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Structural Basis of HIV‐1 Genome Recognition via Viral RNA‐Protein Interaction
Author(s) -
Florwick Alyssa,
Carter Greg,
Keane Sarah,
Summers Michael
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.lb219
Subject(s) - isothermal titration calorimetry , genome , computational biology , rna , genetics , binding site , biology , untranslated region , viral protein , chemistry , virus , gene , biochemistry
According to the World Health Organization, more than 35 million people worldwide are currently living with Human Immunodeficiency Virus (HIV). There are 31 antiretroviral drugs to date approved for treatment, yet there is no cure for infection. To inform new drug development strategies, we study the mechanism of genome packaging, a crucial step in viral proliferation. Packaging is mediated via the highly specific selection of the dimeric genome by the nucleocapsid (NC) domain of viral Gag polyprotein. Previous studies have identified the Core Encapsidation Signal (CES), the minimal area of the 5' untranslated region required to direct genome packaging. Using NMR spectroscopy, we have determined the 3‐D structure of CES. This structure contains tandem three‐way junctions that sequester the splice donor site. We propose that the tandem three‐way junctions, along with the psi hairpin, both of which containing a number of exposed or weakly base‐paired guanosines, have a large number of NC binding sites. To identify NC binding sites, we used site‐directed mutagenesis to mutate specific guanosines (G) to adenosines (A), which should inhibit NC binding. Using isothermal titration calorimetry, we performed binding studies with RNA constructs containing clusters of G to A mutations at the Psi hairpin and junctions. These data confirm the existence of NC binding sites at junction and hairpin Gs. Understanding the determinants of RNA‐protein interaction between CES and NC further elucidates the mechanism of genome selection. Better understanding of the structural mechanism of HIV‐1 genome recognition provides new possibilities for late‐phase viral inhibition. This research was funded by NIH/NIGMS grant 1P50GM103297.