RNA dynamics: perspectives from spin labels

Nguyen, Phuong H.; Qin, Peter Z.

doi:10.1002/wrna.104

Cited by 54 publications

(36 citation statements)

References 74 publications

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“…Several methods have been developed to attach a paramagnetic tag to nucleic acids (Keyes & Bobst, 2002;Nguyen & Qin, 2012;Qin & Dieckmann, 2004;Shelke & Sigurdsson, 2012): these include replacing structural Mg 2+ with paramagnetic Mn 2+ (Bonneau & Legault, 2014;Kisseleva, Khvorova, Westhof, & Schiemann, 2005), attaching paramagnetic groups to 2 0 -amino modified nucleotides or 4-thiouridines (Piton et al, 2007;Qin, Feigon, & Hubbell, 2005), and the use of paramagnetic phosphoramidites during RNA synthesis (Wunderlich et al, 2013). In the case of large RNAs, annealing and ligating short spin-labeled oligonucleotides to a larger RNA scaffold (Helmling et al, 2014), and enzymatically, using T7 RNA polymerase (Lebars et al, 2014) have been reported.…”

Section: Spin Labeling Of Protein-rna Complexesmentioning

confidence: 99%

Structural Analysis of Protein–RNA Complexes in Solution Using NMR Paramagnetic Relaxation Enhancements

Hennig

Warner

Simon

et al. 2015

Methods in Enzymology

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Section: Spin Labeling Of Protein-rna Complexesmentioning

confidence: 99%

Structural Analysis of Protein–RNA Complexes in Solution Using NMR Paramagnetic Relaxation Enhancements

Hennig

Warner

Simon

et al. 2015

Methods in Enzymology

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“…Techniques including NMR and EPR relaxation have been developed to incisively probe motions in the ensemble on different time scales, ranging from picoseconds to milliseconds (3,4). Nonetheless, such dynamic information represents an average of the dynamics of the molecules across the conformational ensemble.…”

Section: Helix-junction-helix | Saxsmentioning

confidence: 99%

From a structural average to the conformational ensemble of a DNA bulge

Shi¹,

Beauchamp

Harbury

et al. 2014

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

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Direct experimental measurements of conformational ensembles are critical for understanding macromolecular function, but traditional biophysical methods do not directly report the solution ensemble of a macromolecule. Small-angle X-ray scattering interferometry has the potential to overcome this limitation by providing the instantaneous distance distribution between pairs of gold-nanocrystal probes conjugated to a macromolecule in solution. Our X-ray interferometry experiments reveal an increasing bend angle of DNA duplexes with bulges of one, three, and five adenosine residues, consistent with previous FRET measurements, and further reveal an increasingly broad conformational ensemble with increasing bulge length. The distance distributions for the AAA bulge duplex (3A-DNA) with six different Au-Au pairs provide strong evidence against a simple elastic model in which fluctuations occur about a single conformational state. Instead, the measured distance distributions suggest a 3A-DNA ensemble with multiple conformational states predominantly across a region of conformational space with bend angles between 24 and 85 degrees and characteristic bend directions and helical twists and displacements. Additional X-ray interferometry experiments revealed perturbations to the ensemble from changes in ionic conditions and the bulge sequence, effects that can be understood in terms of electrostatic and stacking contributions to the ensemble and that demonstrate the sensitivity of X-ray interferometry. Combining X-ray interferometry ensemble data with molecular dynamics simulations gave atomic-level models of representative conformational states and of the molecular interactions that may shape the ensemble, and fluorescence measurements with 2-aminopurinesubstituted 3A-DNA provided initial tests of these atomistic models. More generally, X-ray interferometry will provide powerful benchmarks for testing and developing computational methods. helix-junction-helix | SAXSA grand challenge in biology is to understand the complex free-energy landscape of macromolecules and to decipher the resulting conformational ensembles. To perform their biological functions, macromolecules must adopt a multiplicity of conformations. Balancing and controlling different conformational states is central to biological processes including protein folding, allostery and signaling, and the stepwise assembly and function of macromolecular machines. To understand these complex molecules requires characterization of their free-energy landscapes-i.e., their equilibrium conformational ensembles. Precise measurements of conformational ensembles could allow quantitative modeling of the folding and function of biological macromolecules, would provide valuable experimental data to test current computational models and assumptions, and might facilitate the rational design of specifically acting inhibitors (1, 2).Techniques including NMR and EPR relaxation have been developed to incisively probe motions in the ensemble on different time scales, ranging from pic...

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“…Over the last decade PDS has been widely used to determine distances within doubly labeled oligonucleotide molecules in the range of 1.5-8 nm (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23). Further developments in PDS methods (3) enable even greater distances to be measured.…”

Section: Introductionmentioning

confidence: 98%

Pulse Dipolar ESR of Doubly Labeled Mini TAR DNA and Its Annealing to Mini TAR RNA

Sun

Borbat

Grigoryants

et al. 2015

Biophysical Journal

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Pulse dipolar electron-spin resonance in the form of double electron electron resonance was applied to strategically placed, site-specifically attached pairs of nitroxide spin labels to monitor changes in the mini TAR DNA stem-loop structure brought on by the HIV-1 nucleocapsid protein NCp7. The biophysical structural evidence was at Ångstrom-level resolution under solution conditions not amenable to crystallography or NMR. In the absence of complementary TAR RNA, double labels located in both the upper and the lower stem of mini TAR DNA showed in the presence of NCp7 a broadened distance distribution between the points of attachment, and there was evidence for several conformers. Next, when equimolar amounts of mini TAR DNA and complementary mini TAR RNA were present, NCp7 enhanced the annealing of their stem-loop structures to form duplex DNA-RNA. When duplex TAR DNA-TAR RNA formed, double labels initially located 27.5 Å apart at the 3'- and 5'-termini of the 27-base mini TAR DNA relocated to opposite ends of a 27 bp RNA-DNA duplex with 76.5 Å between labels, a distance which was consistent with the distance between the two labels in a thermally annealed 27-bp TAR DNA-TAR RNA duplex. Different sets of double labels initially located 26-27 Å apart in the mini TAR DNA upper stem, appropriately altered their interlabel distance to ~35 Å when a 27 bp TAR DNA-TAR RNA duplex formed, where the formation was caused either through NCp7-induced annealing or by thermal annealing. In summary, clear structural evidence was obtained for the fraying and destabilization brought on by NCp7 in its biochemical function as an annealing agent and for the detailed structural change from stem-loop to duplex RNA-DNA when complementary RNA was present.

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RNA dynamics: perspectives from spin labels

Cited by 54 publications

References 74 publications

Structural Analysis of Protein–RNA Complexes in Solution Using NMR Paramagnetic Relaxation Enhancements

Structural Analysis of Protein–RNA Complexes in Solution Using NMR Paramagnetic Relaxation Enhancements

From a structural average to the conformational ensemble of a DNA bulge

Pulse Dipolar ESR of Doubly Labeled Mini TAR DNA and Its Annealing to Mini TAR RNA

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