Nuclear Overhauser effect distance constraints

Distance Constraints from Nuclear Overhauser Effects... 

The principle source of experimental conformational data in an NMR structure determination is constraints on short interatomic distances between hydrogen atoms obtained from NMR measurements of the nuclear Overhauser effect (NOE). NOEs result from cross-relaxation mediated by the dipole-dipole interaction between spatially proximate nu-... 

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... 

The basis for the determination of solution conformation from NMR data lies in the determination of cross relaxation rates between pairs of protons from cross peak intensities in two-dimensional nuclear Overhauser effect (NOE) experiments. In the event that pairs of protons may be assumed to be rigidly fixed in an isotopically tumbling sphere, a simple inverse sixth power relationship between interproton distances and cross relaxation rates permits the accurate determination of distances. Determination of a sufficient number of interproton distance constraints can lead to the unambiguous determination of solution conformation, as illustrated in the early work of Kuntz, et al. (25). While distance geometry algorithms remain the basis of much structural work done today (1-4), other approaches exist. For instance, those we intend to apply here represent NMR constraints as pseudoenergies for use in molecular dynamics or molecular mechanics programs (5-9). 

Many other intermolecular and intramolecular contacts are described by distances (hydrogen bond lengths, van der Waals contact, experimentally determined distances from nuclear Overhauser effect (NOE) spectra, fluorescence energy transfer, etc.) so that the distance matrix representation can be used to specify all the known information about a molecular structure. These bounds are entered into a distance geometry program, as are other bounds that specify constraints on modeling problems, such as constraints to superimpose atoms in different molecules. Hypotheses about intra- or intermolecular conformations and interactions are easily specified with distance constraints models can be built quickly to test different hypotheses simply by changing the distance constraints. 

The most important structural probe is the nuclear Overhauser effect (NOE), which provides valuable information on the structure of linear peptides. Briefly, the observation of a direct NOE between a pair of protons indicates the presence of a significant population of conformers in which the distance between these two proteins is relatively short. The overall pattern of connectivity therefore corresponds to a particular conformation. Constraints on both backbone and side chain dihedral angles are obtained from coupling contacts. An example of a two-dimensional NMR spectrum for a synthetic peptide related to residues 6-13 of human growth hormone is shown in Figure 8. 

The bound conformations of ligands determined by transferred-NOE (nuclear Overhauser effect) can also serve as templates for query formulation, even though this implicit approach cannot identify the spatial arrangement of site residue atoms. With these data, one proceeds with the analysis of the bound conformation as in the case of a co-determined complex (see above) however, the excluded volume constraints cannot be specified. One advantage of this data set is that multiple distance geometry solutions for the NOE-determined bound conformation can be used to define some initial tolerances for the distance, angle, and torsional 3D constraint features. 

Distance measurements by relaxation enhancement are much less popular in EPR as compared to NMR spectroscopy, where constraints based on the nuclear Overhauser effect or paramagnetic relaxation enhancement often play an important role in macromolecular structure determination. Such an underrepresentation of RE-based distance measurements in EPR is only partially connected to the fast and broad spread of the DEER technique. The RE approach has its internal difficulties that have to be overcome in order to secure broad and robust applicability of the method. We shall see in the following that while the RE approach is useful to derive qualitative conclusions, in its current state it has to be applied with care for measurement of precise distances. The perspectives for resolving this precision issue will be briefly discussed in the last section. 

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