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Elife. 2017 Jul 20;6. pii: e27055. doi: 10.7554/eLife.27055.

Dissection of specific binding of HIV-1 Gag to the 'packaging signal' in viral RNA.

Author information

1
HIV Dynamics and Replication Program, Center for Cancer Research, National Cancer Institute, Frederick, United States.
2
Xencor Inc., Monrovia, United States.
3
Optical Microscopy and Analysis Laboratory, Cancer Research Technology Program, Leidos Biomedical Research, Inc., Frederick, United States.

Abstract

Selective packaging of HIV-1 genomic RNA (gRNA) requires the presence of a cis-acting RNA element called the 'packaging signal' (Ψ). However, the mechanism by which Ψ promotes selective packaging of the gRNA is not well understood. We used fluorescence correlation spectroscopy and quenching data to monitor the binding of recombinant HIV-1 Gag protein to Cy5-tagged 190-base RNAs. At physiological ionic strength, Gag binds with very similar, nanomolar affinities to both Ψ-containing and control RNAs. We challenged these interactions by adding excess competing tRNA; introducing mutations in Gag; or raising the ionic strength. These modifications all revealed high specificity for Ψ. This specificity is evidently obscured in physiological salt by non-specific, predominantly electrostatic interactions. This nonspecific activity was attenuated by mutations in the MA, CA, and NC domains, including CA mutations disrupting Gag-Gag interaction. We propose that gRNA is selectively packaged because binding to Ψ nucleates virion assembly with particular efficiency.

KEYWORDS:

HIV; RNA; biophysics; infectious disease; microbiology; protein-RNA interaction; retrovirus; structural biology; virus; virus assembly

PMID:
28726630
PMCID:
PMC5531834
DOI:
10.7554/eLife.27055
[Indexed for MEDLINE]
Free PMC Article

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