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Heteronuclear NMR Experiments (heteronuclear + nmr_experiment)
Selected AbstractsNMR Characterization of Complex p- Oligophenyl Scaffolds by Means of Aliasing Techniques to Obtain Resolution-Enhanced Two-Dimensional SpectraHELVETICA CHIMICA ACTA, Issue 9 2004Damien Jeannerat The usefulness of computer-assisted aliasing to secure maximal resolution of signal clusters in 1H- and 13C-NMR spectra (which is essential for structure determination by HMBC 2D NMR spectroscopy) in minimal acquisition time is exemplified by the complete characterization of the two complementary p -octiphenyls 1 and 2 with complex substitution patterns. The need for digital resolution near 1,Hz/pt to dissect the extensive signal clusters in the NMR spectra of these refined oligomers excluded structure determination under routine conditions. High resolution was secured by exploiting the low signal density in the 13C dimension of HMBC spectra by using computer-assisted aliasing to maximize signal density. Based on the observed shifts in DEPT and 1H-decoupled 13C-NMR spectra of 1 and 2, computer-assisted aliasing allowed to reduce the number of required time increments by a factor of 20 to 30 compared to full-width spectra with identical resolution. Without signal-to-noise constraints, this computer-assisted aliasing reduced the acquisition time for high-resolution NMR spectra needed for complete characterization of refined oligomers 1 and 2 by the same factor (e.g., from over a day to about an hour). With resolved signal clusters in fully aliased HSQC and HMBC spectra, unproblematic structure determination of 1 and 2 is demonstrated by unambiguous assignment of all C- and H-atoms. These findings demonstrate that computer-assisted aliasing of the underexploited 13C dimension makes extensive molecular complexity accessible by conventional multidimensional heteronuclear NMR experiments without extraordinary efforts. [source] Orexin-A is composed of a highly conserved C -terminal and a specific, hydrophilic N -terminal region, revealing the structural basis of specific recognition by the orexin-1 receptorJOURNAL OF PEPTIDE SCIENCE, Issue 7 2006Tomoyo Takai Abstract Orexins-A and B, also called hypocretins-1 and 2, respectively, are neuropeptides that regulate feeding and sleep-wakefulness by binding to two orphan G protein-coupled receptors named orexin-1 (OX1R) and orexin-2 (OX2R). The sequences and functions of orexins-A and B are similar to each other, but the high sequence homology (68%) is limited in their C -terminal half regions (residues 15,33). The sequence of the N -terminal half region of orexin-A (residues 1,14), containing two disulfide bonds, is very different from that of orexin-B. The structure of orexin-A was determined using two-dimensional homonuclear and 15N and 13C natural abundance heteronuclear NMR experiments. Orexin-A had a compact conformation in the N -terminal half region, which contained a short helix (III:Cys6-Gln9) and was fixed by the two disulfide bonds, and a helix-turn-helix conformation (I:Leu16-Ala23 and II:Asn25-Thr32) in the remaining C -terminal half region. The C -terminal half region had both hydrophobic and hydrophilic residues, which existed on separate surfaces to provide an amphipathic character in helices I and II. The nine residues on the hydrophobic surface are also well conserved in orexin-B, and it was reported that the substitution of each of them with alanine resulted in a significant drop in the functional potency at the receptors. Therefore, we suggest that they form the surface responsible for the main hydrophobic interaction with the receptors. On the other hand, the residues on the hydrophilic surface, together with the hydrophilic residues in the N -terminal half region that form a cluster, are known to make only small contributions to the binding to the receptors through similar alanine-scan experiments. However, since our structure of orexin-A showed that large conformational and electrostatical differences between orexins-A and B were rather concentrated in the N -terminal half regions, we suggest that the region of orexin-A is important for the preference for orexin-A of OX1R. Copyright © 2006 European Peptide Society and John Wiley & Sons, Ltd. [source] 15N relaxation study of the amyloid ,-peptide: structural propensities and persistence lengthMAGNETIC RESONANCE IN CHEMISTRY, Issue S1 2006Jens Danielsson Abstract The dynamics of monomeric Alzheimer A,(1,40) in aqueous solution was studied using heteronuclear NMR experiments. 15N NMR relaxation rates of amide groups report on the dynamics in the peptide chain and make it possible to estimate structural propensities from temperature-dependent relaxation data and chemical shifts change analysis. The persistence length of the polypeptide chain was determined using a model in which the influence of neighboring residue relaxation is assumed to decay exponentially as a function of distance. The persistence length of the A,(1,40) monomer was found to decrease from eight to three residues when temperature was increased from 3 to 18 °C. At 3 °C the peptide shows structural propensities that correlate well with the suggested secondary structure regions of the peptide to be present in the fibrils, and with the ,-helical structure in membrane-mimicking systems. Our data leads to a structural model for the monomeric soluble ,-peptide with six different regions of secondary structure propensities. The peptide has two regions with ,-strand propensity (residues 16,24 and 31,40), two regions with high PII-helix propensity (residues 1,4 and 11,15) and two unstructured regions with higher mobility (residues 5,10 and 25,30) connecting the structural elements. Copyright © 2006 John Wiley & Sons, Ltd. [source] The Effects of Ligand Exchange and Mobility on the Peroxidase Activity of a Bacterial Cytochrome c upon UnfoldingCHEMBIOCHEM, Issue 4 2005Jonathan A. R. Worrall Dr. Abstract The effect on the heme environment upon unfolding Paracoccus versutus ferricytochrome c-550 and two site-directed variants, K99E and H118Q, has been assessed through a combination of peroxidase activity increase and one-dimensional NMR spectroscopy. At pH 4.5, the data are consistent with a low- to high-spin heme transition, with the K99E mutation resulting in a protein with increased peroxidase activity in the absence of or at low concentrations of denaturant. Furthermore, the mobility of the polypeptide chain at pH 4.5 for the wild-type protein has been monitored in the absence and presence of denaturant through heteronuclear NMR experiments. The results are discussed in terms of local stability differences between bacterial and mitochondrial cytochromes c that are inferred from peroxidase activity assays. At pH 7.0, a mixture of misligated heme states arising from protein-based ligands assigned to lysine and histidine is detected. At low denaturant concentrations, these partially unfolded misligated heme forms inhibit the peroxidase activity. Data from the K99E mutation at pH 7.0 indicate that K99 is not involved in heme misligation, whereas histidine coordination is proven by the data from the H118Q variant. [source] | ||||||||||