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Jacob Schaefer - One of the best experts on this subject based on the ideXlab platform.
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Cross-Link Formation and Peptidoglycan Lattice Assembly in the FemA Mutant of Staphylococcus aureus
Biochemistry, 2014Co-Authors: Sung Joon Kim, Manmilan Singh, Shasad Sharif, Jacob SchaeferAbstract:Staphylococcus aureus FemA mutant grown in the presence of an alanine-racemase inhibitor was labeled with d-[1-13C]alanine, l-[3-13C]alanine, [2-13C]glycine, and l-[5-19F]lysine to characterize some details of the peptidoglycan tertiary structure. Rotational-Echo Double-Resonance (REDOR) NMR of isolated cell walls was used to measure internuclear distances between 13C-labeled alanines and 19F-labeled lysine incorporated in the peptidoglycan. The alanyl 13C labels were preselected for REDOR measurement by their proximity to the glycine label using 13C–13C spin diffusion. The observed 13C–13C and 13C–19F distances are consistent with a tightly packed, hybrid architecture containing both parallel and perpendicular stems in a repeating structural motif within the peptidoglycan.
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Staphylococcus aureus peptidoglycan stem packing by Rotational-Echo double resonance NMR spectroscopy.
Biochemistry, 2013Co-Authors: Sung Joon Kim, Manmilan Singh, Maria N. Preobrazhenskaya, Jacob SchaeferAbstract:Staphylococcus aureus grown in the presence of an alanine-racemase inhibitor was labeled with d-[1-(13)C]alanine and l-[(15)N]alanine to characterize some details of the peptidoglycan tertiary structure. Rotational-Echo Double-Resonance NMR of intact whole cells was used to measure internuclear distances between (13)C and (15)N of labeled amino acids incorporated in the peptidoglycan, and from those labels to (19)F of a glycopeptide drug specifically bound to the peptidoglycan. The observed (13)C-(15)N average distance of 4.1-4.4 Å between d- and l-alanines in nearest-neighbor peptide stems is consistent with a local, tightly packed, parallel-stem architecture for a repeating structural motif within the peptidoglycan of S. aureus.
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"Development of REDOR Rotational-Echo Double-Resonance NMR" by Terry Gullion and Jacob Schaefer [J. Magn. Reson. 81 (1989) 196-200].
Journal of magnetic resonance (San Diego Calif. : 1997), 2011Co-Authors: Jacob SchaeferAbstract:The popularity of Rotational-Echo Double-Resonance (REDOR) NMR arises from its ability to measure weak dipolar couplings and long-range heteronuclear distances accurately. This ability was not anticipated in the first REDOR experiments and resulted from the effectiveness of a simple radiofrequency phase alternation scheme to suppress amplitude and phase distortions in echo trains even after hundreds of pi pulses.
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Rotational-Echo Double-Resonance NMR distance measurements for the tubulin-bound Paclitaxel conformation.
Journal of the American Chemical Society, 2007Co-Authors: Younkee Paik, Chao Yang, Belhu B. Metaferia, Shoubin Tang, Susan Bane, Rudravajhala Ravindra, Natasha Shanker, Ana A. Alcaraz, Scott A. Johnson, Jacob SchaeferAbstract:The important anticancer drug Taxol (paclitaxel, PTX) owes its unique activity to its ability to bind to tubulin in a stoichiometric ratio and promote its assembly into microtubules. The conformation of the microtubule-bound drug has been the focus of numerous research efforts, since the inability of polymerized tubulin to form crystals precludes structure proof by X-ray crystallography. Likewise, although the α,β-tubulin dimer structure has been solved by electron crystallography, the 3.7 A resolution is too low to permit direct determination of either ligand conformation or binding pose. In this article, we present experimental results from 2H{19F} REDOR NMR that provide direct confirmation that paclitaxel adopts a T-shaped conformation when it is bound to tubulin.
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Dipolar double-quantum filtered Rotational-Echo double resonance.
Magnetic resonance in chemistry : MRC, 2007Co-Authors: Shigeru Matsuoka, Jacob SchaeferAbstract:The homonuclear dipolar coupling of a directly bonded (13)C-(13)C pair has been used to create a dipolar double-quantum filter (D-DQF) to remove the natural-abundance (13)C background in (13)C[(2)H] Rotational-Echo Double-Resonance (REDOR) experiments. The most efficient version of this experiment has the D-DQF excitation and reconversion preceding the REDOR evolution period. Calculated and observed (13)C[(2)H]D-DQF-REDOR dephasings were in agreement for a test sample of mixed recrystallized labeled alanines.
Hellmut Eckert - One of the best experts on this subject based on the ideXlab platform.
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Ultraviolet Upconversion Luminescence in a Highly Transparent Triply-Doped Gd3+–Tm3+–Yb3+ Fluoride–Phosphate Glasses
The Journal of Physical Chemistry C, 2018Co-Authors: Gustavo Galleani, Silvia Helena Santagneli, Yannick Ledemi, Younes Messaddeq, Oliver Janka, Rainer Pöttgen, Hellmut EckertAbstract:We report near-infrared to ultraviolet (UV) upconversion emissions in triply-doped Gd3+–Tm3+–Yb3+ fluoride–phosphate glasses. Emission at 310 nm, originated from the Gd3+:6P7/2 → 8S7/2 transition, was observed for the first time in glasses. The high-purity glasses prepared exhibit extended transparency in the UV down to 200–250 nm. The mixed fluoride–phosphate environment of the rare-earth ions was characterized by means of NMR techniques using scandium as a diamagnetic mimic for the luminescent species, for which the ligand distribution was quantified by 45Sc{31P} rotational echo Double-Resonance NMR. Both the intensity of the Gd3+ emission as well as those of the UV emissions at 290, 347, and 363 nm increase with increasing ratio of fluoride to phosphate ligands coordinating to the rare-earth ion.
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Recoupling dipolar interactions with multiple I=1 quadrupolar nuclei: A 11B{6Li} and 31P{6Li} rotational echo double resonance study of lithium borophosphate glasses.
Solid state nuclear magnetic resonance, 2017Co-Authors: Lena Marie Funke, Henrik Bradtmüller, Hellmut EckertAbstract:The case of rotational echo double resonance (REDOR) experiments on the observe nuclei 11B and 31P interacting with multiple I=1 quadrupolar nuclei is analyzed in detail by SIMPSON simulations and experimental studies. The simulations define the region within the parameter space spanned by nutation frequency, quadrupolar coupling constant and spinning frequency where the parabolic analysis of the initial REDOR curve in terms of dipolar second moments has validity. The predictions are tested by experimental studies on the crystalline model compounds lithium diborate and lithium pyrophosphate, which are subsequently extended to measure dipolar second moments M2(11B{6Li}) and M2(31P{6Li}) in three borophosphate glasses. The data indicate that the lithium cations interact significantly more strongly with the phosphate than with the borate species, despite the formally anionic character of four-coordinate boron and the formally neutral character of the ultraphosphate (P(3)) units to which they are linked.
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Site discrimination in the crystalline borophosphate Na5B2P3O13 using advanced solid-state NMR techniques
Solid state nuclear magnetic resonance, 2007Co-Authors: Wenzel Strojek, Hellmut Eckert, Constanze Fehse, Bastian Ewald, Rüdiger KniepAbstract:A comprehensive solid-state NMR investigation on crystalline Na(5)B(2)P(3)O(13) is presented. Triple-quantum magic angle spinning (TQMAS) and rotational echo double resonance (REDOR) studies are used for accurate determinations of the (11)B, (23)Na and (31)P interaction parameters. Based on these results and complementary quantum mechanical calculations, plausible site assignments can be derived. Generally, the results show that detailed, quantitative information about structures in borophosphate compounds can be obtained by investigating both the local site environments characterized by chemical shift and quadrupolar interaction parameters and the correlated dipolar interactions to atoms in the second coordination sphere.
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Medium-range order in sodium phosphate glasses: a quantitative rotational echo double resonance solid state NMR study.
Physical chemistry chemical physics : PCCP, 2006Co-Authors: Wenzel Strojek, Hellmut EckertAbstract:Sodium ultraphosphate glasses (Na(2)O)(x)(P(2)O(5))(1-x) show a strongly non-linear dependence of the glass transition temperatures T(g)(x) on composition. To explore the structural origins of this behaviour, local and medium range ordering processes have been investigated by state-of-the-art (23)Na high-resolution and dipolar NMR spectroscopies. In particular, (31)P(23)Na) and (23)Na((31)P) rotational echo double resonance (REDOR) experiments have been analyzed to yield quantitative constraints for the structural description of these glasses. The sodium ions are found to be randomly distributed and, for x < 0.25, spatially correlated with a single metaphosphate-type Q((2)) unit at a distance of 330 pm. In this region, unusual compositional trends observed for the (23)Na chemical shifts and nuclear electric quadrupolar coupling constants, measured by triple-quantum magic-angle spinning (TQMAS) NMR, suggest a systematic decrease of Na coordination number with x. At higher sodium contents (x > 0.25), the magnitude of the (31)P((23)Na) dipolar interaction increases markedly, indicating a significantly increased extent of Q((2))-Na-Q((2)) crosslinking. Based on these results, a comprehensive description of medium-range order in sodium ultraphosphate glasses is developed, suggesting that the T(g)(x) dependence is closely linked to changes in the relative phosphorus/sodium distance distributions.
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Site connectivities in silver borophosphate glasses: new results from 11B{31P} and 31P{11B} rotational echo double resonance NMR spectroscopy.
Solid state nuclear magnetic resonance, 2005Co-Authors: Stefan Elbers, Wenzel Strojek, Ladislav Koudelka, Hellmut EckertAbstract:Local and medium range order in the glass system 50Ag2O-50[(B2O3)x-(P2O5)(1-x)] (x=0, 0.1, 0.2, 0.3, 0.4, 0.5, and 0.6) have been investigated by high-resolution solid state nuclear magnetic resonance (NMR) techniques. The detailed local site distribution has been derived from deconvolution analysis of the 11B and 31P magic-angle spinning (MAS) NMR signals. Quantitative information regarding the extent of boron-oxygen-phosphorus connectivity has been obtained on the basis of 11B[31P} and 31P{11B} rotational echo double resonance experiments. Incorporation of borate into silver metaphosphate glasses produces four-coordinate BO4/2- sites, which crosslink the metaphosphate chains, resulting in a significant increase in the glass transition temperature. Furthermore, the presence of borate favors the disproportionation of P(2) chain-like units into P(1) and P(3) sites, an effect not observed in binary alkali phosphate glasses. Finally, borate incorporation beyond x=0.3 results in the formation of neutral BO3/2 units, indicating some net charge transfer from the borate to the phosphate network former species. This latter result corresponds to the general metal ion scavenging effect observed for phosphate species in other mixed network former glass systems. In the present system, the effect is relatively moderate, however, suggesting that anionic BO4/2- groups are stabilized by the interaction with the phosphate groups.
Lynda M. Mcdowell - One of the best experts on this subject based on the ideXlab platform.
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Characterization of the complex of a trifluoromethyl-substituted shikimate-based bisubstrate inhibitor and 5-enolpyruvylshikimate-3-phosphate synthase by REDOR NMR.
Biochemistry, 2004Co-Authors: Lynda M. Mcdowell, Daniel R. Studelska, Robert D. O'connor, Barbara Poliks, Jacob SchaeferAbstract:A combination of (15)N[(19)F], (31)P[(15)N], and (31)P[(19)F] Rotational-Echo Double-Resonance NMR has been used to characterize the conformation of a bound trifluoromethylketal, shikimate-based bisubstrate inhibitor of 5-enolpyruvylshikimate-3-phosphate synthase. The solid-state NMR experiments were performed on the complex formed in solution and then lyophilized at low temperatures in the presence of stabilizing lyoprotectants. The results of these experiments indicate that none of the side chains of the six arginines that surround the active site forms a compact salt bridge with the phosphate groups of the bound inhibitor.
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Rotational-Echo Double-Resonance NMR-restrained model of the ternary complex of 5-enolpyruvylshikimate-3-phosphate synthase
Journal of Biomolecular NMR, 2004Co-Authors: Lynda M. Mcdowell, Daniel R. Studelska, Robert D. O'connor, Barbara Poliks, Denise D. Beusen, Jacob SchaeferAbstract:The 46-kD enzyme 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase catalyzes the condensation of shikimate-3-phosphate (S3P) and phosphoenolpyruvate to form EPSP. The reaction is inhibited by N-(phosphonomethyl)-glycine (Glp), which, in the presence of S3P, binds to EPSP synthase to form a stable ternary complex. We have used solid-state NMR and molecular modeling to characterize the EPSP synthase–S3P–Glp ternary complex. Modeling began with the crystal coordinates of the unliganded protein, published distance restraints, and information from the chemical modification and mutagenesis literature on EPSP synthase. New inter-ligand and ligand-protein distances were obtained. These measurements utilized the native ^31P in S3P and Glp, biosynthetically ^13C-labeled S3P, specifically ^13C and ^15N labeled Glp, and a variety of protein-^15N labels. Several models were investigated and tested for accuracy using the results of both new and previously published Rotational-Echo double resonance (REDOR) NMR experiments. The REDOR model is compared with the recently published X-ray crystal structure of the ternary complex, PDB code 1G6S. There is general agreement between the REDOR model and the crystal structure with respect to the global folding of the two domains of EPSP synthase and the relative positioning of S3P and Glp in the binding pocket. However, some of the REDOR data are in disagreement with predictions based on the coordinates of 1G6S, particularly those of the five arginines lining the binding site. We attribute these discrepancies to substantive differences in sample preparation for REDOR and X-ray crystallography. We applied the REDOR restraints to the 1G6S coordinates and created a REDOR-refined xray structure that agrees with the NMR results.
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Conformation of a bound inhibitor of blood coagulant factor Xa.
Biochemistry, 2003Co-Authors: Daniel R. Studelska, Lynda M. Mcdowell, Anil K. Mehta, Robert D. O'connor, David R. Light, William J. Guilford, Arnaiz Damian O, Marc Adler, Jerry L. Dallas, Jacob SchaeferAbstract:13C[(15)N] and (13)C[(19)F] Rotational-Echo Double-Resonance NMR have been used to characterize the enzyme-bound structure of ZK-816042, an amidine-imidazoline inhibitor of human factor Xa (FXa). The NMR experiments were performed on a lyophilized FXa-inhibitor complex. The complex was formed in solution in the presence of stabilizing excipients and frozen after gradual supercooling prior to lyophilization. The results indicate that the inhibitor binds with a distribution of orientations of the imidazoline ring.
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Human factor Xa bound amidine inhibitor conformation by double Rotational-Echo double resonance nuclear magnetic resonance and molecular dynamics simulations.
Journal of medicinal chemistry, 2003Co-Authors: Lynda M. Mcdowell, Margaret A. Mccarrick, Daniel R. Studelska, Robert D. O'connor, David R. Light, William J. Guilford, Arnaiz Damian O, Marc Adler, Jerry L. Dallas, Barbara PoliksAbstract:Double Rotational-Echo double resonance (double REDOR) NMR was used to investigate the conformation of a 13C-, 15N-, and 19F-labeled inhibitor (Berlex Biosciences compound no. ZK-806299) bound to h...
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rotational echo double resonance detection of cross links formed in mussel byssus under high flow stress
Journal of Biological Chemistry, 1999Co-Authors: Lynda M. Mcdowell, Luis A. Burzio, Herbert J Waite, Jacob SchaeferAbstract:Abstract 13C{2H} rotational echo double resonance NMR has been used to provide the first evidence for the formation of quinone-derived cross-links in mussel byssal plaques. Labeling of byssus was achieved by allowing mussels to filter feed from seawater containingl-[phenol-4-13C]tyrosine andl-[ring-d 4]tyrosine for 2 days. Plaques and threads were harvested from two groups of mussels over a period of 28 days. One group was maintained in stationary water while the other was exposed to turbulent flow at 20 cm/s. The flow-stressed byssal plaques exhibited significantly enhanced levels of 5, 5′-di-dihydroxyphenylalanine cross-links. The average concentration of di-dihydroxyphenylalanine cross-links in byssal plaques is 1 per 1800 total protein amino acid residues.
Daniel R. Studelska - One of the best experts on this subject based on the ideXlab platform.
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Characterization of the complex of a trifluoromethyl-substituted shikimate-based bisubstrate inhibitor and 5-enolpyruvylshikimate-3-phosphate synthase by REDOR NMR.
Biochemistry, 2004Co-Authors: Lynda M. Mcdowell, Daniel R. Studelska, Robert D. O'connor, Barbara Poliks, Jacob SchaeferAbstract:A combination of (15)N[(19)F], (31)P[(15)N], and (31)P[(19)F] Rotational-Echo Double-Resonance NMR has been used to characterize the conformation of a bound trifluoromethylketal, shikimate-based bisubstrate inhibitor of 5-enolpyruvylshikimate-3-phosphate synthase. The solid-state NMR experiments were performed on the complex formed in solution and then lyophilized at low temperatures in the presence of stabilizing lyoprotectants. The results of these experiments indicate that none of the side chains of the six arginines that surround the active site forms a compact salt bridge with the phosphate groups of the bound inhibitor.
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Rotational-Echo Double-Resonance NMR-restrained model of the ternary complex of 5-enolpyruvylshikimate-3-phosphate synthase
Journal of Biomolecular NMR, 2004Co-Authors: Lynda M. Mcdowell, Daniel R. Studelska, Robert D. O'connor, Barbara Poliks, Denise D. Beusen, Jacob SchaeferAbstract:The 46-kD enzyme 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase catalyzes the condensation of shikimate-3-phosphate (S3P) and phosphoenolpyruvate to form EPSP. The reaction is inhibited by N-(phosphonomethyl)-glycine (Glp), which, in the presence of S3P, binds to EPSP synthase to form a stable ternary complex. We have used solid-state NMR and molecular modeling to characterize the EPSP synthase–S3P–Glp ternary complex. Modeling began with the crystal coordinates of the unliganded protein, published distance restraints, and information from the chemical modification and mutagenesis literature on EPSP synthase. New inter-ligand and ligand-protein distances were obtained. These measurements utilized the native ^31P in S3P and Glp, biosynthetically ^13C-labeled S3P, specifically ^13C and ^15N labeled Glp, and a variety of protein-^15N labels. Several models were investigated and tested for accuracy using the results of both new and previously published Rotational-Echo double resonance (REDOR) NMR experiments. The REDOR model is compared with the recently published X-ray crystal structure of the ternary complex, PDB code 1G6S. There is general agreement between the REDOR model and the crystal structure with respect to the global folding of the two domains of EPSP synthase and the relative positioning of S3P and Glp in the binding pocket. However, some of the REDOR data are in disagreement with predictions based on the coordinates of 1G6S, particularly those of the five arginines lining the binding site. We attribute these discrepancies to substantive differences in sample preparation for REDOR and X-ray crystallography. We applied the REDOR restraints to the 1G6S coordinates and created a REDOR-refined xray structure that agrees with the NMR results.
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Conformation of a bound inhibitor of blood coagulant factor Xa.
Biochemistry, 2003Co-Authors: Daniel R. Studelska, Lynda M. Mcdowell, Anil K. Mehta, Robert D. O'connor, David R. Light, William J. Guilford, Arnaiz Damian O, Marc Adler, Jerry L. Dallas, Jacob SchaeferAbstract:13C[(15)N] and (13)C[(19)F] Rotational-Echo Double-Resonance NMR have been used to characterize the enzyme-bound structure of ZK-816042, an amidine-imidazoline inhibitor of human factor Xa (FXa). The NMR experiments were performed on a lyophilized FXa-inhibitor complex. The complex was formed in solution in the presence of stabilizing excipients and frozen after gradual supercooling prior to lyophilization. The results indicate that the inhibitor binds with a distribution of orientations of the imidazoline ring.
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Human factor Xa bound amidine inhibitor conformation by double Rotational-Echo double resonance nuclear magnetic resonance and molecular dynamics simulations.
Journal of medicinal chemistry, 2003Co-Authors: Lynda M. Mcdowell, Margaret A. Mccarrick, Daniel R. Studelska, Robert D. O'connor, David R. Light, William J. Guilford, Arnaiz Damian O, Marc Adler, Jerry L. Dallas, Barbara PoliksAbstract:Double Rotational-Echo double resonance (double REDOR) NMR was used to investigate the conformation of a 13C-, 15N-, and 19F-labeled inhibitor (Berlex Biosciences compound no. ZK-806299) bound to h...
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Rotational-Echo double resonance characterization of the effects of vancomycin on cell wall synthesis in Staphylococcus aureus
Biochemistry, 2002Co-Authors: Lynette Cegelski, Daniel R. Studelska, Robert D. O'connor, Sung Joon Kim, And Anil K. Mehta, Andrew W. Hing, Jacob SchaeferAbstract:Cross-polarization magic-angle spinning and Rotational-Echo double resonance 13C and 15N NMR experiments have been performed on intact cells of Staphylococcus aureus labeled with D-[1-13C]alanine and [15N]glycine or with [1-13C]glycine and L-[epsilon-15N]lysine. The cells were harvested during stationary or exponential growth conditions, the latter in media with and without the addition of vancomycin. The results of these experiments allowed the in situ determination of the relative concentrations of peptidoglycan cross-links (the number of peptide-stem D-alanines covalently linked to a pentaglycyl bridge) and bridge-links (the number of peptide-stem lysines covalently linked to a pentaglycyl bridge). The concentration of cross-links remained constant in the presence of vancomycin, whereas the number of bridge-links decreased. These changes suggest that vancomycin (at therapeutic levels) interrupts peptidoglycan synthesis in S. aureus by interference with transglycosylation.
Ray Dupree - One of the best experts on this subject based on the ideXlab platform.
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A High-Resolution 43Ca Solid-State NMR Study of the Calcium Sites of Hydroxyapatite.
Journal of the American Chemical Society, 2008Co-Authors: Danielle Laurencin, Ray Dupree, Alan Wong, John V. Hanna, Mark E. SmithAbstract:High resolution Ca-43 solid-state NMR studies of hydroxyapatite (Ca-10(PO4)(6)(OH2)) were performed at 14.1 T. The two crystallographically distinct calcium sites were unequivocally resolved by a triple-quantum magic angle spinning experiment, and the unambiguous assignment of the signals was possible using H-1-Ca-43 rotational echo double resonance and H-1-Ca-43 cross polarization magic angle spinning experiments.
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A High-Resolution 43Ca Solid-State NMR Study of the Calcium Sites of Hydroxyapatite.
Journal of the American Chemical Society, 2008Co-Authors: Danielle Laurencin, Ray Dupree, Alan Wong, John Hanna, Mark SmithAbstract:High resolution 43Ca solid-state NMR studies of hydroxyapatite (Ca10(PO4)6(OH)2) were performed at 14.1 T. The two crystallog. distinct calcium sites were unequivocally resolved by a triple-quantum magic angle spinning experiment, and the unambiguous assignment of the signals was possible using 1H-43Ca rotational echo double resonance and 1H-43Ca CPMAS experiments.
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Structural Studies of ZrV2-xPxO7 Solid Solutions Using 31P−{51V} and 51V−{31P} Rotational Echo Double Resonance NMR
The Journal of Physical Chemistry, 1996Co-Authors: Christopher Hudalla, Hellmut Eckert, Ray DupreeAbstract:The negative thermal expansion coefficients of ZrV2-xPxO7 solid solutions are attributed to the structural flexibility of M−O−M‘ dimers (M, M‘ = P, V), upon which the structure of these materials is based. The local environments of these M species have been probed by high-resolution 31P and 51V solid state NMR experiments, assisted by 51V−{31P} and 31P−{51V} rotational echo double resonance (REDOR) studies for assignment purposes. At all of the compositions studied, formation of mixed P−O−V dimers is distinctly preferred over the symmetric P−O−P and V−O−V species, indicative of chemical ordering. Analysis of the REDOR experiments, assuming simple spin 1/2 behavior of the 51V nuclei, yields an estimate of the P···V internuclear distance in the mixed dimers of 3.42 A, consistent with a linear geometry of the P−O−V units as required by the space group symmetry. This result is subject to the important caveat that a precise theoretical description of REDOR behavior involving quadrupolar nuclei is not yet avail...
