title: NMR experiments for resonance assignments of 13C, 15N doubly-labeled flexible polypeptides: application to the human prion protein hPrP(23-230).
abstract: A combination of three heteronuclear three-dimensional NMR experiments tailored for sequential resonance assignments in uniformly 15N, 13C-labeled flexible polypeptide chains is described. The 3D (H)N(CO-TOCSY)NH, 3D (H)CA(CO-TOCSY)NH and 3D (H)CBCA(CO-TOCSY)NH schemes make use of the favorable 15N chemical shift dispersion in unfolded polypeptides, exploit the slow transverse 15N relaxation rates of unfolded polypeptides in high resolution constant-time [1H, 15N]-correlation experiments, and use carbonyl carbon homonuclear isotropic mixing to transfer magnetization sequentially along the amino acid sequence. Practical applications are demonstrated with the 100-residue flexible tail of the recombinant human prion protein, making use of spectral resolution up to 0.6 Hz in the 15N dimension, simultaneous correlation with the two adjacent amino acid residues to overcome problems associated with spectral overlap, and the potential of the presently described experiments to establish nearest-neighbor correlations across proline residues in the amino acid sequence.
authors.affiliation: Institut für Molekularbiologie und Biophysik, Eidgenössische Technische Hochschule Hönggerberg, Zürich, Switzerland.
authors.forename: A
authors.initials: A
authors.lastname: Liu
authors.forename: R
authors.initials: R
authors.lastname: Riek
authors.forename: G
authors.initials: G
authors.lastname: Wider
authors.forename: C
authors.initials: C
authors.lastname: von Schroetter
authors.forename: R
authors.initials: R
authors.lastname: Zahn
authors.forename: K
authors.initials: K
authors.lastname: Wüthrich
mesh_terms: D002247:Carbon Isotopes; D006801:Humans; D009038:Motion; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D010455:Peptides / Q000737:chemistry*; D011328:Prions / Q000737:chemistry; D011392:Proline / Q000737:chemistry; D017433:Protein Structure, Secondary
chemical_list: D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D010455:Peptides; D011328:Prions; D011392:Proline