varani, Gabriele; Drobny, Gary P (2010) 13C/15N-19F intermolecular REDOR NMR study of the interaction of TAR RNA with Tat peptides. J Am Chem Soc 132:17643-5
Olsen, Greg L; Bardaro Jr, Michael F; Echodu, Dorothy C et al. (2009) Hydration dependent dynamics in RNA. J Biomol NMR 45:133-42
Olsen, Greg L; Echodu, Dorothy C; Shajani, Zahra et al. (2008) Solid-state deuterium NMR studies reveal micros-ns motions in the HIV-1 transactivation response RNA recognition site. J Am Chem Soc 130:2896-7

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