Abstract Molecular motion and dynamics play an essential role in the biological function of many RNAs. An important source of information on biomolecular motion can be found in residual dipolar couplings which contain dynamics information over the entire ms-ps timescale. However, these methods are not fully applicable to RNA because nucleic acid molecules tend to align in a highly collinear manner in different alignment media. As a consequence, information on dynamics that can be obtained with this method is limited. In order to overcome this limitation, we have generated a chimeric RNA containing both the wild type TAR RNA, the target of our investigation of dynamics, as well as the binding site for U1A protein. When U1A protein was bound to the portion of the chimeric RNA containing its binding site, we obtained independent alignment of TAR by exploiting the physical chemical characteristics of this protein. This technique can allow the extraction of new information on RNA dynamics, which is particularly important for time scales not covered by relaxation methods where important RNA motions occur.
Content Type Journal Article
Category Article
Pages 1-12
DOI 10.1007/s10858-012-9655-0
Authors
Michael F. Bardaro Jr., Department of Chemistry, University of Washington, Box 351700, Seattle, WA 98195, USA
Gabriele Varani, Department of Chemistry, University of Washington, Box 351700, Seattle, WA 98195, USA
Residual dipolar couplings: are multiple independent alignments always possible?
Residual dipolar couplings: are multiple independent alignments always possible?
Abstract RDCs for the 14 kDa protein hen egg-white lysozyme (HEWL) have been measured in eight different alignment media. The elongated shape and strongly positively charged surface of HEWL appear to limit the protein to four main alignment orientations. Furthermore, low levels of alignment and the proteinâ??s interaction with some alignment media increases the experimental error. Together with heterogeneity across the alignment media arising from constraints on temperature, pH and ionic strength for some...
nmrlearner
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12-26-2010 04:43 AM
[NMR paper] Sensitivity of NMR residual dipolar couplings to perturbations in folded and denature
Sensitivity of NMR residual dipolar couplings to perturbations in folded and denatured staphylococcal nuclease.
Related Articles Sensitivity of NMR residual dipolar couplings to perturbations in folded and denatured staphylococcal nuclease.
Biochemistry. 2005 May 3;44(17):6392-403
Authors: Sallum CO, Martel DM, Fournier RS, Matousek WM, Alexandrescu AT
The invariance of NMR residual dipolar couplings (RDCs) in denatured forms of staphylococcal nuclease to changes in denaturant concentration or amino acid sequence has previously been attributed...
nmrlearner
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11-25-2010 08:21 PM
[NMR paper] Residual dipolar couplings in NMR structure analysis.
Residual dipolar couplings in NMR structure analysis.
Related Articles Residual dipolar couplings in NMR structure analysis.
Annu Rev Biophys Biomol Struct. 2004;33:387-413
Authors: Lipsitz RS, Tjandra N
Residual dipolar couplings (RDCs) have recently emerged as a new tool in nuclear magnetic resonance (NMR) with which to study macromolecular structure and function in a solution environment. RDCs are complementary to the more conventional use of NOEs to provide structural information. While NOEs are local-distance restraints, RDCs provide...
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11-24-2010 09:25 PM
[NMR paper] Residual dipolar couplings: synergy between NMR and structural genomics.
Residual dipolar couplings: synergy between NMR and structural genomics.
Related Articles Residual dipolar couplings: synergy between NMR and structural genomics.
J Biomol NMR. 2002 Jan;22(1):1-8
Authors: Al-Hashimi HM, Patel DJ
Structural genomics is on a quest for the structure and function of a significant fraction of gene products. Current efforts are focusing on structure determination of single-domain proteins, which can readily be targeted by X-ray crystallography, NMR spectroscopy and computational homology modeling. However,...
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11-24-2010 08:49 PM
Facile measurement of 1Hâ??15N residual dipolar couplings in larger perdeuterated pro
Abstract We present a simple method, ARTSY, for extracting 1JNH couplings and 1Hâ??15N RDCs from an interleaved set of two-dimensional 1Hâ??15N TROSY-HSQC spectra, based on the principle of quantitative J correlation. The primary advantage of the ARTSY method over other methods is the ability to measure couplings without scaling peak positions or altering the narrow line widths characteristic of TROSY spectra. Accuracy of the method is demonstrated for the model system GB3. Application to the catalytic core domain of HIV integrase, a 36 kDa homodimer with unfavorable spectral...
nmrlearner
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08-14-2010 04:19 AM
Theoretical framework for NMR residual dipolar couplings in unfolded proteins
Theoretical framework for NMR residual dipolar couplings in unfolded proteins
O. I. Obolensky, Kai Schlepckow, Harald Schwalbe and A. V. Solov’yov
Journal of Biomolecular NMR; 2007; 39(1) pp 1-16
Abstract:
A theoretical framework for the prediction of nuclear magnetic resonance (NMR) residual dipolar couplings (RDCs) in unfolded proteins under weakly aligning conditions is presented. The unfolded polypeptide chain is modeled as a random flight chain while the alignment medium is represented by a set of regularly arranged obstacles. For the case of bicelles oriented perpendicular to...
stewart
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08-05-2008 02:26 AM
De novo determination of internuclear vector orientations from residual dipolar couplings measured in three independent alignment media
De novo determination of internuclear vector orientations from residual dipolar couplings measured in three independent alignment media
Ke Ruan, Kathryn B. Briggman and Joel R. Tolman
Journal of Biomolecular NMR; 2008; 41(2) pp 61 - 76
Abstract:
The straightforward interpretation of solution state residual dipolar couplings (RDCs) in terms of internuclear vector orientations generally requires prior knowledge of the alignment tensor, which in turn is normally estimated using a structural model. We have developed a protocol which allows the requirement for prior structural knowledge to...
daniel
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08-03-2008 03:54 AM
Composite Alignment Media for the Measurement of Independent Sets of NMR Residual Dipolar Couplings
Composite Alignment Media for the Measurement of Independent Sets of NMR Residual Dipolar Couplings
Ke Ruan and Joel R. Tolman
J. Am. Chem. Soc.; 2005; 127(43) pp 15032 - 15033;
Abstract:
The measurement of independent sets of NMR residual dipolar couplings (RDCs) in multiple alignment media can provide a detailed view of biomolecular structure and dynamics, yet remains experimentally challenging. It is demonstrated here that independent sets of RDCs can be measured for ubiquitin using just a single alignment medium composed of aligned bacteriophage Pf1 particles embedded in a...
Independent alignment of RNA for dynamic studies using residual dipolar couplings
Linear Mode
