title: Identification of a high affinity nucleocapsid protein binding element from the bovine leukemia virus genome.
abstract: Retroviral genome recognition is mediated by interactions between the nucleocapsid (NC) domain of the virally encoded Gag polyprotein and cognate RNA packaging elements that, for most retroviruses, appear to reside primarily within the 5'-untranslated region (5'-UTR) of the genome. Recent studies suggest that a major packaging determinant of bovine leukemia virus (BLV), a member of the human T-cell leukemia virus (HTLV)/BLV family and a non-primate animal model for HTLV-induced leukemogenesis, resides within the gag open reading frame. We have prepared and purified the recombinant BLV NC protein and conducted electrophoretic mobility shift and isothermal titration calorimetry studies with RNA fragments corresponding to these proposed packaging elements. The gag-derived RNAs did not exhibit significant affinity for NC, suggesting an alternate role in packaging. However, an 83-nucleotide fragment of the 5'-UTR that resides just upstream of the gag start codon binds NC stoichiometrically and with high affinity (K(d)=136±21 nM). These nucleotides were predicted to form tandem hairpin structures, and studies with smaller fragments indicate that the NC binding site resides exclusively within the distal hairpin (residues G369-U399, K(d)=67±8 nM at physiological ionic strength). Unlike all other structurally characterized retroviral NC binding RNAs, this fragment is not expected to contain exposed guanosines, suggesting that RNA binding may be mediated by a previously uncharacterized mechanism.
authors: Howard Hughes Medical Institute (HHMI) and Department of Chemistry and Biochemistry, University of Maryland Baltimore County, 1000 Hilltop Circle, Baltimore, MD 21250, USA.
authors: F Zehra
authors: FZ
authors: Yildiz
authors: Kathlene
authors: K
authors: Babalola
authors: Michael F
authors: MF
authors: Summers
mesh_terms: D020121:5' Untranslated Regions; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D016583:Enzootic Bovine Leukosis / Q000821:virology*; D016679:Genome, Viral; D007700:Kinetics; D001909:Leukemia Virus, Bovine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D019590:Nucleocapsid Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D020121:5' Untranslated Regions; D019590:Nucleocapsid Proteins; D012367:RNA, Viral