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Technische Entwicklungen in biomolekularer NMR Spektroskopie

English title Technical developments in biomolecular NMR Spectroscopy
Applicant Wider Gerhard
Number 140559
Funding scheme Project funding
Research institution Institut für Molekularbiologie und Biophysik ETH Zürich
Institution of higher education ETH Zurich - ETHZ
Main discipline Other disciplines of Physics
Start/End 01.09.2012 - 31.08.2013
Approved amount 64'062.00
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All Disciplines (3)

Discipline
Other disciplines of Physics
Physical Chemistry
Biophysics

Keywords (5)

NMR; assignment; projection; APSY; RNA

Lay Summary (English)

Lead
Lay summary

Summary: Three-dimensional structures of biological macromolecules form the basis for a detailed understanding of biological processes. Nuclear magnetic resonance (NMR) spectroscopy is one method widely used to obtain structural data. In this project we further develop methods that make the application of NMR more efficient.

Background: NMR spectroscopy in solution has found wide spread applications in physical, chemical, biological and medical sciences. A prerequisite for the detailed analysis of the information content of NMR spectra are individual assignments of resonances to specific nuclei (atoms). In biological applications, the necessary data is usually obtained from a number of multi-dimensional NMR experiments which correlate signals of 1H, 13C and 15N nuclei in doubly 13C,15N isotope-labeled macromolecules. Often, the data cannot be completely analyzed by automated computer programs and full resonance assignments can only be obtained interactively by an experienced NMR spectroscopist. Recently, a new technique was introduced: APSY (automated projection spectroscopy). APSY is based on projection NMR spectroscopy and it was shown to provide data that can reliably be analyzed by automated programs. For example, for protein spectra fully automated assignments of resonances of nuclei in the backbone and in aliphatic side chains could be obtained.

Goal: Based on the experience gained with the use of APSY with proteins we want to apply the APSY technique to another class of macromolecules: RNA and DNA. The assignment of resonances in RNA (DNA) is often challenging especially for automated routines because of the small dispersion of resonances in RNA (DNA) spectra. The application of APSY with denatured proteins has shown that this method can deal very well with poorly resolved spectra. Based on APSY correlation experiments in the bases, sugar and the oligonucleotide backbone, we will design a suite of APSY experiments for automated sequence-specific assignments of resonances in spectra of 13C,15N-labeled RNA (DNA).

Significance: The outcome of this work will provide experimental procedures for automated resonance assignment in spectra of biological macromolecules. The methodology will make the application of NMR spectroscopy with this class of molecules more efficient by alleviating a major bottleneck in the analysis of NMR data.

Direct link to Lay Summary Last update: 21.02.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
Automated NMR resonance assignment strategy for RNA via the phosphodiester backbone based on high-dimensional through-bond APSY experiments.
Krähenbühl Barbara, El Bakkali Issam, Schmidt Elena, Güntert Peter, Wider Gerhard (2014), Automated NMR resonance assignment strategy for RNA via the phosphodiester backbone based on high-dimensional through-bond APSY experiments., in Journal of biomolecular NMR, 59(2), 87-93.
Strategy for automated NMR resonance assignment of RNA: application to 48-nucleotide K10.
Krähenbühl Barbara, Lukavsky Peter, Wider Gerhard (2014), Strategy for automated NMR resonance assignment of RNA: application to 48-nucleotide K10., in Journal of biomolecular NMR, 59(4), 231-40.
4D experiments measured with APSY for automated backbone resonance assignments of large proteins
Krähenbühl Barbara, Boudet Julien, Wider Gerhard (2013), 4D experiments measured with APSY for automated backbone resonance assignments of large proteins, in J Biomol NMR, 56, 149-154.

Collaboration

Group / person Country
Types of collaboration
Prof. F. Allain / ETH-Z Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Bruker Biospin Switzerland (Europe)
- Industry/business/other use-inspired collaboration
Prof.S. Hiller / University of Basle Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
ANZMAG 2015 Talk given at a conference Automated Assignment Strategies Based on Through-bond and Through-space High Dimensional Experiments 29.11.2015 Bay of Island, New Zealand Wider Gerhard;
6th Asia-Pacific NMR Symposium Talk given at a conference Automated NMR Resonance Assignment Strategies Exemplified for Nucleic Acids 13.08.2015 Hongkong, China Wider Gerhard;
EUROMAR 2015 Talk given at a conference Automated NMR Resonance Assignment Strategies for Nucleic Acids using through-bond and through-space highdimensional Experiments 05.07.2015 Prague, Czech Republic Wider Gerhard;
NMR Winter Retreat of Protein-RNA Interactions Talk given at a conference APSY with RNA – A roundup 16.03.2014 Parpan, Switzerland Wider Gerhard;
7th NMR Winter Retreat of Protein-RNA Interactions Talk given at a conference APSY experiments on RNA 04.03.2013 Parpan, Switzerland Krähenbühl Barbara; Wider Gerhard;


Associated projects

Number Title Start Funding scheme
120048 Technische Entwicklunen in biomolekularer NMR Spektroskopie 01.06.2009 Project funding
144242 Combined use of NMR and DNA encoded libraries for drug discovery 01.12.2012 Project funding

Abstract

This project of one year duration is an extension of a three year project funded by the SNF. This extension will allow the PhD student working on the current project to complete her PhD degree. Most applications of nuclear magnetic resonance (NMR) spectroscopy in solution require individual assignments of resonances to specific nuclei (atoms) of the molecules under investigation. In biological applications, the necessary data is usually obtained from a number of multi-dimensional NMR experiments with doubly 13C and 15N isotope-labeled macromolecules. Often, these data cannot be fully analyzed by automated computer programs, and complete resonance assignments can only be obtained interactively by an experienced NMR spectroscopist. Recently, it was shown that APSY (automated projection spectroscopy), a method which is based on projection spectroscopy, can reliably provide fully automated assignments of resonance frequencies in protein NMR spectra. In this proposal we want to further develop the APSY methodology and extend its applicability. Based on the experience gained with the use of APSY with proteins we started to apply the APSY technique also to RNA molecules. These molecules produce NMR spectra with a small dispersion of signals, which makes assignments challenging for automated routines. Applications of APSY have shown that this method can deal very well with poorly resolved spectra. Based on APSY correlation experiments in the bases, sugar and the oligonucleotide backbone we want to develop a suite of APSY experiments for automated sequence-specific assignments of resonances in spectra of [13C,15N]-labeled RNA (DNA). The outcome of this work will provide experimental procedures for automated resonance assignment in RNA (DNA) spectra. The methodology will make the application of NMR spectroscopy with this class of molecules more efficient by alleviating a major bottleneck in the analysis of NMR data.
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