STRUCTURE-BASED STUDIES OF RNA BINDING PROTEINS OF HIV

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We are working on RNA-protein Interaction in HIV genome. RNA stemloop called SL1, located near the 5'-end of the HIV-1 genome, mediates several functions crucial for viral assembly, and so is a potential new target for antiviral therapy. By binding the nucleocapsid (NCp7) portion of the HIV Gag protein, SL1 forms part of the packaging signal that targets genomic RNA into virions. Contact between SL1 loop in a pair of HIV RNAs also initiates genomic dimerization- a process that is facilitated by NCp7 binding and is essential for full infectivity. All biological activities of SL1 depend on its three-dimensional structure. Using heteronuclear labeling and multidimensional NMR, we propose to solve the corresponding structures of full-length, authentic SL1, which contains additional features that are biologically important. SL1 dimer is a 70nt RNA with molecular weight nearging 22kD. By determining these structures, we seek to understand the remarkable efficiency with which SL1 dimerizes and the chemical features that stabilize each successive conformation. We will also determine the structures and dynamics of complexes formed when HIV NCp7 protein binds each RNA conformer, revealing the exact molecular contacts involved and the mechanism by which NCp7 facilitates refolding of SL1 and other nucleic acids. In addition, we will solve the complex formed by NCp7 with a heterologous, monomeric RNA ligand (aptamer) that binds with higher affinity than SL1, to identify features that account for its tight binding and could be exploited in rational drug design. Results of these studies will provide a basis for discovering new antivirals that target the SL1/NCp7 interaction.