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Characterizing RNA ensembles from NMR data with kinematic models.

Fonseca R, Pachov DV, Bernauer J, van den Bedem H - Nucleic Acids Res. (2014)

Bottom Line: We found that KGSrna ensembles accurately represent the conformational landscapes of 3D RNA encoded by NMR proton chemical shifts.KGSrna resolves motionally averaged NMR data into structural contributions; when coupled with residual dipolar coupling data, a KGSrna ensemble revealed a previously uncharacterized transient excited state of the HIV-1 trans-activation response element stem-loop.Ensemble-based interpretations of averaged data can aid in formulating and testing dynamic, motion-based hypotheses of functional mechanisms in RNAs with broad implications for RNA engineering and therapeutic intervention.

View Article: PubMed Central - PubMed

Affiliation: AMIB Project, INRIA Saclay-Île de France, 1 rue Honoré d'Estienne d'Orves, Bâtiment Alan Turing, Campus de l'École Polytechnique, 91120 Palaiseau, France Laboratoire d'Informatique de l'École Polytechnique (LIX), CNRS UMR 7161, École Polytechnique, 91128 Palaiseau, France Department of Computer Science, University of Copenhagen, Nørre Campus, Universitetsparken 5, DK-2100 Copenhagen, Denmark.

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Structural characterization of conformational substates of the apical Tat binding loop of HIV-1 TAR. (a) Paired measured RDCs for apical loop nucleotides and those predicted from a 10-member weighted ensemble (inset) obtained from fitting 20 000 KGSrna samples to measured RDCs with a quadratic program. The data points are expected to lie along a 45° line if measured RDCs are accurately predicted. The coefficient of determination for the predicted RDCs equals 0.98. (b) Observed (solid symbols) and predicted (open symbols) RDCs for apical loop nucleotides. Smaller magnitudes for RDCs generally indicate more angular mobility in the bond vectors. (c) Schematic of the GS (top panel, left) and the ES (top panel, right) corresponding to the 3D structures closest to the GS and the ES in the 10-member ensemble. The bottom left panel shows the GS highlighted in the ensemble, with the other members translucent in the background. The bottom right panel shows the ES identified from biasing the sampling toward pairing C30–A35 and U31–G34. (d) Time evolution of the hydrogen bond distances between reverse wobble pair C30–A35 (blue colors) and GU wobble pair U31–G34 (red colors) in the ES of HIV-1 TAR for 80 ns of the molecular dynamics trajectory. The distances shown are between heavy donor and acceptor atoms, sampled every 100 ps. Along the trajectory, the apical loop maintains a helical structure (inset at 35 ns) until, at 65 ns, pairing of U31–G34 and subsequently C30–A35 is disrupted.
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Figure 5: Structural characterization of conformational substates of the apical Tat binding loop of HIV-1 TAR. (a) Paired measured RDCs for apical loop nucleotides and those predicted from a 10-member weighted ensemble (inset) obtained from fitting 20 000 KGSrna samples to measured RDCs with a quadratic program. The data points are expected to lie along a 45° line if measured RDCs are accurately predicted. The coefficient of determination for the predicted RDCs equals 0.98. (b) Observed (solid symbols) and predicted (open symbols) RDCs for apical loop nucleotides. Smaller magnitudes for RDCs generally indicate more angular mobility in the bond vectors. (c) Schematic of the GS (top panel, left) and the ES (top panel, right) corresponding to the 3D structures closest to the GS and the ES in the 10-member ensemble. The bottom left panel shows the GS highlighted in the ensemble, with the other members translucent in the background. The bottom right panel shows the ES identified from biasing the sampling toward pairing C30–A35 and U31–G34. (d) Time evolution of the hydrogen bond distances between reverse wobble pair C30–A35 (blue colors) and GU wobble pair U31–G34 (red colors) in the ES of HIV-1 TAR for 80 ns of the molecular dynamics trajectory. The distances shown are between heavy donor and acceptor atoms, sampled every 100 ps. Along the trajectory, the apical loop maintains a helical structure (inset at 35 ns) until, at 65 ns, pairing of U31–G34 and subsequently C30–A35 is disrupted.

Mentions: This procedure identified a 10-member, weighted ensemble from the sample set starting from model seven in the NMR bundle that agrees extremely well with experimentally observed RDC values (Figure 5a). The coefficient of determination between observed values and those predicted from the weighted ensemble equals 0.98. The predicted values of the ensemble accurately reflect the mobility of riboses and nucleobases, with small in magnitude indicating elevated mobility (Figure 5b). The RMSD between observed and predicted values is 1.55 Hz, below the experimental error of 2–4 Hz (16,18).


Characterizing RNA ensembles from NMR data with kinematic models.

Fonseca R, Pachov DV, Bernauer J, van den Bedem H - Nucleic Acids Res. (2014)

Structural characterization of conformational substates of the apical Tat binding loop of HIV-1 TAR. (a) Paired measured RDCs for apical loop nucleotides and those predicted from a 10-member weighted ensemble (inset) obtained from fitting 20 000 KGSrna samples to measured RDCs with a quadratic program. The data points are expected to lie along a 45° line if measured RDCs are accurately predicted. The coefficient of determination for the predicted RDCs equals 0.98. (b) Observed (solid symbols) and predicted (open symbols) RDCs for apical loop nucleotides. Smaller magnitudes for RDCs generally indicate more angular mobility in the bond vectors. (c) Schematic of the GS (top panel, left) and the ES (top panel, right) corresponding to the 3D structures closest to the GS and the ES in the 10-member ensemble. The bottom left panel shows the GS highlighted in the ensemble, with the other members translucent in the background. The bottom right panel shows the ES identified from biasing the sampling toward pairing C30–A35 and U31–G34. (d) Time evolution of the hydrogen bond distances between reverse wobble pair C30–A35 (blue colors) and GU wobble pair U31–G34 (red colors) in the ES of HIV-1 TAR for 80 ns of the molecular dynamics trajectory. The distances shown are between heavy donor and acceptor atoms, sampled every 100 ps. Along the trajectory, the apical loop maintains a helical structure (inset at 35 ns) until, at 65 ns, pairing of U31–G34 and subsequently C30–A35 is disrupted.
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Related In: Results  -  Collection

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Figure 5: Structural characterization of conformational substates of the apical Tat binding loop of HIV-1 TAR. (a) Paired measured RDCs for apical loop nucleotides and those predicted from a 10-member weighted ensemble (inset) obtained from fitting 20 000 KGSrna samples to measured RDCs with a quadratic program. The data points are expected to lie along a 45° line if measured RDCs are accurately predicted. The coefficient of determination for the predicted RDCs equals 0.98. (b) Observed (solid symbols) and predicted (open symbols) RDCs for apical loop nucleotides. Smaller magnitudes for RDCs generally indicate more angular mobility in the bond vectors. (c) Schematic of the GS (top panel, left) and the ES (top panel, right) corresponding to the 3D structures closest to the GS and the ES in the 10-member ensemble. The bottom left panel shows the GS highlighted in the ensemble, with the other members translucent in the background. The bottom right panel shows the ES identified from biasing the sampling toward pairing C30–A35 and U31–G34. (d) Time evolution of the hydrogen bond distances between reverse wobble pair C30–A35 (blue colors) and GU wobble pair U31–G34 (red colors) in the ES of HIV-1 TAR for 80 ns of the molecular dynamics trajectory. The distances shown are between heavy donor and acceptor atoms, sampled every 100 ps. Along the trajectory, the apical loop maintains a helical structure (inset at 35 ns) until, at 65 ns, pairing of U31–G34 and subsequently C30–A35 is disrupted.
Mentions: This procedure identified a 10-member, weighted ensemble from the sample set starting from model seven in the NMR bundle that agrees extremely well with experimentally observed RDC values (Figure 5a). The coefficient of determination between observed values and those predicted from the weighted ensemble equals 0.98. The predicted values of the ensemble accurately reflect the mobility of riboses and nucleobases, with small in magnitude indicating elevated mobility (Figure 5b). The RMSD between observed and predicted values is 1.55 Hz, below the experimental error of 2–4 Hz (16,18).

Bottom Line: We found that KGSrna ensembles accurately represent the conformational landscapes of 3D RNA encoded by NMR proton chemical shifts.KGSrna resolves motionally averaged NMR data into structural contributions; when coupled with residual dipolar coupling data, a KGSrna ensemble revealed a previously uncharacterized transient excited state of the HIV-1 trans-activation response element stem-loop.Ensemble-based interpretations of averaged data can aid in formulating and testing dynamic, motion-based hypotheses of functional mechanisms in RNAs with broad implications for RNA engineering and therapeutic intervention.

View Article: PubMed Central - PubMed

Affiliation: AMIB Project, INRIA Saclay-Île de France, 1 rue Honoré d'Estienne d'Orves, Bâtiment Alan Turing, Campus de l'École Polytechnique, 91120 Palaiseau, France Laboratoire d'Informatique de l'École Polytechnique (LIX), CNRS UMR 7161, École Polytechnique, 91128 Palaiseau, France Department of Computer Science, University of Copenhagen, Nørre Campus, Universitetsparken 5, DK-2100 Copenhagen, Denmark.

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