13C nuclear overhauser polarization-magic-angle spinning nuclear magnetic resonance spectroscopy in uniformly 13C-labeled solid proteins

A recently introduced (13)C polarization technique based on the nuclear Overhauser effect in rotating solid (nuclear Overhauser polarization-magic-angle spinning, NOP-MAS) (Takegoshi, K.; Terao, T. J. Chem. Phys. 2002, 117, 1700-1707) is applied to uniformly (13)C, (15)N-labeled proteins. NOP enhancement factors per scan of 1.5 approximately 2.0 are obtained, while that by cross polarization (CP) is less than 1.0. We show that uniform enhancement of all (13)C signals by CP is difficult to attain, while it is easily achieved by NOP, thus enabling quantitative comparison of signal intensities. NOP is easy to carry out under fast MAS and works well even for somewhat mobile molecules, for which CP does not work. Moreover, in labeled protein samples containing nonlabeled additives, NOP can eliminate the latter signals. For these features, NOP is superior to CP in many uniformly (13)C labeled proteins.     

13C and 199Hg nuclear magnetic resonance study of alkynyl mercury(II) compounds

Alkynyl mercury compounds of the type Hg(CCR)₂ (I), R′HgCCR (II) and R′HgCCHgR′ (III) have been studied by ¹H, ¹³C and ¹⁹⁹Hg NMR. Chemical shifts (δ¹H, δ¹³C, δ¹⁹⁹Hg) and coupling constants J(¹⁹⁹Hg¹H), J(¹⁹⁹Hg¹³C), J(¹³C¹H) and J(¹³C¹³C) (in natural abundance) are reported. The changes in magnitude of the coupling constants ¹J(¹⁹⁹Hg¹³C) and ¹J(¹³C¹³C) cannot be fully explained in terms of changes in the “s-character” of the HgC bond and the CC bond, respectively. The shielding of the alkynyl carbon linked to mercury in II is decreased by ca. 23 ppm as compared to the analogous carbon in I. This indicates a greatly different degree of polarization for the HgC bonds in I and II in agreement with the behaviour of ¹J(¹⁹⁹Hg¹³C) and ¹J(¹³C¹³C). The solvent and temperature dependence of the ¹⁹⁹Hg chemical shift of I (R = C₆H₅, C₄H₉ⁿ) and II (R = H, R′= CH₃) has been studied. The results indicate covalent interactions of I with amines, pyridine, dimethylsulphide and Br⁻, while the interaction with acetonitrile and oxygen donors (DMSO, DMF, dioxane, acetone) is of a different nature.     

Assignment of juvabione diastereomers by 13C nuclear magnetic resonance spectroscopy

The determination of structures and partial assignments of stereochemistry of juvabione and some of its analogues can be made on the basis of ¹³C nuclear magnetic resonance studies. The complete ¹³C n.m.r. spectral assignments for juvabione and five analogues are reported.     

Stabilization of the cyclopropenum and cyclopropenone ring systems by ferrocene : A 13C nuclear magnetic resonance study

The ¹³C NMR spectra of diferocenylcyclopropenone (1) and the triferrocenylcyclopropenium (2) and ferrocenyldiphenylcyclopropenium (3) cations in non-acidic media are analyzed with special reference to the information they afford concerning charge distribution in these species. A delocalization of the positive charge into the remote unsubstituted five-membered rings of 2 and 3 is indicated. The role of the metal as a conduit for this effect is suggested.     

13C nuclear magnetic resonance studies of cellulose acetate

¹³C-NMR spectra of ring carbons and O-acetyl carbonyl carbons of cellulose acetate (CA) in dimethyl sulfoxide-d₆ were analyzed. The CA samples with the degree of substitution (DS) ranging from 0.84 and 1.91 were prepared by homogeneous acetylation of cellulose with acetic anhydride in a 10% LiCl/dimethyl acetamide solvent. It was found that the use of these low DS samples permitted easier assignments not only of the ring carbon but also of the O-acetyl carbonyl carbon signals. The assignments were confirmed by comparing with the ¹H-NMR spectra of the samples obtained by complete acetylation of the corresponding CA samples with acetyl-d₃ chloride. Two methods for determining the distribution of O-acetyl groups of CA, i.e., the relative DS at the three different types of hydroxyl groups, were developed. One is based on the measurements of the relative intensities of the signals for the ring carbons and the other is based on the measurements of the relative intensities of the signals for the O-acetyl carbonyl carbons.     

13C nuclear magnetic resonance spectroscopy to determine olive oil grades

¹³C nuclear magnetic resonance spectroscopy was used in a first attempt to differentiate olive oil samples by grades. High resolution ¹³C NMR Distortionless Enhancement by Polarization Transfer (DEPT) spectra of 137 olive oil samples from the four grades, extra virgin olive oils, olive oils, olive pomace oils and lampante olive oils, were measured. The data relative to the resonance intensities (variables) of the unsaturated carbons of oleate (C-9 and C-10) and linoleate (L-9, L-10 and L-12) chains attached at the 1,3- and 2-positions of triacylglycerols were analyzed by linear discriminant analysis. The 1,3- and 2- carbons of the glycerol moiety of triacylglycerols along with the C-2, C-16 and C-18 resonance intensities of saturated, oleate and linoleate chains were also analyzed by linear discriminant analysis. The three discriminanting functions, which were calculated by using a stepwise variable selection algorithm, classified in the true group by cross-validation procedure, respectively, 76.9, 70.0, 94.4 and 100% of the extra virgin, olive oil, olive pomace oil and lampante olive oil grades.     

A study of the structure of chemical compounds by nuclear magnetic resonance spectroscopy

The PMR spectra of the condensation products of N-arylsulfonylmonoimino sulfur dioxide with the dienes were studied. It was established that the reaction goes according to the condensation pattern of the Diels-Alder reaction, involving the nitrogen sulfur double bond with the formation of 1-oxo-2-arylsulfonyl-3, 6-dihydro-1, 2-thiazines.     

A study of the structure of chemical compounds by nuclear magnetic resonance spectroscopy

The PMR spectra of the condensation products of N-arylsulfonylmonoimino sulfur dioxide with the dienes were studied. It was established that the reaction goes according to the condensation pattern of the Diels-Alder reaction, involving the nitrogen sulfur double bond with the formation of 1-oxo-2-arylsulfonyl-3, 6-dihydro-1, 2-thiazines.For part V, see [13].     

Inhibition and enhancement of resonance as studied by 13C nuclear magnetic resonance spectroscopy

¹³C chemical shifts are reported for a series of 2-substituted 1,3-dimethylbenzenes: comparisons of these values with those for the corresponding monosubstituted benzenes reveal, in some cases, large differences in the para-carbon substituent chemical shifts, which are attributable to steric hindrance of resonance. The questions of steric enhancement of resonance, and methoxy group conformation in certain anisoles are also studied by the ¹³C NMR technique. Studies of selected 2-substituted fluorenes are also reported, and substituent chemical shifts at carbon-7 (traversing eight bonds) of greater than 2ppm are observed. These effects are consistently greater than those reported for the corresponding biphenyl compounds, and are associated with planarity-enforced enhancement of resonance.     

17O and 13C nuclear magnetic resonance study of polyfluorinated carbonyl compounds

δ¹⁷O- and δ¹³C values are reported for 15 polyfluorinated carbonyl compounds. If the R_f group is separated from the carbonyl group by less than two carbon atoms a marked increase in the nuclear shielding of ¹³C(C=O) and a marked decrease in the nuclear shielding of the ¹⁷O(C=O) nucleus is observed. This is ascribed to the differing effects of the R_f group on the σ and the π system of the carbonyl unit. The effect on the σ-manifold leads to increase in shielding but it may be offset (as in the case of the ¹⁷O nucleus) by destabilization of the π system. UV spectroscopic data for some polyfluorinated carbonyl compounds support these arguments.     

Substrate affinities for membrane transport proteins determined by 13C cross-polarization magic-angle spinning nuclear magnetic resonance spectroscopy

We have devised methods in which cross-polarization magic-angle spinning (CP-MAS) solid-state NMR is exploited to measure rigorous parameters for binding of (13)C-labeled substrates to membrane transport proteins. The methods were applied to two proteins from Escherichia coli: a nucleoside transporter, NupC, and a glucuronide transporter, GusB. A substantial signal for the binding of methyl [1-(13)C]-beta-d-glucuronide to GusB overexpressed in native membranes was achieved with a sample that contained as little as 20 nmol of GusB protein. The data were fitted to yield a K(D) value of 4.17 mM for the labeled ligand and 0.42 mM for an unlabeled ligand, p-nitrophenyl beta-d-glucuronide, which displaced the labeled compound. CP-MAS was also used to measure binding of [1'-(13)C]uridine to overexpressed NupC. The spectrum of NupC-enriched membranes containing [1'-(13)C]uridine exhibited a large peak from substrate bound to undefined sites other than the transport site, which obscured the signal from substrate bound to NupC. In a novel application of a cross-polarization/polarization-inversion (CPPI) NMR experiment, the signal from undefined binding was eliminated by use of appropriate inversion pulse lengths. By use of CPPI in a titration experiment, a K(D) value of 2.6 mM was determined for uridine bound to NupC. These approaches are broadly applicable to quantifying binding of substrates, inhibitors, drugs, and antibiotics to numerous membrane proteins.     

Solid-state 13C nuclear magnetic resonance characterization of MDI-based polyurethanes

¹³C solid-state nuclear magnetic resonance (NMR) experiments on linear polyurethanes and poly(ether-urethane) block copolymers demonstrate that ¹³C spin-lattice relaxation experiments in the laboratory [T₁(C)] and rotating [T_1p(C)] frames provide the most information about domain morphology in these microphase-separated polymer systems. T₁(H) TCH, and T_1p(H) data are less useful in a 4,4′-methylene bis(p-phenyl isocyanate)-1,4-butanediol (MDI/BD) hard-segment material, the MDI bridging methylene and the MDI urethane carbonyl T₁(C and T_1p(C) times fall in characteristic ranges for crystalline, amorphous, interfacial, and dissolved species. BD methylene carbons have short T_1p(C) for crystalline and long T_1p(C) for amorphous hard-segment aggregates. The distinct T_1p(C) and T₁(C) fractins observed are attributed to the presence of several crystalline polymorphs. Both T₁(C) results and DSC endotherms indicate that the crystalline polymorphs present in the poly(ether-urethane) are less ordered than the types seen in the pure hard-segment material. © 1993 John Wiley & Sons, Inc.     

The study of domain structure in polyurethanes by nuclear magnetic resonance

Polyurethanes are block copolymers composed of hard and soft segments which often undergo microphase separation with the resultant formation of hard and soft domains. The nuclear-magnetic-free induction decay of such systems is shown to consist of a fast Gaussian component corresponding to glassy domains and a slow exponential component corresponding to rubbery domains. Thus, by being sensitive to the microscopic mobility of a material, nuclear magnetic resonance (NMR) has the potential to determine when a polyurethane has separated into domains and the relative amounts of material in each domain. After annealing the free-induction decay of a linear polyurethane showed a distribution of relaxation times indicating that mixing of the domains had taken place and a continuum of compositions exist in the material. The free-induction decay of a cross-linked polyurethane was unaffected by annealing, confirming the results of an earlier study.