Concerted motions in HIV-1 TAR RNA may allow access to bound state conformations: RNA dynamics from NMR residual dipolar couplings 1 1Edited by M. F. Summers

Al‐Hashimi, Hashim M.; Gosser, Yuying; Gorin, Andrey; Hu, Weidong; Majumdar, Ananya; Patel, Dinshaw J.

doi:10.1006/jmbi.2001.5235

Cited by 146 publications

(295 citation statements)

References 35 publications

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“…These above results are consistent with the experimental view that the bound TAR RNA is stabilized by weak intermolecular interactions with the ligand 27 and that bound conformations are transiently sampled in the free ensemble, 4,5 following the conformation selection model. The results are also encouraging from an application point of view in that those simulated TAR RNA conformations can be used as receptor structures for docking.…”

Section: Discussionsupporting

confidence: 89%

“…Notably, these numbers are of the same order of magnitude as, but smaller than, the binding free energies of the respective ligands such that weak intermolecular interactions with the ligands should suffice to stabilize bound TAR RNA. 27 Furthermore, the small conformational free energy differences are consistent with the view that bound TAR conformations are transiently sampled in the free ensemble, 4,5 following the conformation selection model.…”

Section: Resultssupporting

confidence: 73%

“…This rigid-cluster decomposition reflects experimental findings that in the free conformation of TAR RNA the two stable helical stems collectively undergo a large-amplitude, hinge-like motion around the flexible bulge region. 4,6 Consequently, when used as input to the constrained geometrical simulation FRODA, TAR RNA motions are successfully predicted based on the identified bonds that limit motion within molecules and their coupling.…”

Section: Resultsmentioning

confidence: 99%

“…In this regard, one of the best investigated RNA structures is the HIV-1 transactivation response element (TAR). [2][3][4][5][6] TAR RNA is a 59-nucleotide hairpin structure located at the 5′-end of all pre-mRNA in a HIV-1 transcript. Binding of the regulatory Tat protein to a bulge region within the TAR RNA structure is an essential step for HIV-1 viral replication.…”

Section: Introductionmentioning

confidence: 99%

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HIV-1 TAR RNA Spontaneously Undergoes Relevant Apo-to-Holo Conformational Transitions in Molecular Dynamics and Constrained Geometrical Simulations

Fulle

Christ

Kestner

et al. 2010

J. Chem. Inf. Model.

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We report all-atom molecular dynamics and replica exchange molecular dynamics simulations on the unbound human immunodeficiency virus type-1 (HIV-1) transactivation responsive region (TAR) RNA structure and three TAR RNA structures in bound conformations of, in total, ∼250 ns length. We compare the extent of observed conformational sampling with that of the conceptually simpler and computationally much cheaper constrained geometrical simulation approach framework rigidity optimized dynamic algorithm (FRODA). Atomic fluctuations obtained by replica-exchange molecular dynamics (REMD) simulations agree quantitatively with those obtained by molecular dynamics (MD) and FRODA simulations for the unbound TAR structure. Regarding the stereochemical quality of the generated conformations, backbone torsion angles and puckering modes of the sugar-phosphate backbone were reproduced equally well by MD and REMD simulations, but further improvement is needed in the case of FRODA simulations. Essential dynamics analysis reveals that all three simulation approaches show a tendency to sample bound conformations when starting from the unbound TAR structure, with MD and REMD simulations being superior with respect to FRODA. These results are consistent with the experimental view that bound TAR RNA conformations are transiently sampled in the free ensemble, following a conformation selection model. The simulation-generated TAR RNA conformations have been successfully used as receptor structures for docking. This finding has important implications for RNA-ligand docking in that docking into an ensemble of simulation-generated RNA structures is shown to be a valuable means to cope with large apo-to-holo conformational transitions of the receptor structure.

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Section: Discussionsupporting

confidence: 89%

Section: Resultssupporting

confidence: 73%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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HIV-1 TAR RNA Spontaneously Undergoes Relevant Apo-to-Holo Conformational Transitions in Molecular Dynamics and Constrained Geometrical Simulations

Fulle

Christ

Kestner

et al. 2010

J. Chem. Inf. Model.

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“…The HIV TAR-Tat interaction has been a subject of major attention in the past two decades, both for understanding the mechanism of transactivation and for development of anti-HIV therapeutics (9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)30), and as a paradigm for the mechanism underlying protein-RNA recognition and signaling observed in a wide range of posttranscriptional regulatory processes. Our results reveal the structure of an intermediate in this interaction, illustrating how the use of RDCs as structural restraints in RAM simulations, particularly with further experimental validation through structure-based mutant design, provides a general strategy for obtaining high-resolution structures of low-population intermediates of RNA-protein complexes, which are very challenging for more conventional structure determination or dynamic techniques.…”

Section: Discussionmentioning

confidence: 99%

Structure of a low-population binding intermediate in protein-RNA recognition

Borkar

Bardaro

Camilloni

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The interaction of the HIV-1 protein transactivator of transcription (Tat) and its cognate transactivation response element (TAR) RNA transactivates viral transcription and represents a paradigm for the widespread occurrence of conformational rearrangements in protein-RNA recognition. Although the structures of free and bound forms of TAR are well characterized, the conformations of the intermediates in the binding process are still unknown. By determining the free energy landscape of the complex using NMR residual dipolar couplings in replica-averaged metadynamics simulations, we observe two low-population intermediates. We then rationally design two mutants, one in the protein and another in the RNA, that weaken specific nonnative interactions that stabilize one of the intermediates. By using surface plasmon resonance, we show that these mutations lower the release rate of Tat, as predicted. These results identify the structure of an intermediate for RNA-protein binding and illustrate a general strategy to achieve this goal with high resolution.RNA structure | NMR spectroscopy | metadynamics | exact RDC restraints | tensor-free method E ssentially all biochemical reactions taking place in living organisms are associated with macromolecular recognition events. A full understanding the molecular mechanisms underlying such events requires the characterization of binding intermediates, which are states that typically have lifetimes of less than a millisecond and may comprise only 5-15% of the conformational space of proteins (1) and nucleic acids (2, 3). Protein-protein and protein-DNA intermediates have recently been characterized at high resolution (4, 5), but despite considerable advances (3, 6-8), high-resolution structures for protein-RNA intermediates have not been reported yet.To address this problem, we focused on the well-studied process by which HIV, like other lentiviruses, hijacks the host transcription machinery to activate transcription of the viral genome (9-13). In HIV, transactivation (Fig. S1) requires binding of the transactivator of transcription (Tat) protein and the host positive transcription elongation factor b (P-TEFb) complex (11) to the transactivation response element (TAR), a 59-residue RNA stem-loop ( Fig. 1 and Fig. S1) with a highly dynamic structure (10, 12, 13). The NMR structures of free TAR (14-16) and of TAR bound to peptide fragments of Tat and to peptide mimetics of and other lentiviruses (21,22) revealed the conformational properties of TAR in its free and bound states, and demonstrated that this RNA molecule undergoes significant dynamic rearrangements associated with its functions. Although the TAR-Tat complex has become a paradigm for the widespread occurrence of conformational rearrangements and molecular adaptation in protein-RNA recognition, the pathway and intermediates linking the free and bound states of TAR are still unknown. Results and DiscussionDetermination of the Tat-TAR Free Energy Landscape. Following an approach recently described for proteins (4) we ident...

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