application) The aim of this research is to identify low molecular weight compounds that bind to the key HIV regulatory RNA elements, TAR and RRE. The structures of the TAR-arginine complex and the Rev peptide-RRE complex that the PI recently determined using will be used as a guide for structure based design of RNA binding ligands. As a complementary effort, a combinatorial approach to screen for inhibitors from several types of peptide bond-based libraries of compounds will be used.
The specific aims are: 1) To determine the structure of HIV-1 Tat peptides in complex with HIV-2 TAR RNA. The HIV-2 TAR has two nucleotide UU-bulge, compared to the HIV-1 TAR bulge, UCU. The smaller bulge improves the dynamic properties of the TAR-arginine complex, and many new NOEs were observed. The complex between HIV-1 and HIV-2 Tat peptides and HIV-2 TAR will be examined, which should also exhibit improved dynamic properties. 2) To conduct small molecule screening of arginine based ligands for binding to TAR. The single amino acid arginine forms a specific complex with HIV TAR RNA with an affinity of about 1mM. A structure-based screen by NMR of small molecules containing guanidinium groups will be employed to identify additional interactions that can be formed close to the arginine binding pocket. 3) To design peptides with increased affinity and specificity for the RRE. The investigators have determined the structure of the complex between a 22 residue peptide derived from the basic region of Rev and a 34 nucleotide RRE RNA, that has a binding constant of about 10mM.
The aim i s to design peptides that include additional specific interactions, taking advantage of """"""""unused"""""""" residues at the RNA-protein interface. 4) To design a left-handed helical Rev peptide analog constructed from D- amino acids. The structure of the RRE binding site includes a deep cleft in which an alpha-helix formed by the Rev peptide binds. There are no contracts to the backbone of Rev from the RNA. An attempt will be made to design a helical peptide from D-amino acids that retains the essential specific contract the Rev peptide.
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