Project

Back to overview

Biomacromolecular structures, dynamics and interactions by NMR and new developments in NMR technology

English title Biomacromolecular structures, dynamics and interactions by NMR and new developments in NMR technology
Applicant Grzesiek Stephan
Number 149927
Funding scheme Project funding (Div. I-III)
Research institution Abteilung Strukturbiologie und Biophysik Biozentrum Universität Basel
Institution of higher education University of Basel - BS
Main discipline Biophysics
Start/End 01.10.2013 - 31.03.2017
Approved amount 809'988.00
Show all

Keywords (11)

protein interaction; GPCR; dynamics; HIV; protein; Abelson kinase; nuclear magnetic resonance; cancer; structure; protein folding; c-di-GMP

Lay Summary (German)

Lead
Trotz enormer Fortschritte im Wissen um Gensequenzen, Proteinstrukturen und Proteinfunktion ist es unklar wie eine Gensequenz eine Proteinstruktur codiert und die Struktur die Funktion bestimmt. Es ist deshalb nötig, Methoden zu entwickeln, um die Lücken zwischen diesen Bereichen von Information über Biomoleküle schliessen. Kernspinresonanz ist dafür ein einzigartiges Werkzeug, da sie atomare Information über Struktur und Dynamik von Biomolekülen unter fast natürlichen Bedingungen liefert.
Lay summary

Inhalt und Ziel

Ziel dieses Projektes ist die speziellen Stärken der Kernspinresonanz (NMR)-Methode anzuwenden und weiterzuentwickeln, um allgemeine Prinzipien von Proteinstrukturfunktionsbeziehungen abzuleiten.

Teilprojekt A zielt auf die Bestimmung von Struktur, Dynamik und Wechselwirkungen von vier biolomolekularen Systemen, für die wir in der letzten Periode Grundlagen erarbeitet und viele Fortschritte erzielt haben: (1) Abl-Kinase, ein Zielmolekül bei der Behandlung von CML-Leukämie. (2) der G-Protein-gekoppelte Rezeptor und HIV-Korezeptor CCR5 und seine Wechselwirkung mit dem Chemokin RANTES. (3) Wechselwirkungen des bakteriellen Virulenzfaktors zyklisches di-GMP. (4) Lipopolysacharid, der Auslöser des endotoxischen Schocks und seine Wechselwirkungen mit dem lipopolysaccharide binding protein.

Teilprojekt B zielt auf die Charakterisierung von ungefalteten Proteinen und ein Verständnis ihrer Beziehung zur gefalteten Struktur. In der vorangegangenen Periode ist es uns gelungen, die Entfaltung eines Modellproteins durch Kombination von hohem Druck und tiefen Temperaturen oder hohem Druck und Alkohol zu induzieren und NMR-spektroskopisch mit atomarer Auflösung zu verfolgen. Diese Experimente sollen nun auf weitere Proteine ausgedehnt werden. Ausserdem haben wir eine Methode entwickelt, um strukturelle Ensembles von ungefalteten Proteinen direkt aus verschiedenen NMR-Daten zu berechnen. Diese Methode soll nun auf chemische Shifts und andere nicht-NMR-Daten ausgeweitet werden, um die Untersuchung einer grösseren Zahl von Proteinen zu ermöglichen.

Kontext

Unsere Arbeit soll grundlegende Zusammenhänge zwischen der Gen/Aminosäuresequenz eines Proteins, seiner Struktur und seiner Funktion aufklären. Dies ist einerseits von hoher allgemeiner Bedeutung für das molekulare Verständnis biologischer Vorgänge und kann andererseits aber auch wichtige Informationen für die Entwicklung neuer Medikamente, z.B. gegen Krebs, AIDS und bakterielle Infektionen, liefern.

Direct link to Lay Summary Last update: 17.10.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
Comprehensive structural and dynamical view of an unfolded protein from the combination of single-molecule FRET, NMR, and SAXS.
Aznauryan Mikayel, Delgado Leonildo, Soranno Andrea, Nettels Daniel, Huang Jie-Rong, Labhardt Alexander M, Grzesiek Stephan, Schuler Benjamin (2016), Comprehensive structural and dynamical view of an unfolded protein from the combination of single-molecule FRET, NMR, and SAXS., in Proceedings of the National Academy of Sciences of the United States of America, 113(37), 5389-98.
Backbone NMR reveals allosteric signal transduction networks in the β1-adrenergic receptor.
Isogai Shin, Deupi Xavier, Opitz Christian, Heydenreich Franziska M, Tsai Ching-Ju, Brueckner Florian, Schertler Gebhard F X, Veprintsev Dmitry B, Grzesiek Stephan (2016), Backbone NMR reveals allosteric signal transduction networks in the β1-adrenergic receptor., in Nature, 530(7589), 237-41.
A Rigorous and Efficient Method To Reweight Very Large Conformational Ensembles Using Average Experimental Data and To Determine Their Relative Information Content.
Leung Hoi Tik Alvin, Bignucolo Olivier, Aregger Regula, Dames Sonja A, Mazur Adam, Bernèche Simon, Grzesiek Stephan (2016), A Rigorous and Efficient Method To Reweight Very Large Conformational Ensembles Using Average Experimental Data and To Determine Their Relative Information Content., in Journal of chemical theory and computation, 12(1), 383-94.
Unraveling the complexity of protein backbone dynamics with combined (13)C and (15)N solid-state NMR relaxation measurements.
Lamley Jonathan M, Lougher Matthew J, Sass Hans Juergen, Rogowski Marco, Grzesiek Stephan, Lewandowski Józef R (2015), Unraveling the complexity of protein backbone dynamics with combined (13)C and (15)N solid-state NMR relaxation measurements., in Physical chemistry chemical physics : PCCP, 17(34), 21997-2008.
An economic approach to efficient isotope labeling in insect cells using homemade 15N-, 13C- and 2H-labeled yeast extracts.
Opitz Christian, Isogai Shin, Grzesiek Stephan (2015), An economic approach to efficient isotope labeling in insect cells using homemade 15N-, 13C- and 2H-labeled yeast extracts., in Journal of biomolecular NMR, 62(3), 373-85.
Backbone hydration determines the folding signature of amino acid residues.
Bignucolo Olivier, Leung Hoi Tik Alvin, Grzesiek Stephan, Bernèche Simon (2015), Backbone hydration determines the folding signature of amino acid residues., in Journal of the American Chemical Society, 137(13), 4300-3.
Structural basis and dynamics of multidrug recognition in a minimal bacterial multidrug resistance system.
Habazettl Judith, Allan Martin, Jensen Pernille Rose, Sass Hans-Jürgen, Thompson Charles J, Grzesiek Stephan (2014), Structural basis and dynamics of multidrug recognition in a minimal bacterial multidrug resistance system., in Proceedings of the National Academy of Sciences of the United States of America, 111(51), 5498-507.
Solid-state NMR of a protein in a precipitated complex with a full-length antibody.
Lamley Jonathan M, Iuga Dinu, Öster Carl, Sass Hans-Juergen, Rogowski Marco, Oss Andres, Past Jaan, Reinhold Andres, Grzesiek Stephan, Samoson Ago, Lewandowski Józef R (2014), Solid-state NMR of a protein in a precipitated complex with a full-length antibody., in Journal of the American Chemical Society, 136(48), 16800-6.
WhiB7, an Fe-S-dependent transcription factor that activates species-specific repertoires of drug resistance determinants in actinobacteria.
Ramón-García Santiago, Ng Carol, Jensen Pernille R, Dosanjh Manisha, Burian Jan, Morris Rowan P, Folcher Marc, Eltis Lindsay D, Grzesiek Stephan, Nguyen Liem, Thompson Charles J (2013), WhiB7, an Fe-S-dependent transcription factor that activates species-specific repertoires of drug resistance determinants in actinobacteria., in The Journal of biological chemistry, 288(48), 34514-28.
NMR reveals the allosteric opening and closing of Abelson tyrosine kinase by ATP-site and myristoyl pocket inhibitors.
Skora Lukasz, Mestan Jürgen, Fabbro Doriano, Jahnke Wolfgang, Grzesiek Stephan (2013), NMR reveals the allosteric opening and closing of Abelson tyrosine kinase by ATP-site and myristoyl pocket inhibitors., in Proceedings of the National Academy of Sciences of the United States of America, 110(47), 4437-45.
Biophysical and structural investigation of bacterially expressed and engineered CCR5, a G protein-coupled receptor.
Wiktor Maciej, Morin Sébastien, Sass Hans-Jürgen, Kebbel Fabian, Grzesiek Stephan (2013), Biophysical and structural investigation of bacterially expressed and engineered CCR5, a G protein-coupled receptor., in Journal of biomolecular NMR, 55(1), 79-95.

Collaboration

Group / person Country
Types of collaboration
Prof. Jozef Lewandowski/U. Warwick Great Britain and Northern Ireland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Research Infrastructure
Dr. Wolfgang Jahnke/Novartis Basel Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Prof. Lyndon Emsley/ENS Lyon France (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Dr. Martin Blackledge/IBS Grenoble France (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Prof. Ben Schuler/U. Zuerich Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Research Infrastructure
Prof. Claus Seidel/U. Duesseldorf Germany (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Research Infrastructure
Prof. Gebhard Schertler/Paul Scherrer Institut Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
Prof. Henning Stahlberg/U. Basel Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Research Infrastructure
Prof. Oliver Hantschel/EPFL Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
Prof. U. Zaehringer/FZ Borstel Germany (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Prof. Cheryl Arrowsmith, U. Toronto Canada (North America)
- in-depth/constructive exchanges on approaches, methods or results

Scientific events

Awards

Title Year
Guest Professorship by the Japan Gateway Program at Kyoto University 2015
Honorary Member of the National Magnetic Resonance Society of India 2014

Associated projects

Number Title Start Funding scheme
132857 Biomacromolecular structures, dynamics and interactions by NMR and new developments in NMR technology 01.10.2010 Project funding (Div. I-III)
141898 NMR studies of GPCRs: Structure, dynamics and interactions with ligands and signaling proteins 01.12.2012 Sinergia
173089 Biomacromolecular structures, dynamics and interactions by NMR and new developments in NMR technology 01.04.2017 Project funding (Div. I-III)
150814 Purchase of a 900 MHz high-resolution NMR instrument 01.12.2013 R'EQUIP

Abstract

Biological function results from time-dependent interactions between biomolecules. It is ultimately encoded within the primary chemical structure of molecules resulting in defined three-dimensional atom positions and movements. NMR spectroscopy is the only experimental method, which yields both structural and dynamical information on biomolecules at atomic resolution with minimal invasiveness and at close to natural conditions. As such it can provide unique information to understand the connection between primary structure, tertiary structure, dynamics and function. It is the goal of this proposal to apply and further develop these strengths of NMR technology with the aim to reveal general principles of protein structure function relations. The proposal is divided into two subprojects:Subproject A is directed towards the determination of structure, dynamics, and interactions in four medically important systems, for which we have made significant progress in recent years: (1) Abl kinase, which is an important leukemia drug target. We have obtained assignment of a large, multidomain Abl construct that is the minimal autoregulatory fragment. Our data show that binding of different classes of inhibitors induces distinct domain rearrangements, which shed light on the mechanism of kinase regulation. We now want to reveal the atomic causes of these allosteric rearrangements, investigate several medically and functionally relevant mutants, and extend the studies to single molecule FRET. (2) the HIV-1 coreceptor and G-protein coupled receptor (GPCR) CCR5. We have obtained large-scale functional expression of CCR5 in insect cells and E. coli, which permits effective isotope labeling. We seek to apply new solid-state and solution NMR techniques for structure determination and to develop general strategies for GPCR analysis by a comparison to other available GPCRs. (3) Interactions of the bacterial virulence factor cyclic di-guanosine-monophosphate (c-di-GMP). We have determined the structure of the PilZ homolog PA4608 in complex with c-di-GMP and the kinetics of c-di-GMP oligomer formation in solution. We now want to solve the structures of a complex c-di-GMP with the chemotactic protein CC3300 and of higher c-di-GMP oligomers. (4) Lipopolysaccharide (LPS), the causative agent of endotoxic shock. We have developed a method to study LPS by solution NMR and obtained expression of the lipopolysaccharide binding protein, which is the first receptor of the endotoxic recognition cascade. We intend to study its interaction with LPS.Subproject B is directed towards the NMR characterization of unfolded states of proteins and their relation to the folded structure with an emphasis on pressure denaturation and structural modeling of unfolded ensembles. The overarching goal is to rationalize protein folding by correlating high resolution experimental data on unfolded states with primary, secondary and tertiary structure information. (1) We have obtained unique data on the pressure/cold-denatured state of ubiquitin, which shows similar subpopulations of partial native structures as an alcohol-denatured state. By addition of non-denaturing concentrations of alcohol, full pressure-induced unfolding can be achieved at room temperature in a completely reversible way. This allows following the unfolding transition at very high resolution. We want to extend this analysis to a larger set of about 10 proteins representing different folds and sequences with the aim to correlate unfolding behavior with sequence or structure and to understand the nature of pressure unfolding. (2) We have developed an effective method to calculate structural ensembles of unfolded proteins from large sets of NMR data. We want to continue these efforts with the aim to include FRET and chemical shift data in order to obtain highly defined quantitative models for larger sets of unfolded proteins.
-