title: 13C/15N-19F intermolecular REDOR NMR study of the interaction of TAR RNA with Tat peptides.
abstract: The complex of the HIV TAR RNA with the viral regulatory protein Tat is of considerable interest, but the plasticity of this interaction has made it impossible so far to establish the structure of that complex. In order to explore a new approach to obtain structural information on protein-RNA complexes, we performed (13)C/(15)N-(19)F REDOR NMR experiments in the solid state on TAR bound to a peptide comprising the RNA-binding section of Tat. A critical arginine in the peptide was uniformly (13)C and (15)N labeled, and 5-fluorouridine was incorporated at the U23 position of TAR. REDOR irradiation resulted in dephasing of the (13)C and (15)N resonances, indicating the proximity of the U23(5F)-C and U23(5F)-N spin pairs. Best fits to the REDOR data show the U23(5F)-C distances and the U23(5F)-N distances are in good agreement with the distances obtained from solution NMR structures of partial complexes of Tat with TAR. These results demonstrate that it is possible to study protein-RNA complexes using solid-state REDOR NMR measurements, adding to a growing list of solid state techniques for studying protein-nucleic acid complexes.
authors.affiliation: Department of Chemistry, University of Washington, Box 351700, Seattle, Washington 98195, United States.
authors.forename: Wei
authors.initials: W
authors.lastname: Huang
authors.forename: Gabriele
authors.initials: G
authors.lastname: Varani
authors.forename: Gary P
authors.initials: GP
authors.lastname: Drobny
mesh_terms: D001483:Base Sequence; D002247:Carbon Isotopes / Q000737:chemistry; D005461:Fluorine / Q000737:chemistry; D015696:Gene Products, tat / Q000737:chemistry* / Q000235:genetics; D015497:HIV-1; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009587:Nitrogen Isotopes / Q000737:chemistry; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D000099253:Trans-Activation Responsive RNA-Binding Protein
chemical_list: D002247:Carbon Isotopes; D005461:Fluorine; D015696:Gene Products, tat; D009587:Nitrogen Isotopes; D016601:RNA-Binding Proteins; D000099253:Trans-Activation Responsive RNA-Binding Protein