Nucleic Acid Chaperone Activity of HIV‐1 Nucleocapsid Protein: Critical Role in Reverse Transcription and Molecular Mechanism

Levin, Joseph; Guo, Jianhui; Rouzina, Ioulia; Musier‐Forsyth, Karin

doi:10.1016/s0079-6603(05)80006-6

Cited by 317 publications

(543 citation statements)

References 391 publications

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“…[124][125][126][127][128][129][130][131][132][133][134][135][136] This chaperone activity, facilitating restructuring of the nucleic acid complex, has been attributed to both an aggregating ability of NC and a facility for duplex destabilization. 125,131,[137][138][139][140][141][142][143][144][145] This destabilization, however, is weak, 143,144,146,147 and will not completely melt DNA without a complementary strand. 6,138,148,149 Stretching and relaxation cycles can also reveal information on the kinetics of protein association and dissociation.…”

Section: Figure 11mentioning

confidence: 99%

Mechanisms of DNA binding determined in optical tweezers experiments

McCauley¹,

Williams²

2006

Biopolymers

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The last decade has seen rapid development in single molecule manipulation of RNA and DNA. Measuring the response force for a particular manipulation has allowed the free energies of various nucleic acid structures and configurations to be determined. Optical tweezers represent a class of single molecule experiments that allows the energies and structural dynamics of DNA to be probed up to and beyond the transition from the double helix to its melted single strands. These experiments are capable of high force resolution over a wide dynamic range. Additionally, these investigations may be compared with results obtained when the nucleic acids are in the presence of proteins or other binding ligands. These ligands may bind into the major or minor groove of the double helix, intercalate between bases or associate with an already melted single strand of DNA. By varying solution conditions and the pulling dynamics, energetic and dynamic information may be deduced about the mechanisms of binding to nucleic acids, providing insight into the function of proteins and the utility of drug treatments. © 2006 Wiley Periodicals, Inc. Biopolymers 85:154–168, 2007.This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com

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Section: Figure 11mentioning

confidence: 99%

Mechanisms of DNA binding determined in optical tweezers experiments

McCauley¹,

Williams²

2006

Biopolymers

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“…Many of these functions rely on the nucleic acid chaperone activity of NC, i.e. its ability to catalyse nucleic acid conformational rearrangements that lead to the most thermodynamically stable structure (for recent reviews [4][5][6]). …”

Section: Introductionmentioning

confidence: 99%

New insights into the formation of HIV-1 reverse transcription initiation complex

et al. 2007

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HIV-1 reverse transcriptase uses the host tRNA 3 Lys as a primer for the synthesis of the minus DNA strand. The first event in viral replication is thus the annealing of tRNA to the primer binding site (PBS) in the 5' UTR of the viral RNA. This event requires a major RNA rearrangement which is chaperoned by the viral NC protein. The binding of NC to nucleic acids is essentially non-specific, however, NC is known to bind selectively to hairpins located in the 5' region of the viral RNA. In a previous study, using an NMR approach in which the reaction is slowed down by controlling temperature, we were able to follow details in this RNA unfolding/refolding process and to uncover an intermediate state. We showed that annealing initiates at the junction between the acceptor and the TΨC stems, and that, at physiological temperature, complete annealing is reached only in the presence of NC, probably when the zinc fingers contact the TΨC/D loops.In the present work, we have refined our model of the formation of the tRNA 3 Lys /PBS duplex. First, we show that annealing can initiate both from the single-stranded CCA 3'-end bases of the acceptor stem and from the bases in the TΨC stem. Secondly, by NMR and fluorescence spectroscopy, we have studied the complex between the NC protein and RNA hairpins that mimic the D and T-arms of the tRNA 3 Lys . Interestingly, the NC protein shows strong and specific binding to the D-arm of tRNA 3 Lys , which could explain the overall annealing mechanism.

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“…It plays essential roles in nearly every step of the viral replication cycle, including genomic RNA packaging, reverse transcription initiation, minus strand transfer, and DNA integration (2,31,32). Two distinct and opposing features of NC-nucleic acid interactions have been described previously in detail: a duplex-stabilizing effect through its nonspecific interaction with phosphates and a duplex-destabilizing effect derived from the preferential binding of its zinc fingers to unpaired bases (33). Efficient viral strand transfer relies on both abilities of NC.…”

mentioning

confidence: 97%

“…Efficient viral strand transfer relies on both abilities of NC. The duplex-destabilization activity of NC disrupts RNA secondary structures that may interfere with the complementary viral genomic RNA and newly synthesized DNA annealing, and then the duplex-stabilizing activity of NC facilitates the intermolecular interactions between the complementary viral RNA and DNA (33). NC promotes strand transfer at all stages of reverse transcription: minus strand transfer, template switches during minus strand synthesis, and plus strand transfer (34 -36).…”

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confidence: 99%

HIV-1 Nucleocapsid Protein Increases Strand Transfer Recombination by Promoting Dimeric G-quartet Formation

Shen

Gorelick

Bambara

2011

Journal of Biological Chemistry

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A preferred site for HIV-1 recombination was identified in vivo and in vitro surrounding the beginning of the HIV-1 gag gene. This G-rich gag hotspot for recombination contains three evenly spaced G-runs that stalled reverse transcriptase. Disruption of the G-runs suppressed both the associated pausing and strand transfer in vitro. Significantly, this same gag sequence was able to fold into a G-quartet monomer, dimer, and tetramer, depending on the cations employed. The pause band at the G-run (nucleotide (nt) 405-409), which was predicted to be involved in forming a G-quartet monomer, diminished with increased HIV-1 nucleocapsid (NC) protein. More NC induced stronger pauses at other G-runs (nt 363-367 and nt 382-384), a region that forms a G-quartet dimer, adhering the two RNA templates. We hypothesized that NC induces the unfolding of the monomeric G-quartet but stabilizes the dimeric interaction. We tested this by inserting a known G-quartet formation sequence, 5-(UGGGGU) 4 -3, into a relatively structure-free template from the HIV-1 pol gene. Strand transfer assays were performed with cations that either encourage (K ؉ ) or discourage (Li ؉ ) G-quartet formation with or without NC. Strikingly, a G-quartet monomer was observed without NC, whereas a G-quartet dimer was observed with NC, both only in the presence of K ؉ . Moreover, the transfer efficiency of the dimerized template (with K ؉ and NC) reached about 90%, approximately 2.5-fold of that of the non-dimerized template. Evidently, template dimerization induced by NC creates a proximity effect, leading to the unique high peak of transfer at the gag recombination hotspot.

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Nucleic Acid Chaperone Activity of HIV‐1 Nucleocapsid Protein: Critical Role in Reverse Transcription and Molecular Mechanism

Cited by 317 publications

References 391 publications

Mechanisms of DNA binding determined in optical tweezers experiments

Mechanisms of DNA binding determined in optical tweezers experiments

New insights into the formation of HIV-1 reverse transcription initiation complex

HIV-1 Nucleocapsid Protein Increases Strand Transfer Recombination by Promoting Dimeric G-quartet Formation

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