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Technische Entwicklunen in biomolekularer NMR Spektroskopie

English title Technical developments in biomolecular NMR Spectroscopy
Applicant Wider Gerhard
Number 120048
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 01.06.2009 - 31.08.2012
Approved amount 180'525.00
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All Disciplines (3)

Discipline
Other disciplines of Physics
Biophysics
Physical Chemistry

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 of the two methods widely used to obtain structural data. In this project we want to develop methods to make the application of NMR more efficient.

Background:Nuclear magnetic resonance (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 coherences 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.
Intramolecular Donor Strand Complementation in the E. coli Type 1 Pilus Subunit FimA Explains the Existence of FimA Monomers As Off-Pathway Products of Pilus Assembly That Inhibit Host Cell Apoptosis
Walczak Michal J., Puorger Chasper, Glockshuber Rudi, Wider Gerhard (2014), Intramolecular Donor Strand Complementation in the E. coli Type 1 Pilus Subunit FimA Explains the Existence of FimA Monomers As Off-Pathway Products of Pilus Assembly That Inhibit Host Cell Apoptosis, in JOURNAL OF MOLECULAR BIOLOGY, 426(3), 542-549.
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 Journal of biomolecular NMR, 56(2), 149-54.
Automated Projection Spectroscopy (APSY) for the Assignment of NMR Resonances of Biological Macromolecules
Krähenbühl B, Wider G (2012), Automated Projection Spectroscopy (APSY) for the Assignment of NMR Resonances of Biological Macromolecules, in CHIMIA, 66(10), 767-771.
Sugar-to-base correlation in nucleic acids with a 5D APSY-HCNCH or two 3D APSY-HCN experiments
Krahenbuhl B, Hofmann D, Maris C, Wider G (2012), Sugar-to-base correlation in nucleic acids with a 5D APSY-HCNCH or two 3D APSY-HCN experiments, in JOURNAL OF BIOMOLECULAR NMR, 52(2), 141-150.
4D APSY-HBCB(CG)CDHD experiment for automated assignment of aromatic amino acid side chains in proteins
Krahenbuhl B, Hiller S, Wider G (2011), 4D APSY-HBCB(CG)CDHD experiment for automated assignment of aromatic amino acid side chains in proteins, in JOURNAL OF BIOMOLECULAR NMR, 51(3), 313-318.
Automated Projection Spectroscopy and Its Applications
Hiller S, Wider G (2011), Automated Projection Spectroscopy and Its Applications, in Top. Curr. Chem., 316, 21-48.

Collaboration

Group / person Country
Types of collaboration
Universität Basel Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication

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 (APNMR 2015) 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 Prag, Czech Republic Wider Gerhard;
7th NMR Winter Retreat of Protein-RNA Interactions Talk given at a conference APSY with RNA – A roundup 16.03.2014 Parpan, Switzerland Wider Gerhard;
XXVth ICMRBS Poster Advances in Automated Projection Spectroscopy (APSY) 19.04.2012 Lyon, France Krähenbühl Barbara;
6th NMR Winter Retreat of Protein-RNA Interactions Talk given at a conference Advances in automated projection spectroscopy (APSY) 11.03.2012 Parpan, Schweiz, Switzerland Krähenbühl Barbara;
Indo Swiss NMR symposium Talk given at a conference Automated resonance assignments in biological macromolecules 23.01.2012 Pune, India Wider Gerhard;
Indian Biophysical Society Meeting Talk given at a conference Improved efficiency of NMR studies of biological macromolecules 19.01.2012 Chennai, India Wider Gerhard;
XIXth Swiss NMR Symposium Talk given at a conference Advances in automated projection spectroscopy (APSY) 17.01.2012 Bern, Switzerland Krähenbühl Barbara;
4th BSM retreat Talk given at a conference High dimensional NMR experimetns using projection spectroscopy 01.07.2011 Lungern, Switzerland Krähenbühl Barbara;
5th NMR Winter Retreat of Protein-RNA Interactions Talk given at a conference Protein and RNA NMR spectra assignment with automated projection spectroscopy (APSY) 06.03.2011 Parpan, Switzerland Krähenbühl Barbara;
5th NMR Winter Retreat of Protein-RNA Interactions Poster 5D APSY HCNCH and 3D APSY HCN experiments for 13C, 15N labeled RNA 06.03.2011 Parpan, Switzerland Krähenbühl Barbara;
3rd BSM retreat Poster Automated NMR Projection Spectroscopy Experiments for RNA Ribose-to-Base Correlation 13.08.2010 Ligerz, Switzerland Krähenbühl Barbara;
3rd Annual Protein & Peptide Conference (PepCon-2010) Talk given at a conference Efficient studies of protein-protein interactions by NMR spectroscopy 21.03.2010 Beijing, China Wider Gerhard;
Advanced Course in New Methods in Biomolecular NMR Talk given at a conference Automated projection spectroscopy for RNA 20.09.2009 Göteborg, Sweden Krähenbühl Barbara;
18th Swiss Symposium on NMR Spectroscopy Talk given at a conference Assignments and interactions 09.09.2009 Genf, Switzerland Wider Gerhard;
2nd BSM retreat Poster High dimensional NMR experimetns using pojection spectroscopy 04.09.2009 Filzbach, Switzerland Krähenbühl Barbara;


Associated projects

Number Title Start Funding scheme
139221 New electronics for 900MHz NMR spectrometer 01.12.2011 R'EQUIP
140559 Technische Entwicklungen in biomolekularer NMR Spektroskopie 01.09.2012 Project funding
113730 NMR in structural biology: protein structure determination and studies of protein-protein interactions 01.01.2007 Project funding

Abstract

Nuclear magnetic resonance (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 coherences of 1H, 13C and 15N nuclei in doubly 13C,15N isotope-labeled macromolecules. Often, the data cannot be analyzed by automated computer programs and full resonance assignments can only be obtained interactively by an experienced NMR spectroscopist. The aim of this research project is the development of efficient methods for NMR spectroscopy applied to biological macromolecules. Recently, in collaboration with Prof. Wüthrich (ETH-Z), we introduced a new technique, APSY (automated projection spectroscopy), which is based on projection NMR spectroscopy. APSY 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.In this proposal we want to further develop the APSY methodology and extend its applicability. In a first step, the available APSY techniques for resonance assignments in protein spectra will be complemented by APSY experiments for automated assignment of resonances of aromatic side chain nuclei. With the successful implementation of these experiments a set of APSY experiments will be available for complete and automated assignment of resonances in spectra of small and medium size proteins. For larger proteins transverse relaxation reduces the sensitivity of NMR spectra and usually these proteins are perdeuterated to reduce relaxation. We want to extend the applicability of the APSY technology to this class of proteins. For maximal sensitivity APSY experiments will be designed that use the concept of transverse relaxation optimizes spectroscopy (TROSY). The result of this development will be a set of APSY experiments for automated resonance assignment of nuclei in the polypeptide backbone (including the ?-carbon) of deuterated 13C,15N-labeled proteins.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). 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. We anticipate that this methodology progress will have a considerable impact on current biophysical research with NMR spectroscopy.
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