Nuclear Overhauser effect spectra

Keepers, J.W. and James, T.L., A theoretical smdy of distance determinations from NMR two-dimensional nuclear Overhauser effect spectra, J. Mag. Resort, 57, 404, 1984. Kemp, W., Organic Spectroscopy, 3rd ed., W.H. Ereeman, New York, 1991. Figueiredo, P. et al., New aspects of anthocyanin complexation intramolecular copigmentation as a means for colour loss Phytochemistry, 41, 301, 1996. 

Distance Determinations from NMR. Two-Dimensional Nuclear Overhauser Effect Spectra. 

P. Yip and D. A. Case, y. Magn. Reson., 83, 643 (1989). A New Method for Refinement of Macromolecular Structures Based on Nuclear Overhauser Effect Spectra. 

Structure Determination via Complete Relaxation Matrix Analysis (CORMA) of Two-Dimensional Nuclear Overhauser Effect Spectra DNA Fragments... 

Most of protein structural information from NMR is obtained in the form of nuclear Overhauser effects or NOEs between pairs of protons that are less than 6 A apart through space. An NOE between a spin pair carries distance information, but only short distances are observed because NOEs have an inverse sixth power dependence on distance. However, the distance cannot be uniquely determined given a measured NOE intensity without making some assumption about the environment of the spin pair and the motion of the vector between them. The simplest model for obtaining a distance from cross peak intensities in a nuclear Overhauser effect spectrum (NOESY) is the isolated ri d spin pair (RRNN - rigid rotor nearest neither) approximation (Jardetzky and Roberts, 1981). In this approximation the observed cross peak intensity, which is proportional to the cross relaxation rate, is related to a sini e intemudear distance, r. 

Methods of disturbing the Boltzmann distribution of nuclear spin states were known long before the phenomenon of CIDNP was recognized. All of these involve multiple resonance techniques (e.g. INDOR, the Nuclear Overhauser Effect) and all depend on spin-lattice relaxation processes for the development of polarization. The effect is referred to as dynamic nuclear polarization (DNP) (for a review, see Hausser and Stehlik, 1968). The observed changes in the intensity of lines in the n.m.r. spectrum are small, however, reflecting the small changes induced in the Boltzmann distribution. 

Gated decoupling The decoupler is gated during certain pulse NMR experiments, so spin decoupling occurs only when the decoupler is switched on and not when it is switched off used to eliminate either H- C spincoupling or nuclear Overhauser effect in a ID C spectrum, and employed as a standard technique in many other H-NMR experiments, such as APT and y-resolved. 

As with the COSY experiment, the sequence starts with a pulse followed by an evolution period, but now the mechanism that couples the two spins (which must be in close proximity, typically <6 A) is the Nuclear Overhauser Effect (NOE). The second pulse converts magnetization into population disturbances, and cross-relaxation is allowed during the mixing time. Finally, the third pulse transfers the spins back to the x-y-plane, where detection takes place. The spectrum will resemble a COSY spectrum, but the off-diagonal peaks now indicate through-space rather than through-bond interactions. 

When one resonance in an NMR spectrum is perturbed by saturation or inversion, the net intensities of other resonances in the spectrum may change. This phenomenon is called the nuclear Overhauser effect (NOE). The change in resonance intensities is caused by spins close in space to those directly affected by the perturbation. In an ideal NOE experiment, the target resonance is completely saturated by selected irradiation, while all other signals are completely unaffected. An NOE study of a rigid molecule or molecular residue often gives both structural and conformational information, whereas for highly flexible molecules or residues NOE studies are less useful. 

In de novo three-dimensional structure determinations of proteins in solution by NMR spectroscopy, the key conformational data are upper distance limits derived from nuclear Overhauser effects (NOEs) [11, 14]. In order to extract distance constraints from a NOESY spectrum, its cross peaks have to be assigned, i.e. the pairs of hydrogen atoms that give rise to cross peaks have to be identified. The basis for the NOESY assignment... 

FIGURE 10. Contour plot of two-dimensional nuclear Overhauser effect ll NMR (NOESY) of the protonated Schiff base of all-traos-retinal, in chloroform, with formate as the counterion. The intermolecular NOE cross-peak observed between H15 of the retinal and the counterion proton, at a mixing time of 0.4 s, is shown. Top trace f2 projection of the 2D NOE spectrum. Reproduced by permission of John Wiley Sons from Reference 36... 

The NOESY spectrum relies on the Nuclear Overhauser Effect and shows which pairs of nuclei in a molecule are close together in space. The NOESY spectrum is very similar in appearance to a COSY spectrum. It is a symmetrical spectmm that has the Iff NMR spectmm of the substance as both of the chemical shift axes (Fi and F2). A schematic representation of NOESY spectmm is given below. Again, it is usual to plot a normal (one-dimensional) NMR spectmm along each of the axes to give reference spectra for the peaks that appear in the two-dimensional spectmm. 

An important first step in interpreting the C-13 spectra is to distinguish a-carbons from 3-carbons, i.e. methine from methylene. Observation of multiplicity when the proton decoupler is off is one way, but this is not always easy if the lines are broadened by chemical shift multiplicity. Measurement of has been used for this purpose since the 3-carbon with two bonded protons relaxes about twice as fast as the a-carbon with only one. A very positive way is by deuterium labelling. In Fig. 3 is shown the main-chain 25 MHz carbon spectrum of two styrene-S02 copolymers containing 58 mol% styrene, or a ratio of styrene to SO2 of 1.38 (7 ). In the bottom one, 3,3-d2-styrene has been used, cind all the 3-carbon resonances are distinguishable from the a-carbon resonances since the presence of deuterium has eliminated their nuclear Overhauser effect because of this eind the deuterium J coupling ( 20 Hz), they are markedly smaller eind broader than the a-carbon resonances. 

The use of reference deconvolution for the correction of artefacts in nuclear Overhauser effect difference spectroscopy [9] is illustrated by the spectra of fig. 3. The experimental technique used here differs slightly from that normally encountered in using a control spectrum in which the preirradiation is gated off rather than shifted in frequency, and in keeping the decoupler and transmitter at the same frequency. These modifications were... 

The most conclusive evidence for the presence of duplexes la la and 2 2 came from 2-D H-NMR [nuclear Overhauser effect spectroscopy (NOESY)] and X-ray crystallography. The NOESY spectrum of lb in CDCI3 contains interstrand NOEs (Fig. 9.2a) between protons c and e, c and i, and c and j, which are consistent with an H bonded dimer. The crystal structures of la and 2 both revealed the expected dimeric stmctures held together by intermolecular H bonds (Fig. 9.2b). 

Steric interaction between the 1- and 9-positions in dibenzofurans has, however, been observed by NMR techniques. Thus saturation of the signal for the 1-methyl group in the H-NMR spectrum of 1,2,3,4-tetramethyl-dibenzofuran (11) produced a 38% nuclear Overhauser effect at the 9-H. This may be compared qualitatively with the 33% nuclear Overhauser effect produced on the 5-H in the spectrum of 1,2,3,4-tetramethylphenanthrene by... 

The nuclear Overhauser method has been used to demonstrate that in the dibenzofuranol 14 the position ortho to the hydroxy group is vacant. Saturation of the signal due to the benzylic protons of the benzyl group in the spectrum of the benzyl ether 15 gave a 39% nuclear Overhauser effect at the aromatic proton. 

Nuclear Overhauser effect (NOE) difference measurements were used to assign structure 79 for the product of reaction of diphenylnitrile imine with 5-ethylsulfonyl-2-methyl(27/)pyridazinone. Thus in the H NMR spectrum the ot/, o-protons of the arylhydrazino moiety (which were identified by two-dimensional heteronuclear multiple quantum correlation (2-D HMQC) spectroscopy) were shown in differential NOE (DNOE) experiment to be significantly enhanced on irradiation of pyridazine hydrogen H-7, proving their steric proximity <2000JST13>. 

H is particularly important in NMR experiments because of its high sensitivity and natural abundance. For macromolecules, 1H NMR spectra can become quite complicated. Even a small protein has hundreds of 1H atoms, typically resulting in a one-dimensional NMR spectrum too complex for analysis. Structural analysis of proteins became possible with the advent of two-dimensional NMR techniques (Fig. 3). These methods allow measurement of distance-dependent coupling of nuclear spins in nearby atoms through space (the nuclear Overhauser effect (NOE), in a method dubbed NOESY) or the coupling of nuclear spins in atoms connected by covalent bonds (total correlation spectroscopy, or TOCSY). 

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