Nucleic Acids Research Advance Access originally published online on April 9, 2008
Nucleic Acids Research 2008 36(8):2799-2810; doi:10.1093/nar/gkn129
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Nucleic Acids Research, 2008, Vol. 36, No. 8 2799-2810
© 2008 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Structural Biology |
Structural probing of the HIV-1 polypurine tract RNA:DNA hybrid using classic nucleic acid ligands
1University of Maryland Baltimore County, Baltimore, 2Center for Advanced Research in Biotechnology of the University of Maryland Biotechnology Institute and the National Institute of Standards and Technology, Rockville and 3HIV Drug Resistance Program, NCI, National Institutes of Health, Frederick, MD, USA
*To whom correspondence should be addressed. Tel: +1 410 455 3053; Fax: +1 410 455 2608; Email: fabris{at}umbc.edu Correspondence may also be addressed to John P. Marino. Tel: +1 240 314 6160; Fax: +1 240 314 6255; Email: marino{at}umbi.umd.edu
Received January 5, 2008. Revised February 22, 2008. Accepted March 9, 2008.
The interactions of archetypical nucleic acid ligands with the HIV-1 polypurine tract (PPT) RNA:DNA hybrid, as well as analogous DNA:DNA, RNA:RNA and swapped hybrid substrates, were used to probe structural features of the PPT that contribute to its specific recognition and processing by reverse transcriptase (RT). Results from intercalative and groove-binding ligands indicate that the wild-type PPT hybrid does not contain any strikingly unique groove geometries and/or stacking arrangements that might contribute to the specificity of its interaction with RT. In contrast, neomycin bound preferentially and selectively to the PPT near the 5'(rA)4:(dT)4 tract and the 3' PPT-U3 junction. Nuclear magnetic resonance data from a complex between HIV-1 RT and the PPT indicate RT contacts within the same regions highlighted on the PPT by neomycin. These observations, together with the fact that the sites are correctly spaced to allow interaction with residues in the ribonuclease H (RNase H) active site and thumb subdomain of the p66 RT subunit, suggest that despite the long cleft employed by RT to make contact with nucleic acids substrates, these sites provide discrete binding units working in concert to determine not only specific PPT recognition, but also its orientation on the hybrid structure.
The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors
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