1. Side chain resonances in static oriented proton-decoupled 15N solid-state NMR spectra of membrane proteins
- Author
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Lydia Prongidi-Fix, Burkhard Bechinger, Alexandre Chenal, Christopher Aisenbrey, Daniel Gillet, Institut de Chimie de Strasbourg, Centre National de la Recherche Scientifique (CNRS)-Université Louis Pasteur - Strasbourg I-Institut de Chimie du CNRS (INC), Max-Planck-Institut für Biochemie (MPIB), Max-Planck-Gesellschaft, Biochimie des Interactions Macromoléculaires / Biochemistry of Macromolecular Interactions, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Service d'Ingénierie Moléculaire pour la Santé (ex SIMOPRO) (SIMoS), Médicaments et Technologies pour la Santé (MTS), Université Paris-Saclay-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)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Paris-Saclay-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)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), The financial support by the ANR, VLM, the EU (MCRTN-33439), and ARC is gratefully acknowledged., Université Louis Pasteur - Strasbourg I-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Max-Planck-Institut für Biochemie = Max Planck Institute of Biochemistry (MPIB), and Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)
- Subjects
Magnetic Resonance Spectroscopy ,MESH: Isotope Labeling ,Protein Conformation ,bcl-X Protein ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Diphtheria Toxin/chemistry ,Catalysis ,MESH: bcl-X Protein ,03 medical and health sciences ,Colloid and Surface Chemistry ,Nuclear magnetic resonance ,MESH: Protein Conformation ,Purple Membrane ,Side chain ,Diphtheria Toxin ,[SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology ,030304 developmental biology ,0303 health sciences ,Nitrogen Isotopes ,Chemistry ,MESH: Magnetic Resonance Spectroscopy ,Tryptophan ,Membrane Proteins ,General Chemistry ,Nuclear magnetic resonance spectroscopy ,MESH: Purple Membrane ,MESH: Diphtheria Toxin ,0104 chemical sciences ,NMR spectra database ,Crystallography ,Solid-state nuclear magnetic resonance ,Membrane protein ,Membrane topology ,Isotope Labeling ,Spin echo ,MESH: Membrane Proteins ,MESH: Nitrogen Isotopes - Abstract
International audience; Proton-decoupled (15)N solid-state NMR spectra are used to analyze the structure, dynamics, and membrane topology of proteins uniformly labeled with (15)N. Preparation of the proteins by bacterial overexpression results in the labeling not only of the backbone amides but also of nitrogens localized within the side chains of arginine, glutamine, tryptophan, asparagines, lysines, and histidines. Most of these side chain resonances appear in the spectral region of the anisotropic backbone amides, and residual intensities have been observed also in cross-polarization spectra. In the past this issue has received little attention although it can cause ambiguities during assignment. Here we show that by combining cross-polarization and Hahn echo solid-state NMR experiments, it is possible to differentiate between side chain and backbone resonances. This is demonstrated using experimental and simulated (15)N spectra of oriented purple membranes, diphtheria toxin T domain and Bcl-x(L).
- Published
- 2009
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