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An optical pumping technique was used to enhance and localize nuclear magnetic resonance (NMR) signals from an n-doped GaAs/Al0.1Ga0.9As multiple quantum well structure, permitting direct radio-frequency measurements of gallium-71 NMR spectra and nuclear spin-lattice relaxation rates (1/T1) as functions of temperature (1.6 K < T < 4.2 K) and the Landau level filling factor (0.66 < v < 1.76). The measurements reveal effects of electron-electron interactions on the energy levels and spin states of the two-dimensional electron system confined in the GaAs wells. Minima in 1/T1 at v approximately 1 and v approximately 2/3 indicate energy gaps for electronic excitations in both integer and fractional quantum Hall states. Rapid, temperature-independent relaxation at intermediate v values indicates a manifold of low-lying electronic states with mixed spin polarizations.
Assuntos
Arsenicais/química , Elétrons , Gálio/química , Fenômenos Químicos , Físico-Química , Espectroscopia de Ressonância Magnética , TemperaturaRESUMO
The results of (13)C nuclear magnetic resonance (NMR) measurements on alkali fullerides KxC(60) are reported. The NMR spectra demonstrate that material with 0 < x < 3 is in fact a two-phase system at equilibrium, with x = 0 and x = 3. NMR lineshapes indicate that C(3-)(60) ions rotate rapidly in the K(3)C(60) phase at 300 K, while C(6)-(60) ions in the insulating K(6)C(60) phase are static on the time scale of the lineshape measurement. The temperature dependence of the (13)C spin-lattice relaxation rate in the normal state of K(3)C(60) is found to be characteristic of a metal, indicating the important role of the C(3-)(60) ions in the conductivity. From the relaxation measurements, an estimate of the density of electronic states at the Fermi level is derived.
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The fullerene C(60) can be converted into two different structures by high pressure and temperature. They are metastable and revert to pristine C(60) on reheating to 300 degrees C at ambient pressure. For synthesis temperatures between 300 degrees and 400 degrees C and pressures of 5 gigapascals, a nominal face-centered-cubic structure is produced with a lattice parameter a(o) = 13.6 angstroms. When treated at 500 degrees to 800 degrees C at the same pressure, C(60) transforms into a rhombohedral structure with hexagonal lattice parameters of a(o) = 9.22 angstroms and c(o) = 24.6 angstroms. The intermolecular distance is small enough that a chemical bond can form, in accord with the reduced solubility of the pressure-induced phases. Infrared, Raman, and nuclear magnetic resonance studies show a drastic reduction of icosahedral symmetry, as might occur if the C(60) molecules are linked.
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Amyloid fibrils are intrinsically noncrystalline, insoluble, high-molecular-weight aggregates of peptides and proteins, with considerable biomedical and biophysical significance. Solid-state NMR techniques are uniquely capable of providing high-resolution, site-specific structural constraints for amyloid fibrils, at the level of specific interatomic distances and torsion angles. So far, a relatively small number of solid-state NMR studies of amyloid fibrils have been reported. These have addressed issues about the supramolecular organization of beta-sheets in the fibrils and the peptide conformation in the fibrils, and have concentrated on the beta-amyloid peptide of Alzheimer's disease. Many additional applications of solid-state NMR to amyloid fibrils from a variety of sources are anticipated in the near future, as these systems are ideally suited for the technique and are of widespread current interest.
Assuntos
Amiloide/química , Ressonância Magnética Nuclear Biomolecular/métodos , Doença de Alzheimer/metabolismo , Amiloide/metabolismo , Humanos , Sondas Moleculares , Conformação ProteicaRESUMO
We report solid state nuclear magnetic resonance (NMR) measurements that probe the supramolecular organization of beta-sheets in the cross-beta motif of amyloid fibrils formed by residues 11-25 of the beta-amyloid peptide associated with Alzheimer's disease (Abeta(11-25)). Fibrils were prepared at pH 7.4 and pH 2.4. The solid state NMR data indicate that the central hydrophobic segment of Abeta(11-25) (sequence LVFFA) adopts a beta-strand conformation and participates in antiparallel beta-sheets at both pH values, but that the registry of intermolecular hydrogen bonds is pH-dependent. Moreover, both registries determined for Abeta(11-25) fibrils are different from the hydrogen bond registry in the antiparallel beta-sheets of Abeta(16-22) fibrils at pH 7.4 determined in earlier solid state NMR studies. In all three cases, the hydrogen bond registry is highly ordered, with no detectable "registry-shift" defects. These results suggest that the supramolecular organization of beta-sheets in amyloid fibrils is determined by a sensitive balance of multiple side-chain-side-chain interactions. Recent structural models for Abeta(11-25) fibrils based on X-ray fiber diffraction data are inconsistent with the solid state NMR data at both pH values.
Assuntos
Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/ultraestrutura , Ressonância Magnética Nuclear Biomolecular/métodos , Sequência de Aminoácidos , Humanos , Ligação de Hidrogênio , Concentração de Íons de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Microscopia Eletrônica , Modelos Moleculares , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/ultraestrutura , Estrutura Secundária de ProteínaRESUMO
Rev is a 116 residue basic protein encoded by the genome of human immunodeficiency virus type 1 (HIV-1) that binds to multiple sites in the Rev response element (RRE) of viral mRNA transcripts in nuclei of host cells, leading to transport of incompletely spliced and unspliced viral mRNA to the cytoplasm of host cells in the latter phases of the HIV-1 life cycle. Rev is absolutely required for viral replication. Because Rev aggregates and fibrillizes in solution at concentrations required for crystal growth or liquid state NMR measurements, high-resolution structural characterization of full-length Rev has not been possible. Previously, circular dichroism studies have shown that approximately 50 % of the Rev sequence adopts helical secondary structure, predicted to correspond to a helix-loop-helix structural motif in the N-terminal half of the protein. We describe the application of solid-state NMR techniques to Rev fibrils as a means of obtaining site-specific, atomic-level structural constraints without requiring a high degree of solubility or crystallinity. Solid-state NMR measurements, using the double-quantum chemical shift anisotropy and constant-time double-quantum-filtered dipolar recoupling techniques, provide constraints on the phi and psi backbone dihedral angles at sites in which consecutive backbone carbonyl groups are labeled with (13)C. Quantitative analysis of the solid-state NMR data, by comparison with numerical simulations, indicates helical phi and psi angles at residues Leu13 and Val16 in the predicted helix 1 segment, and at residues Arg39, Arg 42, Arg43, and Arg44 in the predicted helix 2 segment. These data represent the first site-specific structural constraints from NMR spectroscopy on full-length Rev, and support the helix-loop-helix structural model for its N-terminal half.
Assuntos
Produtos do Gene rev/química , Produtos do Gene rev/metabolismo , HIV-1/química , Sequências Hélice-Alça-Hélice , Ressonância Magnética Nuclear Biomolecular , Sequência de Aminoácidos , Amiloide/química , Anisotropia , Dicroísmo Circular , Produtos do Gene rev/ultraestrutura , Microscopia Eletrônica , Dados de Sequência Molecular , Ligação Proteica , Estrutura Quaternária de Proteína , Estrutura Secundária de Proteína , Solubilidade , Produtos do Gene rev do Vírus da Imunodeficiência HumanaRESUMO
New derivations of selection rules for excitation and detection of multiple quantum coherences in coupled spin-1/2 systems are presented. The selection rules apply to experiments in which the effective coupling Hamiltonian used for multiple quantum excitation is both time-reversal invariant and time-reversible by a phase shift of the radiofrequency pulse sequence that generates the effective couplings. The selection rules are shown to be consequences of time-reversal invariance and time-reversibility and otherwise independent of the specific form of the effective coupling Hamiltonian. Numerical simulations of multiple quantum NMR signal amplitudes and experimental multiple quantum excitation spectra are presented for the case of a multiply (13)C-labeled helical polypeptide. The simulations and experiments confirm the selection rules and demonstrate their impact on multiple quantum (13)C NMR spectra in this biochemically relevant case.
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Espectroscopia de Ressonância Magnética/métodos , Isótopos de Carbono , Humanos , Peptídeos/química , Análise de Sequência de ProteínaRESUMO
Enhancement of sensitivity in solid state (15)N NMR by indirect detection through (1)H NMR signals under high-speed magic angle spinning and high-field conditions is demonstrated experimentally on two (15)N-labeled peptides, polycrystalline AlaGlyGly and the helix-forming, 17-residue peptide MB(i + 4)EK in lyophilized form. Sensitivity enhancement factors ranging from 2.0 to 3.2 are observed experimentally, depending on the (15)N and (1)H linewidths and polarization transfer efficiencies. The (1)H-detected two-dimensional (1)H/(15)N correlation spectrum of AlaGlyGly illustrates the possibility of increased spectral resolution and resonance assignments in indirectly detected experiments, in addition to the sensitivity enhancement.
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Espectroscopia de Ressonância Magnética/métodos , Humanos , Isótopos de Nitrogênio , Peptídeos/químicaRESUMO
We discuss procedures for processing data in rotor-synchronized two-dimensional magic angle spinning (2D MAS) NMR exchange measurements for both structural and dynamical studies. We show, both mathematically and experimentally, that there are two distinct data processing procedures that lead to 2D MAS exchange spectra with purely absorptive crosspeaks. One procedure is that described previously by Hagemeyer, Schmidt-Rohr, and Spiess (HSS). The other procedure is related, but different, and leads to crosspeak intensities given by the formulae of Herzfeld, Roberts, and Griffin (HRG). In 2D MAS exchange experiments on doubly (13)C-labeled l-alanylglycylglycine, we demonstrate that the HSS and HRG crosspeak intensities can be extracted separately from the same data set and contain independent information. Processing and analysis of 2D MAS exchange data with both the HSS and the HRG procedures may enhance utilization of the information content of 2D MAS exchange measurements.