The ultimate goal of this work is to understand RNA structure and function relations. Despite biological examples of the importance of RNA structure, there are few physical studies which describe the influence of RNA structure on systems such as RNA:protein complexes or RNA:RNA interactions. The work proposed here has two components: 1) investigation of the biochemical and biophysical parameters of the association of a sequence-specific RNA binding protein, human U1A, with its RNA substrate. U1A is one of the proteins of U1 snRNP, which binds to the U1 snRNA stem/loop II. The U1A protein contains a structural and sequence-dependent motif which is common to many RNA binding proteins, the so-called 'RNP conserved motif', or alternatively, 'RRM'. Understanding the interactions of this system will provide more information not only on how the proteins and RNA of the snRNP assemble, but should also serve as a model for the general properties of this class of RNA binding protein. 2) RNA:RNA interactions include base pairing in duplexes and in structural motifs containing unpaired bases, tertiary interactions in loop structures, and nonstandard interactions using modified bases. RNAs containing these elements will be investigated here to determine their thermodynamic and structural properties, again with the goal of understanding these features of RNA when they are found in biological sequences. All RNAs will be studied by NMR and chemical and enzymatic probing to determine their structure, as well as characterized thermodynamically. RNA molecules will be synthesized enzymatically or chemically; when enzymatic synthesis is used, it will be possible to incorporate isotopically labelled nucleotides to facilitate NMR structural analysis.
| Hall, Kathleen B (2012) Spectroscopic probes of RNA structure and dynamics. Methods Mol Biol 875:67-84 |
