A fourier transform infrared spectroscopic and solid-state NMR study of bone mineral in osteogenesis imperfecta.

Fourier transform infrared spectroscopy and 31P solid-state nuclear magnetic resonance spectroscopy were used to determine if any structural or compositional differences in osteogenesis imperfecta (OI) bone mineral could be detected that might help to explain the bone fragility observed in this disease. A previous study by Cassella et al. used an electron probe X-ray microanalytical technique to compare the calcium to phosphorus (Ca/P) molar ratios in normal bone and bone from patients with OI. It was demonstrated that bone from OI patients had a lower Ca/P molar ratio. This study demonstrated that OI bone mineral had a general hydroxyapatite structure and that isomorphous substitutions in the carbanoapatite lattice could account for the low Ca/P molar ratio.

13C CP/MAS NMR Studies of Hemoprotein Models with and without an Axial Hindered Base: (13)C Shielding Tensors and Comparison with Hemoproteins and X-ray Structural Data.

13C cross-polarization magic-angle-spinning (CP/MAS) NMR spectra of several carbonmonoxide (93-99% (13)C enriched) hemoprotein models with 1,2-dimethylimidazole (1,2-diMeIm) and 1-methylimidazole (1-MeIm) as axial ligands are reported. This enables the (13)CO spinning sideband manifold to be measured and hence the principal components of the (13)CO chemical shift tensor to be obtained. Negative polar interactions in the binding pocket of the cap porphyrin model and inhibition of Fe-->CO back-donation result in a reduction in shielding anisotropy; on the contrary, positive distal polar interactions result in an increase in the shielding anisotropy and asymmetry parameter in some models. It appears that the axial hindered base 1,2-dimethylimidazole has little direct effect on the local geometry at the CO site, despite higher rates of CO desorption being observed for such complexes. This suggests that the mechanism by which steric interactions are released for the 1,2-diMeIm complexes compared to 1-MeIm complexes does not involve a significant increase in bending of the Fe-C-O unit. The asymmetry of the shielding tensor of all the heme model compounds studied is smaller than that found for horse myoglobin and rabbit hemoglobin.

31P solid-state NMR measurements used to detect interactions between NADPH and water and to determine the ionisation state of NADPH in a protein-ligand complex subjected to low-level hydration.

31P-NMR spectra of NADPH and NADPH bound to Lactobacillus casei dihydrofolate reductase have been recorded using the techniques of cross-polarization, magic-angle spinning and high-power proton-decoupling on both lyophilized and hydrated samples. Previous studies on the lyophilized complex of L. casei dihydrofolate reductase with NADPH and methotrexate, measuring the isotropic shifts and principal components of the chemical shift tensors, have shown that the 2'-phosphate group of bound NADPH exists as a mixture of the dianionic and monoanionic states [Gerothanassis, I. P, Barrie, P. J., Birdsall, B. & Feeney, J. (1994) Eur J. Biochem. 226, 211-218]. In the present study on hydrated samples, the characterization of the isotropic shift and chemical shift tensors of the 2'-phosphate signal indicates that the 2'-phosphate is almost exclusively in the dianionic state. This is in agreement with earlier 31P-NMR studies in solution [Feeney, J., Birdsall, B., Roberts, G. C. K. & Burgen, A. S. V. (1975) Nature 257, 564-566]. In experiments examining progressively hydrated (6%, 12%, 15%, by mass) samples, the observed signals become increasingly narrower probably because the microenvironments of the 31P nuclei become more homogeneous upon sample hydration. Chemical exchange between mobile water molecules and bound protons close to individual sites on NADPH has been indirectly monitored on a hydrated sample (15% water, by mass) using a pulse sequence proposed by Harbison and coworkers [Harbison, G. S., Roberts, J. E., Herzfeld, J. & Griffin, R. G. (1988) J. Am. Chem. Soc. 110, 7221-7223]. In this experiment, the two diphosphate signals are totally suppressed while the 2'-phosphate phosphorus signal remains: this indicates a significant polarization of the 2'-phosphate nuclei from protons in exchange with those of mobile water molecules.

31P-NMR studies of NADPH, NADP+ and the complex of NADPH and methotrexate with Lactobacillus casei dihydrofolate reductase in the solid state.

31P-NMR spectra on solid samples of NADP+, NADPH and NADPH bound to Lactobacillus casei dihydrofolate reductase have been recorded using the techniques of cross polarisation, magic angle spinning and high power proton decoupling. The isotropic chemical shifts, the principal components of the shielding tensors and the asymmetry parameters for the 31P nuclei in the 2'-phosphate and pyrophosphate groups have been measured. The isotropic shifts show similar trends to the chemical shifts measured in solution. The isotropic shifts and the shielding tensors for the dianionic and monoanionic states of the 2'-phosphate group have been determined and the presence of both ionisation states has been detected in a solid sample of the lyophilised complex of L. casei dihydrofolate reductase with NADPH and methotrexate. This contrasts with the behaviour in solution, where only the dianionic form is bound to the enzyme. The signals from the two pyrophosphates 31P nuclei in bound NADPH were resolved and identified. The asymmetry parameters in the different ionisation states and the orientations of the shielding tensors within the molecular framework are considered in the context of previous 31P studies on phosphate-containing compounds.

Solid-state 27Al NMR studies of aluminophosphate molecular sieves. Enhanced resolution by quadrupole nutation and double-rotation.

Solid-state 27Al NMR spectra of several aluminophosphate molecular sieves have been recorded with conventional magic-angle spinning (MAS), double-rotation (DOR) and quadrupole nutation with fast MAS. Enhanced resolution was obtained in the quadrupole nutation experiment at certain radiofrequency pulse strengths. This extra resolution can be comparable to that attainable using DOR, and does not introduce spinning sidebands.