title: In-Cell Protein Structures from 2D NMR Experiments.
abstract: In-cell NMR spectroscopy provides atomic resolution insights into the structural properties of proteins in cells, but it is rarely used to solve entire protein structures de novo. Here, we introduce a paramagnetic lanthanide-tag to simultaneously measure protein pseudocontact shifts (PCSs) and residual dipolar couplings (RDCs) to be used as input for structure calculation routines within the Rosetta program. We employ this approach to determine the structure of the protein G B1 domain (GB1) in intact Xenopus laevis oocytes from a single set of 2D in-cell NMR experiments. Specifically, we derive well-defined GB1 ensembles from low concentration in-cell NMR samples (∼50 μM) measured at moderate magnetic field strengths (600 MHz), thus offering an easily accessible alternative for determining intracellular protein structures.
authors.affiliation: Department of Chemistry, University of Basel , St. Johanns-Ring 19, 4056 Basel, Switzerland.
authors.forename: Thomas
authors.initials: T
authors.lastname: Müntener
authors.affiliation: Department of Chemistry, University of Basel , St. Johanns-Ring 19, 4056 Basel, Switzerland.
authors.forename: Daniel
authors.initials: D
authors.lastname: Häussinger
authors.affiliation: Department of Structural Biology, Leibniz Institute of Molecular Pharmacology (FMP Berlin) , Robert Roessle Straße 10, 13125 Berlin, Germany.
authors.forename: Philipp
authors.initials: P
authors.lastname: Selenko
authors.affiliation: Department of Structural Biology, Leibniz Institute of Molecular Pharmacology (FMP Berlin) , Robert Roessle Straße 10, 13125 Berlin, Germany.
authors.forename: Francois-Xavier
authors.initials: FX
authors.lastname: Theillet
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D028581:Lanthanoid Series Elements / Q000737:chemistry; D019906:Nuclear Magnetic Resonance, Biomolecular; D009865:Oocytes / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D014982:Xenopus laevis / Q000254:growth & development
chemical_list: D001426:Bacterial Proteins; C034284:IgG Fc-binding protein, Streptococcus; D028581:Lanthanoid Series Elements