Recognition of RNA by Peptides and Design Molecules

Scripps Research Institute, La Jolla CA

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Abstract

This proposal seeks to enhance our understanding of the thermodynamics of protein-RNA interactions through the study of model peptides based on the arginine-rich domain of the human immunodeficiency virus (HIV)Rev protein. The specific binding of ions and conformation changes in the virus (HIV)Rev protein. The specific binding of ions and conformation changes in the macromolecules can make substantial energetic contributions to protein-RNA binding; however, these thermodynamic effects have not been well studied in most systems. To evaluate how these equilibria are linked to RNA recognition, I will perform a systematic analysis of the interaction of Rev peptides with their RNA target site, the Rev response element (RRE). By studying the thermodynamics of association as a function of pH, salt concentration, and peptide stability, the effects of both ion binding and peptide folding on RNA recognition will be assessed. In addition, I will examine peptide-RNA interactions engineered to have increased affinity and specificity in which the Rev helix is stabilized in a zinc finger context and dimerized. Related strategies have resulted in the successful development of other high affinity DNA and RNA binders, but a detailed analysis of how dimerization affects the linked equilibria has not been conducted. Binding and calorimetric experiments will be used to assess the effects of ion binding and conformational changes in this designed system, further enhancing our understanding of dimeric protein-nucleic acid interactions.