Abstract

The replication of HIV-1 occurs through the reverse-transcription of its RNA genome into DMA. This complex process is chaperoned by the nucleocapsid protein (NC), and occurs through a series of intermediate steps. One sub-step in the replication is minus-strand transfer, in which NC assists the annealing of transactivation response (TAR) DNA to the complementary 5' end of the viral RNA in preparation for the completion of minus-strand synthesis. This step in the reverse transcription is in itself complex, because it involves alternate pathways and intermediates, and requires the presence of NC to achieve the thermodynamically optimal product: fully annealed DNA/RNA. Single molecule spectroscopy (SMS) is an ideal technique to study such an innately heterogeneous thermodynamic process. I propose to use SMS to characterize the pathway of this step in the reverse-transcription process, and to elucidate any crucial intermediates in the annealing of TAR DNA/RNA. Confocal, wide-field, and correlation techniques will be used to study native and mutated nucleic acid/NC interactions. Specifically, the experiments I have proposed are designed to determine the.rate-limiting step in the annealing process, and the nature of the nucleation event.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
1F32GM073534-01
Application #
6885138
Study Section
AIDS Molecular and Cellular Biology Study Section (AMCB)
Program Officer
Flicker, Paula F
Project Start
2004-12-11
Project End
2006-06-10
Budget Start
2004-12-11
Budget End
2005-12-10
Support Year
1
Fiscal Year
2005
Total Cost
$43,976
Indirect Cost

  Institution

Name
University of Texas Austin
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
170230239
City
Austin
State
TX
Country
United States
Zip Code
78712

  Publications

Shuang, Bo; Byers, Chad P; Kisley, Lydia et al. (2013) Improved analysis for determining diffusion coefficients from short, single-molecule trajectories with photoblinking. Langmuir 29:228-34
Daniels, C R; Tauzin, L J; Foster, E et al. (2013) On the pH-responsive, charge-selective, polymer-brush-mediated transport probed by traditional and scanning fluorescence correlation spectroscopy. J Phys Chem B 117:4284-90
Reznik, Carmen; Landes, Christy F (2012) Transport in supported polyelectrolyte brushes. Acc Chem Res 45:1927-35
Reznik, Carmen; Berg, Rosalie; Foster, Ed et al. (2011) Transient 3-Dimensional Orientation of Molecular Ions in an Ordered Polyelectrolyte Membrane. J Phys Chem Lett 2:
Daniels, Charlisa R; Reznik, Carmen; Kilmer, Rachel et al. (2011) Permeability of anti-fouling PEGylated surfaces probed by fluorescence correlation spectroscopy. Colloids Surf B Biointerfaces 88:31-8
Daniels, Charlisa R; Reznik, Carmen; Landes, Christy F (2010) Dye diffusion at surfaces: charge matters. Langmuir 26:4807-12
Zeng, Yining; Liu, Hsiao-Wei; Landes, Christy F et al. (2007) Probing nucleation, reverse annealing, and chaperone function along the reaction path of HIV-1 single-strand transfer. Proc Natl Acad Sci U S A 104:12651-6
Landes, Christy F; Zeng, Yining; Liu, Hsiao-Wei et al. (2007) Single-molecule study of the inhibition of HIV-1 transactivation response region DNA/DNA annealing by argininamide. J Am Chem Soc 129:10181-8
Liu, Hsiao-Wei; Zeng, Yining; Landes, Christy F et al. (2007) Insights on the role of nucleic acid/protein interactions in chaperoned nucleic acid rearrangements of HIV-1 reverse transcription. Proc Natl Acad Sci U S A 104:5261-7
Cosa, Gonzalo; Zeng, Yining; Liu, Hsiao-Wei et al. (2006) Evidence for non-two-state kinetics in the nucleocapsid protein chaperoned opening of DNA hairpins. J Phys Chem B 110:2419-26

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