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16 results on '"Dept Chem Ugo Schiff"'

1. Metabolite and lipoprotein profiles reveal sex-related oxidative stress imbalance in de novo drug-naive Parkinson's disease patients

Author

Meoni, Gaia, Tenori, Leonardo, Schade, Sebastian, Licari, Cristina, Pirazzini, Chiara, Bacalini, Maria Giulia, Garagnani, Paolo, Turano, Paola, Baldelli, Luca, Jesús, Silvia, Schreglmann, Sebastian R, Sambati, Luisa, Gómez-Garre, Pilar, Halsband, Claire, Calandra-Buonaura, Giovanna, Adarmes-Gómez, Astrid Daniela, Sixel-Döring, Friederike, Zenesini, Corrado, Bhatia, Kailash P., Cortelli, Pietro, Mollenhauer, Brit, Franceschi, Claudio, Houlden, Henry, Liò, Pietro, Luchinat, Claudio, Delledonne, Massimo, Mills, Kevin, Pedersen, Nancy L., Azevedo, Tiago, Bartoletti-Stella, Anna, Bonilla-Toribio, Marta, Buiza-Rueda, Dolores, Capellari, Sabina, Carriòn-Claro, Mario, Clayton, Robert, Dal Molin, Alessandra, Dimitri, Giovanna Maria, Doykov, Ivan, Giuliani, Cristina, Hägg, Sara, Hällqvist, Jenny, Heywood, Wendy, Huertas, Ismael, Jylhävä, Juulia, Labrador-Espinosa, Miguel A., Macias, Daniel, Magrinelli, Francesca, Rodríguez, Juan Francisco Martín, Maturo, Maria Giovanna, Mengozzi, Giacomo, Milazzo, Maddalena, Nardini, Christine, Periñán-Tocino, Maria Teresa, Ravaioli, Francesco, Sala, Claudia, Spasov, Simeon, Tejera-Parrado, Cristina, Paola, Turano, Williams, Dylan, Xumerle, Luciano, Trenkwalder, Claudia, European Commission, Meoni, Gaia, Tenori, Leonardo, Schade, Sebastian, Turano, Paola, Licari, Cristina, Pirazzini, Chiara, Bacalini, Maria Giulia, Garagnani, Paolo, PROPAG-AGEING Consortium[.., Giuliani, Cristina, Provini, Federica, Calandra Buonaura, Giovanna, Cortelli, Pietro, Capellari, Sabina, ], Trenkwalder, Claudia, Franceschi, Claudio, Mollenhauer, Brit, Luchinat, Claudio, [Meoni, Gaia] Univ Florence, Magnet Resonance Ctr CERM, Florence, Italy, [Tenori, Leonardo] Univ Florence, Magnet Resonance Ctr CERM, Florence, Italy, [Licari, Cristina] Univ Florence, Magnet Resonance Ctr CERM, Florence, Italy, [Turano, Paola] Univ Florence, Magnet Resonance Ctr CERM, Florence, Italy, [Luchinat, Claudio] Univ Florence, Magnet Resonance Ctr CERM, Florence, Italy, [Meoni, Gaia] Univ Florence, Dept Chem Ugo Schiff, Florence, Italy, [Tenori, Leonardo] Univ Florence, Dept Chem Ugo Schiff, Florence, Italy, [Licari, Cristina] Univ Florence, Dept Chem Ugo Schiff, Florence, Italy, [Turano, Paola] Univ Florence, Dept Chem Ugo Schiff, Florence, Italy, [Luchinat, Claudio] Univ Florence, Dept Chem Ugo Schiff, Florence, Italy, [Tenori, Leonardo] Consorzio Interuniv Risonanze Magnet Met Prot CIR, Florence, Italy, [Turano, Paola] Consorzio Interuniv Risonanze Magnet Met Prot CIR, Florence, Italy, [Luchinat, Claudio] Consorzio Interuniv Risonanze Magnet Met Prot CIR, Florence, Italy, [Schade, Sebastian] Univ Med Ctr Goettingen, Dept Clin Neurophysiol, Gottingen, Germany, [Pirazzini, Chiara] IRCCS Ist Sci Neurol Bologna, Bologna, Italy, [Bacalini, Maria Giulia] IRCCS Ist Sci Neurol Bologna, Bologna, Italy, [Garagnani, Paolo] Univ Bologna, Dept Expt Diagnost & Specialty Med DIMES, Bologna, Italy, [Franceschi, Claudio] Univ Bologna, Dept Expt Diagnost & Specialty Med DIMES, Bologna, Italy, [Trenkwalder, Claudia] Univ Med Ctr Goettingen, Dept Neurol & Paracelsus Elena Klin, Kassel, Germany, [Mollenhauer, Brit] Univ Med Ctr Goettingen, Dept Neurol & Paracelsus Elena Klin, Kassel, Germany, [Franceschi, Claudio] Lobachevsky Univ, Lab Syst Med Hlth Aging, Nizhnii Novgorod, Russia, [Franceschi, Claudio] Lobachevsky Univ, Dept Appl Math, Nizhnii Novgorod, Russia, Instruct-ERIC, a Landmark ESFRI project, CERM/CIRMMP Italy Centre, Horizon 2020 Framework Programme (PROPAG-AGEING), Meoni, Gaia [0000-0002-8608-4641], Tenori, Leonardo [0000-0001-6438-059X], Schade, Sebastian [0000-0002-6316-6804], Turano, Paola [0000-0002-7683-8614], Franceschi, Claudio [0000-0001-9841-6386], Luchinat, Claudio [0000-0003-2271-8921], and Apollo - University of Cambridge Repository

Subjects
Risk, Metabolites, ipoproteins, sex-related, oxidative stress imbalance, Parkinson’s disease, NMR, Parkinson's disease, Serum-cholesterol, Predictive markers, Levodopa, Cellular and Molecular Neuroscience, Plasma, RC346-429, Homocysteine, Cholesterol levels, PROPAG-AGEING Consortium, Diagnostic markers, metabolomics, Nmr, Phenotypes, Neurology, PD, Neurology. Diseases of the nervous system, Neurology (clinical), Biomarkers
Abstract

Parkinson's disease (PD) is the neurological disorder showing the greatest rise in prevalence from 1990 to 2016. Despite clinical definition criteria and a tremendous effort to develop objective biomarkers, precise diagnosis of PD is still unavailable at early stage. In recent years, an increasing number of studies have used omic methods to unveil the molecular basis of PD, providing a detailed characterization of potentially pathological alterations in various biological specimens. Metabolomics could provide useful insights to deepen our knowledge of PD aetiopathogenesis, to identify signatures that distinguish groups of patients and uncover responsive biomarkers of PD that may be significant in early detection and in tracking the disease progression and drug treatment efficacy. The present work is the first large metabolomic study based on nuclear magnetic resonance (NMR) with an independent validation cohort aiming at the serum characterization of de novo drug-naive PD patients. Here, NMR is applied to sera from large training and independent validation cohorts of German subjects. Multivariate and univariate approaches are used to infer metabolic differences that characterize the metabolite and the lipoprotein profiles of newly diagnosed de novo drug-naive PD patients also in relation to the biological sex of the subjects in the study, evidencing a more pronounced fingerprint of the pathology in male patients. The presence of a validation cohort allowed us to confirm altered levels of acetone and cholesterol in male PD patients. By comparing the metabolites and lipoproteins levels among de novo drug-naive PD patients, age- and sex-matched healthy controls, and a group of advanced PD patients, we detected several descriptors of stronger oxidative stress., This work was supported by the Horizon 2020 Framework Programme (grant number 634821, PROPAG-AGEING).

Published
2022

2. Phase transition and amyloid formation by a viral protein as an additional molecular mechanism of virus-induced cell toxicity

Author

C. Debarnot, Vincent Delauzun, Sonia Longhi, Roberta Pierattelli, Silvia Spinelli, Christophe Bignon, Priscila Sutto-Ortiz, Maria Grazia Murrali, Edoardo Salladini, Architecture et fonction des macromolécules biologiques (AFMB), Institut National de la Recherche Agronomique (INRA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Centre de recherche en neurobiologie - neurophysiologie de Marseille (CRN2M), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Bioénergétique et Ingénierie des Protéines (BIP ), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Dept Chem Ugo Schiff, University of Florence (UNIFI), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Institut National de la Recherche Agronomique (INRA), Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), and Università degli Studi di Firenze = University of Florence (UniFI)

Subjects
0303 health sciences, Innate immune system, Amyloid, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, Viral protein, 030302 biochemistry & molecular biology, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Transfection, medicine.disease_cause, biology.organism_classification, Virus, Cell biology, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], [SDV.BBM.BP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biophysics, 03 medical and health sciences, DDB1, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], medicine, Hendra Virus, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], 030304 developmental biology, Henipavirus
Abstract

Henipaviruses are severe human pathogens responsible for severe encephalitis. Their V protein is a key player in the evasion of the host innate immune response. We have previously reported a biophysical characterization of the Henipavirus V proteins and shown that they interact with DDB1, a cellular protein that is a component of the ubiquitin ligase E3 complex. Here, we serendipitously discovered that the Hendra virus V protein undergoes a liquidhydrogel phase transition. By combining experimental and bioinformatics approaches, we have identified the V region responsible for this phenomenon. This region (referred to as PNT3), which falls within the long intrinsically disordered region of V, was further investigated using a combination of biophysical and structural approaches. ThioflavinT and Congo red binding assays, together with negative-staining electron microscopy studies, show that this region forms amyloid-like, β-enriched structures. Such structures are also formed in mammal cells transfected to express PNT3. Those cells also exhibit a reduced viability in the presence of a stress agent. Interestingly, mammal cells expressing a rationally designed, non-amyloidogenic PNT3 variant (PNT33A), appear to be much less sensitive to the stress agent, thus enabling the establishment of a link between fibril formation and cell toxicity. The present findings therefore pinpoint a so far never reported possible mechanism of virus-induced cell toxicity.

Published
2019

3. Paramagnetic Properties of a Crystalline Iron-Sulfur Protein by Magic-Angle Spinning NMR Spectroscopy

Author

Giacomo Parigi, Andrea Bertarello, Lyndon Emsley, Carmelo Fuccio, Enrico Ravera, Marco Fragai, Claudio Luchinat, Guido Pintacuda, Tobias Schubeis, Biological Solid-State NMR Methods - Méthodes de RMN à l'état solide en biologie, Institut des Sciences Analytiques (ISA), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Giotto Biotech SRL, Magnet Resonance Ctr CERM & Interunivers Consoriu, Dept Chem Ugo Schiff, Università degli Studi di Firenze = University of Florence (UniFI), Institut des Sciences et Ingénierie Chimiques (ISIC), This research was supported by the CNRS (IR-RMN FR3050), the Fondazione Cassa di Risparmio di Firenze, the MIUR PRIN 2012SK7ASN, the People Programme of the European Union's FP7 (FP7-PEOPLE-2012-ITN no. 317127 'pNMR'), the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation programme (grant no. 648974 'P-MEM-NMR'), the European Commission projects BioMedBridges No. 284209, iNEXT No. 653706, and Instruct, part of the European Strategy Forum on Research Infrastructures (ESFRI). Specifically, we thank the EU ESFRI Instruct Core Centre CERM, Italy. E.R. holds a FIRC triennial fellowship 'Gino Mazzega and Guglielmina Lucatello' (17941)., European Project: 317127,EC:FP7:PEOPLE,FP7-PEOPLE-2012-ITN,PNMR(2013), European Project: 653706,H2020,H2020-INFRAIA-2014-2015,iNEXT(2015), European Project: 648974,H2020,ERC-2014-CoG,P-MEM-NMR(2015), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), and Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI)

Subjects
Iron-Sulfur Proteins, chemistry.chemical_classification, 010405 organic chemistry, Chemistry, Nuclear magnetic resonance spectroscopy, 010402 general chemistry, 01 natural sciences, 0104 chemical sciences, Inorganic Chemistry, Crystallography, Paramagnetism, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, Magic angle spinning, Cluster (physics), Metalloprotein, Irradiation, Physical and Theoretical Chemistry, Crystallization, Nuclear Magnetic Resonance, Biomolecular, Spinning, Cysteine
Abstract

International audience; We present the first solid-state NMR study of an iron sulfur protein. The combined use of very fast (60 kHz) magic-angle spinning and tailored radiofrequency irradiation schemes allows the detection and the assignment of most of the H-1 and C-13 resonances of the oxidized high-potential iron sulfur protein I from Ectothiorhodospira halophila (EhHiPIP I), including those in residues coordinating the Fe4S4 cluster. For these residues, contact shifts as large as 100 and 400 ppm for H-1 and C-13 resonances, respectively, were observed, which represent the most shifted solid-state NMR signals ever measured in metalloproteins. Interestingly, by targeting EhHiPIP I in a crystalline environment, we were able to capture distinct paramagnetic signatures from the two conformations present in the asymmetric unit. The magnetic properties of the system were verified by following the temperature dependence of the contact-shifted cysteine resonances.

Published
2017
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4. Magic Angle Spinning NMR of Paramagnetic Proteins

Author

Lyndon Emsley, Roberta Pierattelli, Michael J. Knight, Guido Pintacuda, Isabella C. Felli, ISA - Centre de RMN à très hauts champs, Institut des Sciences Analytiques (ISA), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Dept Chem Ugo Schiff, Università degli Studi di Firenze = University of Florence (UniFI), Magnet Resonance Ctr CERM, Agence Nationale de la Recherche (ANR 08-BLAN-0035-01 , 10-BLAN-713-01), Ente Cassa di Risparmio di Firenze, Egide (programmes Galiee 22397RJ , 26000XF), the Universita Italo-francese (programma Galileo 2011/2012, Progetto 26000XF), Joint Research Activity and Access to Research Infrastructures in the 7th Framework program of the EC (BioNMR no. 261863)., ANR-08-BLAN-0035,PARA-NMR,Structure and reactivity of paramagnetic molecules by solid-state NMR(2008), European Project, Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), and Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI)

Subjects
Models, Molecular, Magnetic Resonance Spectroscopy, Carbon-13 NMR satellite, 010402 general chemistry, 01 natural sciences, SENSITIVITY ENHANCEMENT, Magnetics, Paramagnetism, LATTICE-RELAXATION, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, Metalloproteins, Magic angle spinning, SUPEROXIDE-DISMUTASE, Molecule, Spectroscopy, SPECTROSCOPY, 010405 organic chemistry, Chemistry, Proteins, CRYSTALLINE PROTEIN, General Medicine, General Chemistry, Nuclear magnetic resonance spectroscopy, QUANTITATIVE-ANALYSIS, SOLID-STATE NMR, 0104 chemical sciences, RANGE STRUCTURAL RESTRAINTS, [CHIM.THEO]Chemical Sciences/Theoretical and/or physical chemistry, Crystallography, RESOLUTION, Unpaired electron, Solid-state nuclear magnetic resonance, BACKBONE DYNAMICS, Condensed Matter::Strongly Correlated Electrons
Abstract

International audience; Metal ions are ubiquitous in biochemical and cellular processes. Since many metal ions are paramagnetic due to the presence of unpaired electrons, paramagnetic molecules are an important class of targets for research in structural biology and related fields. Today, NMR spectroscopy plays a central role in the investigation of the structure and chemical properties of paramagnetic metalloproteins, linking the observed paramagnetic phenomena directly to electronic and molecular structure. A major step forward in the study of proteins by solid-state NMR came with the advent of ultrafast magic angle spinning (MAS) and the ability to use (1)H detection. Combined, these techniques have allowed investigators to observe nuclei that previously were invisible in highly paramagnetic metalloproteins. In addition, these techniques have enabled quantitative site-specific measurement of a variety of long-range paramagnetic effects. Instead of limiting solid-state NMR studies of biological systems, paramagnetism provides an information-rich phenomenon that can be exploited in these studies. This Account emphasizes state-of-the-art methods and applications of solid-state NMR in paramagnetic systems in biological chemistry. In particular, we discuss the use of ultrafast MAS and (1)H-detection in perdeuterated paramagnetic metalloproteins. Current methodology allows us to determine the structure and dynamics of metalloenzymes, and, as an example, we describe solid-state NMR studies of microcrystalline superoxide dismutase, a 32 kDa dimer. Data were acquired with remarkably short times, and these experiments required only a few milligrams of sample.

Published
2013
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5. Rapid Measurement of Pseudocontact Shifts in Metalloproteins by Proton-Detected Solid-State NMR Spectroscopy

Author

Isabella C. Felli, Lyndon Emsley, Roberta Pierattelli, Guido Pintacuda, Michael J. Knight, Ivano Bertini, Torsten Herrmann, ISA - Centre de RMN à très hauts champs, Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Dept Chem Ugo Schiff, Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), Magnet Resonance Ctr CERM, Agence Nationale de la Recherche (ANR08-BLAN-0035-01) (ANR10-BLAN-713-01), Ente Cassa di Risparmio di Firenze, Egide (Galiee 22397RJ) (26000XF), Universita Italo Francese (26000XF), Joint Research Activity, EC (EAST-NMR 228461) (BioNMR 261863), European Project, Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), and Università degli Studi di Firenze = University of Florence (UniFI)

Subjects
DYNAMICS, IONS, Models, Molecular, Magnetic Resonance Spectroscopy, Proton, ASSIGNMENT, Analytical chemistry, PROTEIN, RELAXATION, Dihedral angle, 010402 general chemistry, 01 natural sciences, Biochemistry, Catalysis, Ion, 03 medical and health sciences, Paramagnetism, ENHANCEMENT, Colloid and Surface Chemistry, Nuclear magnetic resonance, DOMAIN, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, Metalloproteins, SUPEROXIDE-DISMUTASE, Spectroscopy, 030304 developmental biology, 0303 health sciences, PARAMAGNETIC METALLOPROTEINS, Superoxide Dismutase, Chemistry, Relaxation (NMR), Cobalt, General Chemistry, Reference Standards, 0104 chemical sciences, [CHIM.THEO]Chemical Sciences/Theoretical and/or physical chemistry, Solid-state nuclear magnetic resonance, Diamagnetism, COMPLEXES, Protons
Abstract

International audience; Pseudocontact shifts (PCSs) arise in para-magnetic systems in which the susceptibility tensor is anisotropic. PCSs depend upon the distance from the paramagnetic center and the position relative to the susceptibility tensor, and they can be used as structural restraints in protein structure determination. We show that the use of H-1-detected solid-state correlations provides facile and rapid detection and assignment of site-specific PCSs, including resolved H-1 PCSs, in a large metalloprotein, Co2+-substituted superoxide dismutase (Co2+-SOD). With only 3 mg of sample and a small set of experiments, several hundred PCSs were measured and assigned, and these PCSs were subsequently used in combination with H-1-H-1 distance and dihedral angle restraints to determine the protein backbone geometry with a precision paralleling those of state-of-the-art liquid-state determinations of diamagnetic proteins, including a well-defined active site.

Published
2012
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6. Longitudinal relaxation properties of (1)H(N) and (1)H(α) determined by direct-detected (13)C NMR experiments to study intrinsically disordered proteins (IDPs)

Author

Sergi Gil-Caballero, Bernhard Brutscher, Tomáš Hošek, Isabella C. Felli, Roberta Pierattelli, Magnet Resonance Ctr CERM, Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), Dept Chem Ugo Schiff, Bruker Biospin AG, Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Thomas, Frank, Università degli Studi di Firenze = University of Florence (UniFI), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)

Subjects
Nuclear and High Energy Physics, Magnetic Resonance Spectroscopy, Proton, Protein Conformation, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Biophysics, Intrinsically disordered proteins, Biochemistry, chemistry.chemical_compound, Protein structure, Amide, Humans, Carbon Radioisotopes, Nuclear Magnetic Resonance, Biomolecular, chemistry.chemical_classification, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, Ubiquitin, Relaxation (NMR), Nuclear magnetic resonance spectroscopy, Carbon-13 NMR, Condensed Matter Physics, Amides, Amino acid, Intrinsically Disordered Proteins, Crystallography, Solvents, alpha-Synuclein, Protons, Hydrogen
Abstract

International audience; Intrinsically disordered proteins (IDPs) are functional proteins containing large fragments characterized by high local mobility. Bioinformatic studies have suggested that a significant fraction (more than 30%) of eukaryotic proteins has disordered regions of more than 50 amino acids in length. Hence, NMR methods for the characterization of local compactness and solvent accessibility in such highly disordered proteins are of high importance. Among the available approaches, the HET-SOFAST/BEST experiments (Schanda et al., 2006, Rennella et al., 2014) provide semi-quantitative information by monitoring longitudinal (1)H relaxation of amide protons under different initial conditions. However, when approaching physiological sample conditions, the potential of these amide (1)H detected experiments is reduced due to rapid amide proton solvent exchange. (13)C direct detection methods therefore provide a valuable alternative thanks to a higher chemical shift dispersion and their intrinsic insensitivity toward solvent exchange. Here we present two sets of (13)C-detected experiments, which indirectly measure (1)H(N) and (1)H(α) inversion recovery profiles. The experiments consist of an initial spin inversion-recovery block optimized for selective manipulation of different types of proton spins followed by a CON read-out scheme. The proposed experiments were tested on human α-synuclein and ubiquitin, two representative examples of unfolded and folded proteins.

Published
2015

7. NMR Methods for the Study of Instrinsically Disordered Proteins Structure, Dynamics, and Interactions: General Overview and Practical Guidelines

Author

Brutscher, Bernhard, Felli, Isabella C, Gil-Caballero, Sergio, Hošek, Tomáš, Kümmerle, Rainer, Piai, Alessandro, Pierattelli, Roberta, Sólyom, Zsófia, Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Magnet Resonance Ctr CERM, Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), Dept Chem Ugo Schiff, Bruker Biospin AG, Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), and Università degli Studi di Firenze = University of Florence (UniFI)

Subjects
Intrinsically Disordered Proteins, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Protein Conformation, Nuclear Magnetic Resonance, Biomolecular
Abstract

International audience; Thanks to recent improvements in NMR instrumentation, pulse sequence design, and sample preparation, a panoply of new NMR tools has become available for atomic resolution characterization of intrinsically disordered proteins (IDPs) that are optimized for the particular chemical and spectroscopic properties of these molecules. A wide range of NMR observables can now be measured on increasingly complex IDPs that report on their structural and dynamic properties in isolation, as part of a larger complex, or even inside an entire living cell. Herein we present basic NMR concepts, as well as optimised tools available for the study of IDPs in solution. In particular, the following sections are discussed hereafter: a short introduction to NMR spectroscopy and instrumentation (Sect. 3.1), the effect of order and disorder on NMR observables (Sect. 3.2), particular challenges and bottlenecks for NMR studies of IDPs (Sect. 3.3), 2D HN and CON NMR experiments: the fingerprint of an IDP (Sect. 3.4), tools for overcoming major bottlenecks of IDP NMR studies (Sect. 3.5), (13)C detected experiments (Sect. 3.6), from 2D to 3D: from simple snapshots to site-resolved characterization of IDPs (Sect. 3.7), sequential NMR assignment: 3D experiments (Sect. 3.8), high-dimensional NMR experiments (nD, with n > 3) (Sect. 3.9) and conclusions and perspectives (Sect. 3.10).

Published
2015

8. pE-DB: a database of structural ensembles of intrinsically disordered and of unfolded proteins

Author

A. Keith Dunker, Dmitri I. Svergun, Joel L. Sussman, Michele Vendruscolo, Richard W. Kriwacki, Martin Blackledge, Peter E. Wright, Isabella C. Felli, Roberta Pierattelli, Peter Tompa, Pierre Lebrun, Mihaly Varadi, David S. Wishart, Erica Valentini, Simone Kosol, Julie D. Forman-Kay, Vladimir N. Uversky, Structural Biology Brussels, Department of Bio-engineering Sciences, Groupe Flexibilité et Dynamique des Protéines par RMN (IBS-FDP ), Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Magnet Resonance Ctr CERM, Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), Dept Chem Ugo Schiff, Department of Structural Biology, St Jude Children's Research Hospital, Weizmann Institute of Science [Rehovot, Israël], European Molecular Biology Laboratory [Hamburg] (EMBL), Department of Chemistry and Biochemistry, University of California [Santa Cruz] (UCSC), University of California-University of California, Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indiana University System-Indiana University System, Institute for Biological Instrumentation, Russian Academy of Sciences [Moscow] (RAS), Department of Chemistry, University of Cambridge [UK] (CAM), Department of Biological Sciences [Edmonton], University of Alberta, Department of Computing Science [Edmonton], Department of Computer Science [York] (CS-YORK), University of York [York, UK], Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Università degli Studi di Firenze = University of Florence (UniFI), University of California [Santa Cruz] (UC Santa Cruz), and University of California (UC)-University of California (UC)

Subjects
MESH: Databases, Protein, Intrinsically dosordered proteins, IDP, NMR, modeling, Biology, 010402 general chemistry, Intrinsically disordered proteins, computer.software_genre, 01 natural sciences, Set (abstract data type), II. Protein sequence and structure, motifs and domains, 03 medical and health sciences, Modelling methods, MESH: Nuclear Magnetic Resonance, Biomolecular, Scattering, Small Angle, Genetics, Data bank, Conformational sampling, Databases, Protein, Nuclear Magnetic Resonance, Biomolecular, MESH: Scattering, Small Angle, 030304 developmental biology, Protein Unfolding, MESH: Intrinsically Disordered Proteins, Flexibility (engineering), 0303 health sciences, Internet, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Database, MESH: X-Ray Diffraction, A protein, Experimental data, 0104 chemical sciences, X-ray diffraction, MESH: Internet, MESH: Protein Unfolding, ddc:540, intrinsically disordered proteins, computer
Abstract

International audience; The goal of pE-DB (http://pedb.vib.be) is to serve as an openly accessible database for the deposition of structural ensembles of intrinsically disordered proteins (IDPs) and of denatured proteins based on nuclear magnetic resonance spectroscopy, small-angle X-ray scattering and other data measured in solution. Owing to the inherent flexibility of IDPs, solution techniques are particularly appropriate for characterizing their biophysical properties, and structural ensembles in agreement with these data provide a convenient tool for describing the underlying conformational sampling. Database entries consist of (i) primary experimental data with descriptions of the acquisition methods and algorithms used for the ensemble calculations, and (ii) the structural ensembles consistent with these data, provided as a set of models in a Protein Data Bank format. PE-DB is open for submissions from the community, and is intended as a forum for disseminating the structural ensembles and the methodologies used to generate them. While the need to represent the IDP structures is clear, methods for determining and evaluating the structural ensembles are still evolving. The availability of the pE-DB database is expected to promote the development of new modeling methods and leads to a better understanding of how function arises from disordered states.

Published
2014
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9. Frontiers in Solid-State NMR Technology

Author

Lyndon Emsley, Ivano Bertini, ISA - Centre de RMN à très hauts champs, Institut des Sciences Analytiques (ISA), Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon, Dept Chem Ugo Schiff, Università degli Studi di Firenze = University of Florence [Firenze], Magnet Resonance Ctr CERM, Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), and Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI)

Subjects
Engineering, Web of science, 010405 organic chemistry, business.industry, MathematicsofComputing_GENERAL, Nanotechnology, General Medicine, General Chemistry, 010402 general chemistry, 01 natural sciences, GeneralLiterature_MISCELLANEOUS, 0104 chemical sciences, World Wide Web, InformationSystems_GENERAL, Solid-state nuclear magnetic resonance, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, business, ComputingMilieux_MISCELLANEOUS
Abstract

Reference EPFL-ARTICLE-204293doi:10.1021/ar4001718View record in Web of Science Record created on 2015-01-08, modified on 2016-08-09

Published
2013
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10. 13C-detected through-bond correlation experiments for protein resonance assignment by ultra-fast MAS solid-state NMR

Author

Lyndon Emsley, Isabella C. Felli, Anne Lesage, Roberta Pierattelli, Guido Pintacuda, Amy L. Webber, Emeline Barbet-Massin, Michael J. Knight, Andrew J. Pell, ISA - Centre de RMN à très hauts champs, Institut des Sciences Analytiques (ISA), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Dept Chem Ugo Schiff, Università degli Studi di Firenze = University of Florence (UniFI), Magnet Resonance Ctr CERM, Agence Nationale de la Recherche (ANR 08-BLAN-0035-01 et 10-BLAN-713-01), Ente Cassa di Risparmio di Firenze ( Egide 22397RJ et 26000XF), Universita Italo-Francese 26000XF, European Commission, EC (BioNMR) 261863, European Project, Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), and Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI)

Subjects
J-coupling, Time Factors, C-13, Analytical chemistry, 010402 general chemistry, 01 natural sciences, Homonuclear molecule, paramagnetism, MICROCRYSTALLINE PROTEIN, Paramagnetism, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, solid-state nuclear magnetic resonance spectroscopy, magic angle spinning, Magic angle spinning, Peptide bond, Physical and Theoretical Chemistry, Nuclear Magnetic Resonance, Biomolecular, J COUPLING-CONSTANTS, PROTONLESS NMR, Carbon Isotopes, PARAMAGNETIC METALLOPROTEINS, 010405 organic chemistry, Chemistry, Resonance, Proteins, CORRELATION SPECTROSCOPY, superoxide dismutase, Atomic and Molecular Physics, and Optics, 3. Good health, 0104 chemical sciences, [CHIM.THEO]Chemical Sciences/Theoretical and/or physical chemistry, Crystallography, ANGLE-SPINNING NMR, Solid-state nuclear magnetic resonance, RESOLUTION, CROSS-POLARIZATION, ZINC SUPEROXIDE-DISMUTASE, Two-dimensional nuclear magnetic resonance spectroscopy
Abstract

International audience; We present two sequences which combine ((1)H,(15)N) and ((15)N,(13)C) selective cross-polarization steps with an efficient variant of the J-based homonuclear transfer scheme, in which a spin-state-selective (S(3)E) block is incorporated to improve both resolution and sensitivity in the direct (13)C dimension. We propose these two sequences as a part of a suite of four N-C correlation experiments allowing for the assignment of protein backbone resonances in the solid state. We illustrate these experiments under ultra-fast magic angle spinning conditions on two samples of microcrystalline dimeric human superoxide dismutase (SOD, 153×2 amino acids), in its diamagnetic ("empty", Zn(II)) and paramagnetic (Cu(II), Zn(II)) states.

Published
2012
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11. Combination of DQ and ZQ Coherences for Sensitive Through-Bond NMR Correlation Experiments in Biosolids under Ultra-Fast MAS

Author

Ivano Bertini, Anne Lesage, Emeline Barbet-Massin, Andrew J. Pell, Lyndon Emsley, Isabella C. Felli, Roberta Pierattelli, Guido Pintacuda, Amy L. Webber, Michael J. Knight, ISA - Centre de RMN à très hauts champs, Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Dept Chem Ugo Schiff, Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), Magnet Resonance Ctr CERM, Agence Nationale de la Recherche (ANR) 08-BLAN-0035-01 10-BLAN-713-01, Ente Cassa di Risparmio di Firenze, Egide (programmes Galiee) 22397RJ 26000XF : Universita Italo-francese 11/12, and EC 261863

Subjects
PROTEINS, Dephasing, Analytical chemistry, 010402 general chemistry, 01 natural sciences, Molecular physics, Spectral line, Magnetization, NMR spectroscopy, Author Keywords: double and zero quantum coherences, magic angle spinning, through-bond interactions, Magic angle spinning, Humans, Physical and Theoretical Chemistry, Nuclear Magnetic Resonance, Biomolecular, DOUBLE-QUANTUM COHERENCE, ZERO-QUANTUM, Spin-½, J COUPLING-CONSTANTS, PROTONLESS NMR, Carbon Isotopes, Nitrogen Isotopes, 010405 organic chemistry, Chemistry, Superoxide Dismutase, Nuclear magnetic resonance spectroscopy, solid state, CORRELATION SPECTROSCOPY, Atomic and Molecular Physics, and Optics, 0104 chemical sciences, SOLID-STATE NMR, [CHIM.THEO]Chemical Sciences/Theoretical and/or physical chemistry, ANGLE-SPINNING NMR, Solid-state nuclear magnetic resonance, COMPLETE ASSIGNMENT, Quantum Theory, CROSS-POLARIZATION, Crystallization, double and zero quantum coherences, Two-dimensional nuclear magnetic resonance spectroscopy
Abstract

International audience; A double-zero quantum (DZQ)-refocused INADEQUATE experiment is introduced for J-based NMR correlations under ultra-fast (60 kHz) magic angle spinning (MAS). The experiment records two spectra in the same dataset, a double quantumsingle quantum (DQ-SQ) and zero quantumsingle quantum (ZQ-SQ) spectrum, whereby the corresponding signals appear at different chemical shifts in ?1. Furthermore, the spin-state selective excitation (S3E) J-decoupling block is incorporated in place of the second refocusing echo of the INADEQUATE scheme, providing an additional gain in sensitivity and resolution. The two sub-spectra acquired in this way can be treated separately by a shearing transformation, producing two diagonal-free single quantum (SQ-SQ)-type spectra, which are subsequently recombined to give an additional sensitivity enhancement, thus offering an improvement greater than a factor of two as compared to the original refocused INADEQUATE experiment. The combined DZQ scheme retains transverse magnetization on the initially polarized (I) spin, which typically exhibits a longer transverse dephasing time (T2') than its through-bond neighbour (S). By doing so, less magnetization is lost during the refocusing periods in the sequence to give even further gains in sensitivity for the J correlations. The experiment is demonstrated for the correlation between the carbonyl (CO) and alpha (CA) carbons in a microcrystalline sample of fully protonated, [15N,13C]-labelled CuII,?ZnII superoxide dismutase, and its efficiency is discussed with respect to other J-based schemes.

Published
2012
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12. Fast resonance assignment and fold determination of human superoxide dismutase by high-resolution proton-detected solid-state MAS NMR spectroscopy

Author

Andrew J. Pell, Anne Lesage, Ivano Bertini, Torsten Herrmann, Lyndon Emsley, Guido Pintacuda, Amy L. Webber, Paul Guerry, Leonardo Gonnelli, Emeline Barbet-Massin, Michael J. Knight, Roberta Pierattelli, Isabella C. Felli, ISA - Centre de RMN à très hauts champs, Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Dept Chem Ugo Schiff, Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), Magnet Resonance Ctr CERM, Magnetic Resonance Center, Agence Nationale de la Recherche (ANR 08-BLAN-0035-01 and 10-BLAN-713-01), Ente Cassa di Risparmio di Firenze, Egide (programme Galiee 22397RJ), the Universita Italo-francese (programma Galileo 09/10), Joint Research Activity and Access to Research Infrastructures in the 7th Framework program of the EC (BioNMR n. 261863)., European Project, Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), and Università degli Studi di Firenze = University of Florence (UniFI)

Subjects
Models, Molecular, Protein Folding, C-13, Protein Conformation, Analytical chemistry, Solid-state, 010402 general chemistry, 01 natural sciences, Catalysis, Spectral line, 3D STRUCTURE DETERMINATION, Superoxide dismutase, Protein structure, NMR spectroscopy, RESTRAINTS, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, LARGER PROTEINS, structural constraints, COHERENCE, SPECTRA, Humans, protein structure, WILD-TYPE, Nuclear Magnetic Resonance, Biomolecular, ComputingMilieux_MISCELLANEOUS, 1H detection, Mas nmr spectroscopy, biology, Chemistry, 010405 organic chemistry, MEMBRANE-PROTEINS, Superoxide Dismutase, General Chemistry, Nuclear magnetic resonance spectroscopy, General Medicine, 0104 chemical sciences, NMR spectra database, Microcrystalline, PROTEIN-STRUCTURE DETERMINATION, biology.protein, PERDEUTERATED PROTEINS, Protons
Abstract

Re-protonation is key: A combination of a high magnetic field (1 GHz) and ultra-fast magic-angle spinning (60 kHz) allows easy detection of NMR spectra revealing details of secondary and tertiary structures of medium-sized proteins. The technique was applied to the 153-residue microcrystalline Zn II-loaded superoxide dismutase (ZnII-SOD) fully [ 2H,13C,15N]-labeled and 100% re-protonated at the exchangeable sites. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Published
2011
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13. High-resolution and sensitivity through-bond correlations in ultra-fast magic angle spinning (MAS) solid-state NMR

Author

Lyndon Emsley, Ivano Bertini, Józef R. Lewandowski, Ségolène Laage, Isabella C. Felli, Anne Lesage, Guido Pintacuda, Roberta Pierattelli, Alessandro Marchetti, Dept Chem Ugo Schiff, Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), Magnet Resonance Ctr CERM, ISA - Centre de RMN à très hauts champs, Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Scuola Normale Superiore di Pisa (SNS), Agence Nationale de la Recherche (ANR 08-BLAN-0035-01), Ente Cassa di Risparmio di Firenze, from Egide (programme Galilee 22397RJ), the Universita Italo-Francese (Programma Galileo 2009/2010), Joint Research Activity and Access to Research Infrastructures activity in the 6th Framework Program of the EC (RII3-026145, EU-NMR), EU IRG (PIRG03-GA-2008-231026)., European Project, Università degli Studi di Firenze = University of Florence (UniFI), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)

Subjects
SELECTION, Spin states, PROTEINS, C-13, Analytical chemistry, 010402 general chemistry, 01 natural sciences, Homonuclear molecule, ASSIGNMENT STRATEGIES, ENHANCEMENT, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, DISORDERED SOLIDS, Magic angle spinning, Molecule, HETERONUCLEAR CORRELATION, Spectroscopy, HOMONUCLEAR, chemistry.chemical_classification, SPECTROSCOPY, 010405 organic chemistry, Biomolecule, General Chemistry, 0104 chemical sciences, Microcrystalline, Solid-state nuclear magnetic resonance, chemistry, Chemical physics, CROSS-POLARIZATION
Abstract

International audience; We introduce a new experiment, which makes use of Spin State Selective manipulations to perform sensitive and resolved through-bond correlations in organic and biological solids at high-fields and under ultra-fast MAS. The scheme is the shortest and most sensitive through-bond correlation method introduced so far in solids, yields resolved fingerprints of uniformly (13)C-labeled biomolecules, and constitutes a tool to highlight slight static structural disorder around crystallographically equivalent molecules in microcrystalline samples.

Published
2011
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14. Cytochrome c and SDS: a molten globule protein with altered axial ligation

Author

Arnaud Bondon, Paul R. Vasos, Paola Turano, Gérard Simonneaux, Ivano Bertini, Soizic Chevance, Dept Chem Ugo Schiff, Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), Magnet Resonance Ctr CERM, Magnet Resonance Center (CERM), Laboratoire de Chimie et de Biochimie Pharmacologiques et Toxicologiques (LCBPT - UMR 8601), Université Paris Descartes - Paris 5 (UPD5)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Institut des Sciences Chimiques de Rennes (ISCR), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Institut National des Sciences Appliquées - Rennes (INSA Rennes), Institut National des Sciences Appliquées (INSA)-Université de Rennes (UNIV-RENNES)-Institut National des Sciences Appliquées (INSA)-Ecole Nationale Supérieure de Chimie de Rennes (ENSCR)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Università degli Studi di Firenze = University of Florence (UniFI), Université de Rennes (UR)-Institut National des Sciences Appliquées - Rennes (INSA Rennes), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Ecole Nationale Supérieure de Chimie de Rennes (ENSCR)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), University of Florence (UNIFI), Université Paris Descartes - Paris 5 (UPD5)-Centre National de la Recherche Scientifique (CNRS), Institut National des Sciences Appliquées - Rennes (INSA Rennes), Institut National des Sciences Appliquées (INSA)-Université de Rennes (UNIV-RENNES)-Institut National des Sciences Appliquées (INSA)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS)-Ecole Nationale Supérieure de Chimie de Rennes-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES), University of Florence, Magnet Resonance Center ( CERM ), Laboratoire de Chimie et de Biochimie Pharmacologiques et Toxicologiques ( LCBPT - UMR 8601 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Centre National de la Recherche Scientifique ( CNRS ), Institut des Sciences Chimiques de Rennes ( ISCR ), Université de Rennes 1 ( UR1 ), and Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Ecole Nationale Supérieure de Chimie de Rennes-Institut National des Sciences Appliquées ( INSA ) -Centre National de la Recherche Scientifique ( CNRS )

Subjects
Magnetic Resonance Spectroscopy, Time Factors, Cytochrome, MESH : Cytochromes c, Protein Conformation, MESH : Saccharomyces cerevisiae, MESH : Rotation, 01 natural sciences, Protein structure, MESH: Protein Conformation, MESH: Rotation, MESH: Saccharomyces cerevisiae Proteins, Structural Biology, MESH : Myocardium, MESH: Animals, MESH : Protein Conformation, MESH : Viscosity, 0303 health sciences, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, Viscosity, Cytochrome c, Cytochromes c, Sodium Dodecyl Sulfate, Nuclear magnetic resonance spectroscopy, MESH: Cytochromes c, Ligand (biochemistry), MESH: Saccharomyces cerevisiae, Molten globule, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Protons, MESH : Time Factors, MESH: Sodium Dodecyl Sulfate, Saccharomyces cerevisiae Proteins, MESH: Myocardium, Rotation, MESH: Viscosity, Saccharomyces cerevisiae, 010402 general chemistry, MESH : Saccharomyces cerevisiae Proteins, 03 medical and health sciences, MESH : Protons, Animals, Horses, MESH: Horses, Molecular Biology, Rotational correlation time, MESH : Sodium Dodecyl Sulfate, 030304 developmental biology, MESH: Magnetic Resonance Spectroscopy, Myocardium, MESH: Time Factors, Random coil, 0104 chemical sciences, Crystallography, biology.protein, MESH : Horses, MESH : Magnetic Resonance Spectroscopy, MESH : Animals, MESH: Protons, [ SDV.BBM.BS ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM]
Abstract

International audience; Saccharomices cerevisiae (yeast iso-1) cytochrome c has been investigated in the presence of 100 mM SDS in order to simulate the interaction of cytochrome c with membrane. Under these circumstances, a high spin species with detached methionine axial ligand is observed through NMR, in analogy to findings on the horse heart protein. However, at variance with the latter system, for the yeast protein also a low spin species is detected, which appears to be present with a concentration of about 40% with respect to that of the high spin species. The R(1), R(2), [1H]-15N NOE of backbone amides which are not affected by paramagnetism are homogeneous and allow a simultaneous analysis of the data for the two species. The result is that the rotational correlation time is larger than in water and larger than expected on the basis of viscosity of the SDS-containing solution. This finding suggests interactions of cytochrome c with SDS. Furthermore, it appears that there is subnanosecond backbone mobility, which also accounts for the decreased intensity of NOE cross-peaks and may be associated with equilibria between helical and random coil structure. The dynamic behavior appears to be a common feature of the high spin and low spin species and is consistent with the presence of a molten globule state. The molten globule nature of the protein could account for the presence of the different axial coordination of the heme iron. Such findings are meaningful with respect to the physiology of cytochrome c as electron transfer protein and as promoter of apoptosis.

Published
2004

15. Structure and backbone dynamics of a microcrystalline metalloprotein by solid-state NMR

Author

Lyndon Emsley, Andrew J. Pell, Guido Pintacuda, Isabella C. Felli, Leonardo Gonnelli, Michael J. Knight, Torsten Herrmann, Ivano Bertini, Roberta Pierattelli, ISA - Centre de RMN à très hauts champs, Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Dept Chem Ugo Schiff, Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), Magnet Resonance Ctr CERM, Magnetic Resonance Center, Agence Nationale de la Recherche (ANR 08-BLAN-0035-01 and 10-BLAN-713-01), Ente Cassa di Risparmio di Firenze, Egide (programme Galiee 22397RJ), the Universita Italo-francese (programma Galileo 11/12), Joint Research Activity and Access to Research Infrastructures in the seventh Framework Programme of the European Community (EAST-NMR 228461, BioNMR 261863)., European Project, Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), and Università degli Studi di Firenze = University of Florence (UniFI)

Subjects
Models, Molecular, Magnetic Resonance Spectroscopy, Nitrogen, Molecular Conformation, Normal Distribution, 010402 general chemistry, 01 natural sciences, nuclear relaxation rates, microcrystal, Catalysis, paramagnetism, Paramagnetism, LATTICE-RELAXATION, RESTRAINTS, Catalytic Domain, Metalloproteins, Magic angle spinning, Molecule, WILD-TYPE, Nuclear Magnetic Resonance, Biomolecular, NUCLEAR-SPIN RELAXATION, Multidisciplinary, PARAMAGNETIC METALLOPROTEINS, SPECTROSCOPY, 010405 organic chemistry, Chemistry, Relaxation (NMR), CRYSTALLINE PROTEIN, Nuclear magnetic resonance spectroscopy, QUANTITATIVE-ANALYSIS, Carbon, 0104 chemical sciences, [CHIM.THEO]Chemical Sciences/Theoretical and/or physical chemistry, Crystallography, Solid-state nuclear magnetic resonance, RESOLUTION, Chemical physics, ZN SUPEROXIDE-DISMUTASE, Yield (chemistry), copper, Physical Sciences, Diamagnetism, Protons, Crystallization, CU, Hydrogen
Abstract

We introduce a new approach to improve structural and dynamical determination of large metalloproteins using solid-state nuclear magnetic resonance (NMR) with 1 H detection under ultrafast magic angle spinning (MAS). The approach is based on the rapid and sensitive acquisition of an extensive set of 15 N and 13 C nuclear relaxation rates. The system on which we demonstrate these methods is the enzyme Cu, Zn superoxide dismutase (SOD), which coordinates a Cu ion available either in Cu + (diamagnetic) or Cu 2+ (paramagnetic) form. Paramagnetic relaxation enhancements are obtained from the difference in rates measured in the two forms and are employed as structural constraints for the determination of the protein structure. When added to 1 H- 1 H distance restraints, they are shown to yield a twofold improvement of the precision of the structure. Site-specific order parameters and timescales of motion are obtained by a Gaussian axial fluctuation (GAF) analysis of the relaxation rates of the diamagnetic molecule, and interpreted in relation to backbone structure and metal binding. Timescales for motion are found to be in the range of the overall correlation time in solution, where internal motions characterized here would not be observable.

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16. Protein residue linking in a single spectrum for magic-angle spinning NMR assignment

Author

Tanguy Le Marchand, Guido Pintacuda, Loren B. Andreas, Sebastian Wegner, Diane Cala de Paepe, Andrea Bertarello, Carl Öster, Isabella C. Felli, Torsten Herrmann, Nicholas E. Dixon, Camille Doyen, Lyndon Emsley, Roberta Pierattelli, Daniela Lalli, Magdaléna Krejčíková, Benno Knott, Frank Engelke, Jan Stanek, Biological Solid-State NMR Methods - Méthodes de RMN à l'état solide en biologie, Institut des Sciences Analytiques (ISA), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Magnet Resonance Center (CERM), Università degli Studi di Firenze = University of Florence [Firenze] (UNIFI), Cent European Inst Technol CEITEC, Masaryk University [Brno] (MUNI), Dept Chem, University of Warwick [Coventry], Bruker BioSpin GmbH, D-76287 Rheinstetten, Germany, affiliation inconnue, Magnet Resonance Ctr CERM, Dept Chem Ugo Schiff, School of Chemistry [Wollongong], University of Wollongong [Australia], Ecole Polytech Fed Lausanne, Inst Sci & Ingn Chim, BCH, CH-1015 Lausanne, Switzerland, Solid-State NMR Methods for Materials - Méthodes de RMN à l'état solide pour les matériaux, Computational Methods for Biomolecular Studies, We acknowledge support from CNRS (Fondation pour la Chimie des Substances Naturelles) and from the People Programme of the European Union's FP7 (FP7-PEOPLE-2012-ITN REA Grant agreement No 317127 'pNMR' and 316630 'CAS-IDP'). LBA is supported by a MC incoming fellowship (REA Grant agreement No 624918 'MEM-MAS'), and JS by an EMBO fellowship (ALTF 1506-2014) and by the Marie Curie Actions of the European Commission (LTFCO-FUND2013, GA-2013-609409)., European Project: 317127,EC:FP7:PEOPLE,FP7-PEOPLE-2012-ITN,PNMR(2013), European Project: 316630,EC:FP7:PEOPLE,FP7-PEOPLE-2012-ITN,CAS-IDP(2013), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), and Università degli Studi di Firenze = University of Florence (UniFI)

Subjects
Automation, Magic-angle spinning, Protein resonance assignment, Proton detection, Nuclear Magnetic Resonance, Biomolecular, Proteins, Protons, Spectroscopy, Biochemistry, 010402 general chemistry, 01 natural sciences, Spectral line, chemistry.chemical_compound, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, Amide, Magic angle spinning, Spinning, 010405 organic chemistry, Chemistry, Resonance, Pulse sequence, 0104 chemical sciences, Crystallography, Microcrystalline
Abstract

Here we introduce a new pulse sequence for resonance assignment that halves the number of data sets required for sequential linking by directly correlating sequential amide resonances in a single diagonal-free spectrum. The method is demonstrated with both microcrystalline and sedimented deuterated proteins spinning at 60 and 111 kHz, and a fully protonated microcrystalline protein spinning at 111 kHz, with as little as 0.5 mg protein sample. We find that amide signals have a low chance of ambiguous linkage, which is further improved by linking in both forward and backward directions. The spectra obtained are amenable to automated resonance assignment using general-purpose software such as UNIO-MATCH.

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