Measurement of long-range cross-correlation rates using a combination of single- and multiple-quantum NMR spectroscopy in one experiment

A method is described to determine long-range cross-correlations between the modulations of an anisotropic chemical shift (e.g., of a C' carbonyl carbon in a protein) and the fluctuations of a weak long-range dipolar interaction (e.g., in cross-correlation between the same C' carbonyl and the H(N) proton of the neighboring amide group). Such long-range correlations are difficult to measure because the corresponding long-range scalar couplings are so small that Redfield's secular approximation is often violated. The method, which combines features of single- and double-quantum NMR spectroscopy, allows one to cancel the effects of dominant short-range dipolar interactions (e.g., between the CSA of the amide nitrogen N and the dipolar coupling to its attached proton H(N)) and is designed so that the secular approximation is rescued even if the scalar coupling between the long-range dipolar coupling partners is very small. The cross-correlation rates thus determined in ubiquitin cover a wide range because of local motions and variations of the CSA tensors.     

A new spin probe of protein dynamics: nitrogen relaxation in 15N-2H amide groups

(15)N spin relaxation data have provided a wealth of information on protein dynamics in solution. Standard R(1), R(1)(rho), and NOE experiments aimed at (15)N[(1)H] amide moieties are complemented in this work by HA(CACO)N-type experiments allowing the measurement of nitrogen R(1) and R(1)(rho) rates at deuterated (15)N[(2)D] sites. Difference rates obtained using this approach, R(1)((15)N[(1)H]) - R(1)((15)N[(2)D]) and R(2)((15)N[(1)H]) - R(2)((15)N[(2)D]), depend exclusively on dipolar interactions and are insensitive to (15)N CSA and R(ex) relaxation mechanisms. The methodology has been tested on a sample of peptostreptococcal protein L (63 residues) prepared in 50% H(2)O-50% D(2)O solvent. The results from the new and conventional experiments are found to be consistent, with respect to both local backbone dynamics and overall protein tumbling. Combining several data sets permits evaluation of the spectral density J(omega(D) + omega(N)) for each amide site. This spectral density samples a uniquely low frequency (26 MHz at a 500 MHz field) and, therefore, is expected to be highly useful for characterizing nanosecond time scale local motions. The spectral density mapping demonstrates that, in the case of protein L, J(omega(D) + omega(N)) values are compatible with the Lipari-Szabo interpretation of backbone dynamics based on the conventional (15)N relaxation data.     

Evidence for slow motion in proteins by multiple refocusing of heteronuclear nitrogen/proton multiple quantum coherences in NMR

A novel NMR method characterizes slow motions in proteins by multiple refocusing of double- and zero-quantum coherences of amide protons and nitrogen-15 nuclei. If both nuclei experience changes in their isotropic chemical shifts because of internal motions on slow time scales (mus - ms), this leads to a difference in the relaxation rates of double- and zero-quantum coherences. This is due to CSM/CSM (chemical shift modulation) cross-correlation effects that are related to the well-known chemical exchange contribution Rex to the decay rate R2 = 1/T2 of nitrogen-15 nuclei. The CSM/CSM contributions can be distinguished from other mechanisms through their dependence on the repetition rate of a Carr-Purcell-Meiboom-Gill (CPMG) multiple refocusing sequence. In ubiquitin, motional processes can be identified that could hitherto not be observed by conventional CPMG nitrogen-15 NMR.     

Determination of chemical shift anisotropy tensors of carbonyl nuclei in proteins through cross-correlated relaxation in NMR.

The principal components and orientations of the chemical shift anisotropy (CSA) tensors of nearly all 13C carbonyl nuclei in a small protein have been determined in isotropic solution by a combination of three complementary cross-correlation measurements.     

CAESURA: measurement of slow molecular dynamics by solid-state nuclear magnetic resonance chemical shift anisotropy modulation amplification

An alternative magic angle spinning (MAS) exchange NMR experiment based on chemical shift anisotropy (CSA) amplification is described. The CSA amplification experiment correlates a standard MAS spectrum in the omega(2) dimension with a sideband pattern in omega(1) in which the intensities are identical to those expected for a sample spinning at some fraction 1N of the actual rate omega(r). In common with 2D-PASS, the isotropic shift appears only in the omega(2) dimension, and long acquisition times can be avoided without loss of resolution of different chemical sites. The new CSA amplification exchange experiment provides information about the time scale and geometry of molecular motions via their effect on the sideband intensities in a one-dimensional pattern. The one-dimensional patterns from different chemical sites are separated across two frequency dimensions according to the isotropic shifts.     

Determinations of 15N chemical shift anisotropy magnitudes in a uniformly 15N,13C-labeled microcrystalline protein by three-dimensional magic-angle spinning nuclear magnetic resonance spectroscopy

Amide 15N chemical shift anisotropy (CSA) tensors provide quantitative insight into protein structure and dynamics. Experimental determinations of 15N CSA tensors in biologically relevant molecules have typically been performed by NMR relaxation studies in solution, goniometric analysis of single-crystal spectra, or slow magic-angle spinning (MAS) NMR experiments of microcrystalline samples. Here we present measurements of 15N CSA tensor magnitudes in a protein of known structure by three-dimensional MAS solid-state NMR. Isotropic 15N, 13C alpha, and 13C' chemical shifts in two dimensions resolve site-specific backbone amide recoupled CSA line shapes in the third dimension. Application of the experiments to the 56-residue beta1 immunoglobulin binding domain of protein G (GB1) enabled 91 independent determinations of 15N tensors at 51 of the 55 backbone amide sites, for which 15N-13C alpha and/or 15N-13C' cross-peaks were resolved in the two-dimensional experiment. For 37 15N signals, both intra- and interresidue correlations were resolved, enabling direct comparison of two experimental data sets to enhance measurement precision. Systematic variations between beta-sheet and alpha-helix residues are observed; the average value for the anisotropy parameter, delta (delta = delta(zz) - delta(iso)), for alpha-helical residues is 6 ppm greater than that for the beta-sheet residues. The results show a variation in delta of 15N amide backbone sites between -77 and -115 ppm, with an average value of -103.5 ppm. Some sites (e.g., G41) display smaller anisotropy due to backbone dynamics. In contrast, we observe an unusually large 15N tensor for K50, a residue that has an atypical, positive value for the backbone phi torsion angle. To our knowledge, this is the most complete experimental analysis of 15N CSA magnitude to date in a solid protein. The availability of previous high-resolution crystal and solution NMR structures, as well as detailed solid-state NMR studies, will enhance the value of these measurements as a benchmark for the development of ab initio calculations of amide 15N shielding tensor magnitudes.     

Pure and Disiline-doped (6,0) AIN Nanotube: A Computational DFT Study on the NMR Parameters

Nuclear Magnetic Resonance (NMR) parameters including isotropic and anisotropic chemical shielding parameters (CSI, CSA) and electronic structures were calculated using Density Functional Theory (DFT) for Disiline-doped Aluminum Nitride Nanotubes (Disiline-AlNNTs). The 27Al and 15N nuclear magnetic resonance (NMR) was calculated by means of the GIAO, CSGT, and IGAIM methods. Geometry optimizations were carried out at the B3LYP/6-311+G* level of theory using the Gaussian 98 program suite. The calculated parameters indicate that the Al and N atoms located at the mouths of nanotube have the smallest and largest chemical shielding isotropic (CSI) values among those of other identical ones, respectively. In the Disiline-doped model, the NMR parameters of those nuclei directly bonded to the C and Si atoms show significant changes, while other nuclei changes are inferior.     

Synthesis, structure and electrochemistry of ferrocene-peptide macrocycles

Redox active cyclopeptides Fc[CSA]2 (5), Fc[Gly-CSA]2 (6), Fc[Ala-CSA]2 (7), Fc[Val-CSA](2) and Fc[Leu-CSA]2 (9) (CSA = cysteamine) which are formed by the reaction of ferrocenedicarboxylic acid with peptide cystamines at high dilutions. These systems exhibit H-bonding involving the amide NH in solution as shown by their temperature dependent NMR spectra. With the exception of 5, the ferrocene macrocycles display intramolecular N...O cross-ring H-bonding in the solid state involving the amino acids proximal to the ferrocene.     

113Cd Shielding Tensors of Monomeric Cadmium Compounds Containing Nitrogen Donor Atoms. 3. CP/MAS Studies on Five-Coordinate Cadmium Complexes Having N(3)X(2) (X = H, N, O, and S) Donor Atoms

The principal elements of the (113)Cd shielding tensor for a set of five- coordinate compounds having mixed donor atoms coordinating to the cadmium were determined via CP/MAS NMR experiments. The first complex, [HB(3,5-Me(2)pz)(3)]CdBH(4) (where pz = pyrazolyl), has a CdN(3)H(2) inner coordination sphere. The isotropic chemical shift in the solid state is 355.1 ppm, and its chemical shift anisotropy (CSA, Deltasigma) is -596 ppm with an asymmetry parameter (eta) of 0.64. The second complex, [HB(3,5-Me(2)pz)(3)]Cd[H(2)B(pz)(2)], has five nitrogen donor atoms bonded to the cadmium. This N(5) or N(3)N(2) compound was the only material of this study to manifest dipolar splitting of the cadmium resonance from the quadrupolar (14)N. The isotropic chemical shift, CSA, and the value of eta for this material were therefore determined at higher field where the dipolar splitting was less than the linewidth, yielding values of 226.6 ppm, -247 ppm, and 0.32, respectively. A second N(5) material, [HB(3-Phpz)(3)]Cd[H(2)B(3,5-Me(2)pz)(2)], was also investigated and has an isotropic shift of 190.2 ppm, a CSA of 254 ppm, and an eta of 0.86. Also studied was [HB(3-Phpz)(3)]Cd[(Bu(t)CO)(2)CH], which has an CdN(3)O(2) inner core. The isotropic chemical shift of this complex is 173.6 ppm, and the values of Deltasigma and eta were determined to be -258 ppm and 0.38, respectively. The final compound, [HB(3,5-Me(2)pz)(3)]Cd[S(2)CNEt(2)], with N(3)S(2) donor atoms, has an isotropic shift of 275.8 ppm, an eta of 0.51, and a CSA of +375 ppm. Utilizing previous assignments, the most shielded tensor element was determined to be oriented normal to the plane of the tridentate ligand. The shielding tensor information is used to speculate on the coordination geometry of the CdN(3)O(2) inner core complex.     

Interaction of Cd and Zn with biologically important ligands characterized using solid-state NMR and ab initio calculations

For the first time, coordination geometry and structure of metal binding sites in biologically relevant systems are studied using chemical shift parameters obtained from solid-state NMR experiments and quantum chemical calculations. It is also the first extensive report looking at metal-imidazole interaction in the solid state. The principal values of the (113)Cd chemical shift anisotropy (CSA) tensor in crystalline cadmium histidinate and two different cadmium formates (hydrate and anhydrate) were experimentally measured to understand the effect of coordination number and geometry on (113)Cd CSA. Further, (13)C and (15)N chemical shifts have also been experimentally determined to examine the influence of cadmium on the chemical shifts of (15)N and (13)C nuclei present near the metal site in the cadmium-histidine complex. These values were then compared with the chemical shift values obtained from the isostructural bis(histidinato)zinc(II) complex as well as from the unbound histidine. The results show that the isotropic chemical shift values of the carboxyl carbons shift downfield and those of amino and imidazolic nitrogens shift upfield in the metal (Zn,Cd)-histidine complexes relative to the values of the unbound histidine sample. These shifts are in correspondence with the anticipated values based on the crystal structure. Ab initio calculations on the cadmium histidinate molecule show good agreement with the (113)Cd CSA tensors determined from solid-state NMR experiments on powder samples. (15)N chemical shifts for other model complexes, namely, zinc glycinate and zinc hexaimidazole chloride, are also considered to comprehend the effect of zinc binding on (15)N chemical shifts.     

14N NMR studies on an optically isotropic liquid crystalline D phase of 4'-n-alkoxy-3'-nitrobiphenyl-4-carboxylic acids (ANBC)

14N nuclear magnetic resonance (NMR) measurements have been carried out for three members of 4'-n-alkoxy-3'-nitrobiphenyl-4-carboxylic acids (ANBC-n, where the number of carbon atoms in the alkoxy group, n, is 14, 16, and 22) in the temperature range 400-500 K. ANBC-16 and-22 show an optically isotropic D phase. The 14N NMR spectrum of the D phase showed a single peak, which may result from isotropically averaged quadrupole interactions around the 14N nucleus. Relaxation time measurements indicate the existence of two relaxational processes, faster anisotropic and slower isotropic motions, and suggest that in both cases ANBC molecules act as a dimer. The present 14N NMR results may be interpreted in the framework of the IPJR model, indicating that the structure of the D phase is a three-dimensional network continuous over the unit lattice.     

Predicting 15N amide chemical shifts in proteins. I. An additive model for the backbone contribution

Because proteins adopt unique structures, chemically identical nuclei in proteins exhibit different chemical shifts. Amide ¹⁵N chemical shifts have been shown to vary over 20 ppm. The cause of these chemical shift inequivalencies is the different intra‐ and intermolecular interactions that individual nuclei experience at different locations in the protein structure. These chemical shift inequivalencies can be described as structural shifts, the difference between the actual chemical shift and the random coil chemical shift. As a first step toward the prediction of these amide ¹⁵N structural shifts, calculations have been carried out on acetyl‐glycine‐methyl amide to examine how a neighboring peptide group influences the amide ¹⁵N structural shifts. The ϕ,ψ dihedral angle space is completely surveyed, while all other geometrical variables are held fixed, to isolate the effect of the backbone conformation. Similar calculations for a limited number of conformations of acetyl‐glycine‐glycine‐methyl amide were carried out, where the effects of the two terminal peptide groups on the central amide ¹⁵N structural shift are examined. It is shown that the effect of the two adjacent groups can be accurately modeled by combining their individual effects additively. This provides a quite simple method to predict the backbone influence on amide ¹⁵N structural shifts in proteins. © 2001 John Wiley & Sons, Inc. J Comput Chem 22: 366–372, 2001     

Reintroducing anisotropic interactions in magic-angle-spinning NMR of half-integer quadrupolar nuclei: 3D MQMAS

Selective reintroduction of anisotropic interactions such as the chemical shift anisotropy (CSA) and homonucler dipolar (HMD) coupling were implemented in a high-resolution NMR spectroscopy for half-integer quadrupolar nuclei. Rotary resonance recoupling (R(3)) combined with the multiple-quantum magic-angle spinning (MQMAS) in a three-dimensional (3D) experiment provides not only site-specific high-resolution spectra to yield the quadrupolar interaction parameters but also the CSA or HMD interaction parameters. This 3D experiment provides an avenue for the complete local structural information of half-integer quadrupolar nuclei. Three-dimensional MQMAS experiments incorporating R(3) of HMD and CSA interactions were demonstrated on model compounds containing (11)B, (23)Na, and (87)Rb nuclei.     

Using the 19F NMR chemical shift anisotropy tensor to differentiate between the zigzag and chiral forms of fluorinated single-walled carbon nanotubes

The structural characterization of different kinds of zigzag and chiral single-walled carbon nanotubes (SWNTs) has been investigated theoretically using (19)F NMR spectroscopy. The chemical shift anisotropy (CSA) tensor is computed at different levels of theory for the (19)F nuclei in different forms of functionalized fluorinated carbon nanotubes (CNT). A set of fluorine CSA parameters comprising the span, skew, and isotropic chemical shift is computed for each form of the fluoronanotubes and multidimensional CSA parameter correlation maps are constructed. We show that these correlations are able to clearly distinguish between the chiral and zigzag forms of fluorinated carbon nanotubes (F-SWNTs). Implications for solid-state and liquid-state NMR experiments are discussed.     

Time-domain calculations of the infrared and polarized Raman spectra of tetraalanine in aqueous solution

The IR and polarized (isotropic and anisotropic) Raman spectra are calculated for the amide I band of tetraalanine ((Ala)4) in aqueous solution by using a time-domain computational method, which includes the effects of the diagonal frequency modulations (of individual peptide groups), the off-diagonal (interpeptide) vibrational couplings, and structural dynamics. It is shown that the calculated band profiles, especially the existence of a large negative noncoincidence effect (i.e., large frequency separations between the IR, isotropic Raman, and anisotropic Raman bands, with the isotropic Raman being higher in frequency), are in reasonable agreement with the experimental results. This negative noncoincidence effect derives from two conditions: the positive coupling between the amide I vibrations of peptide groups and the angle larger than 90 degrees between the transition dipoles of the coupled vibrations. This result means that the dynamically changing structures mainly in the polyproline II and beta-type conformations containing some repeated interconversions obtained from the molecular dynamics calculation are consistent with the existence of a large negative noncoincidence effect, as far as the structures satisfy the above two conditions. It is also shown that the electric fields from solvent water molecules induce larger frequency shifts than those of intrachain interactions, with rapid underdamped oscillatory modulations ( approximately 43 fs) due to the librational motions of water molecules that give rise to motional narrowing effect on the spectra. The reason for the difference from the behavior seen for the O-H stretching mode of liquid water is discussed. The time-domain analysis of the mode identity shows that the system proceeds halfway to complete mode mixing with a similar time scale ( approximately 60 fs), suggesting the importance of the nonadiabatic effect, which is included in a natural way in the present computational method.     

Dynamics of large elongated RNA by NMR carbon relaxation

We present an NMR strategy for characterizing picosecond-to-nanosecond internal motions in uniformly 13C/15N-labeled RNAs that combines measurements of R1, R1rho, and heteronuclear 13C{1H} NOEs for protonated base (C2, C5, C6, and C8) and sugar (C1') carbons with a domain elongation strategy for decoupling internal from overall motions and residual dipolar coupling (RDC) measurements for determining the average RNA global conformation and orientation of the principal axis of the axially symmetric rotational diffusion. TROSY-detected pulse sequences are presented for the accurate measurement of nucleobase carbon R1 and R1rho rates in large RNAs. The relaxation data is analyzed using a model free formalism which takes into account the very high anisotropy of overall rotational diffusion (Dratio approximately 4.7), asymmetry of the nucleobase CSAs and noncollinearity of C-C, C-H dipolar and CSA interactions under the assumption that all interaction tensors for a given carbon experience identical isotropic internal motions. The approach is demonstrated and validated on an elongated HIV-1 TAR RNA (taum approximately 18 ns) both in free form and bound to the ligand argininamide (ARG). Results show that, while ARG binding reduces the amplitude of collective helix motions and local mobility at the binding pocket, it leads to a drastic increase in the local mobility of "spacer" bulge residues linking the two helices which undergo virtually unrestricted internal motions (S2 approximately 0.2) in the ARG bound state. Our results establish the ability to quantitatively study the dynamics of RNAs which are significantly larger and more anisotropic than customarily studied by NMR carbon relaxation.     

Exact, Born-Oppenheimer, and quantum-chemistry-like calculations in helium clusters doped with light molecules: The He2N2(X) system

Helium clusters doped with diatomic molecules, He(N)-BC, have been recently studied by means of a quantum-chemistry-like approach. The model treats He atoms as "electrons" and dopants as "nuclei" in standard electronic structure calculations. Due to the large mass difference between He atoms and electrons, and to the replacement of Coulomb interactions by intermolecular potentials, it is worth assessing up to what extent are the approximations involved in this model, i.e., decoupling of the BC rotation from the He-atom orbital angular momenta and Born-Oppenheimer separation of the BC stretch versus the He motions, accurate enough. These issues have been previously tackled elsewhere for the (4)He(2)-Br(2)(X) system, which contains a heavy dopant [Roncero et al., Int. J. Quantum Chem. 107, 2756 (2007)]. Here, we consider a similar cluster but with a much lighter dopant such as N(2)(X). Although the model does not provide the correct energy levels for the cluster, positions and intensities of the main detectable lines of the vibrotational Raman spectrum at low temperature are accurately reproduced.     

Electron spin echo envelope modulation of trapped radicals in disordered systems: nitrogen modulation from isotopically substituted chlorophyll a cations

The electron spin echo envelope modulation of the chlorophyll a radical cation has been examined for radicals containing ¹⁴N and ¹⁵N. The modulation is found to be due primarily to the nitrogen nuclei in the heterocycle and the nuclear quadrupole interaction plays a large part in determining the modulation from ¹⁴N. The modulation from ¹⁵N allows limits to be set on the isotropic and anisotropic hyperfine interactions.     

1H, 2H and 13C NMR spectra and molecular dynamics of some solid alkylene-aromatic polyesters

Molecular dynamics of some mesomorphic main–chain alkylene–aromatic polyesters have been investigated by means of NMR spectra of various nuclei over a wide temperature range. In solid polymers regions of different molecular mobilities coexist and their fractions are determined by the sample temperature and thermal history. The sample annealing leads to the growth of rigid fraction. It was found that below the glass transition temperature the only forms of large–scale mobility are the torsional vibrations and flips of para–phenylene groups, while spacer groups are virtually rigid. Above the glass temperature almost all phenylene rings undergo flipping motions and methylene groups of the spacer take part in complicated motions of both anisotropic and isotropic character.     

Oxygen decorating at the one ring and at the N-mouth of (10, 0) aluminum nitride nanotube: A DFT investigation

We have investigated oxygen decorating in the (10, 0) aluminum nitride nanotube (AlNNT) by density functional theory. Band gaps, total (TDOS) and partial (PDOS) densities of state and chemical-shielding isotropic (CSI) and chemical-shielding anisotropic (CSA) have been calculated or determined in three models of the investigated (10, 0) AlNNT: pristine (model.0), O-decorating at the one ring in the middle of AlNNT (Model.1) and O-decorating at the nitrogen mouth of AlNNT (Model.2). The results indicated that the dipole moment does not detect the significant effects of dopant whereas TDOS, PDOS and band gap energies detect notable effects. The CSI and CSA values for the Al and N atoms-contributed to the Al-O bonds or those atoms close to the decorated region, in both models of O-decorated AlNNTs are changed.