Determination of solution conformation

Ragg, E., Tagliavini, F., Malesani, P., Monticelli, L., Bugiani, O., Forloni, G., and Salmona, M. (1999). Determination of solution conformations of PrP106-126, a neurotoxic fragment of prion protein, by 1H NMR and restrained molecular dynamics. Eur.J. Biochem. 266, 1192-1201. 

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). 

There is a variety of methods for the determination of the structure of oligosaccharides. Different techniques are available for the determination of solution conformations and crystal structures respectively. The following section will deal with the most frequently used techniques. 

The different VCD spectra calculated for the NMR and X-ray structures demonstrate just how sensitive VCD is toward structural changes. It is premature, at this stage, to attempt to use VCD as a tool to distinguish or even judge the quality of either of the previously obtained structures. However, further computational efforts may well establish VCD as a complementary tool to NMR for the determination of solution conformation. VCD will certainly not challenge NMR in large molecules, where the superior resolution allows detailed structures to be derived. However, the much faster time frame of vibrational spectroscopy over nuclear magnetic resonance techniques allows distinct structures to be discerned when NMR measurements perceive little structure. 

We note that the calculation of At/ will depend primarily on local information about solute-solvent interactions i.c., the magnitude of A U is of molecular order. An accurate determination of this partition function is therefore possible based on the molecular details of the solution in the vicinity of the solute. The success of the test-particle method can be attributed to this property. A second feature of these relations, apparent in Eq. (4), is the evaluation of solute conformational stability in solution by separately calculating the equilibrium distribution of solute conformations for an isolated molecule and the solvent response to this distribution. This evaluation will likewise depend on primarily local interactions between the solute and solvent. For macromolecular solutes, simple physical approximations involving only partially hydrated solutes might be sufficient. 

A general account of the NMR spectroscopy of six-membered ring heterocycles is given in Chapter 2.01. Proton and carbon NMR spectroscopy have been used extensively to determine the solution conformations of saturated heterocyclic rings containing three or... 

Techniques and methods for the study of protein adsorption have been well reviewed 4). It is now generally recognized that it is not necessarily the type and amount of protein present at the surface which is most important, but rather the orientation and conformational state of those proteins. At present it is virtually impossible to predict the specific conformation of an adsorbed protein at a particular interface. The techniques used in the determination of protein conformation in solution or in the solid state do not usually apply to adsorbed proteins. Hence, the difference between adsorbed and bulk solution protein conformation has to be inferred indirectly. 

A combination of the two techniques was shown to be a useful method for the determination of solution structures of weakly coupled dicopper(II) complexes (Fig. 9.4)[119]. The MM-EPR approach involves a conformational analysis of the dimeric structure, the simulation of the EPR spectrum with the geometric parameters resulting from the calculated structures and spin hamiltonian parameters derived from similar complexes, and the refinement of the structure by successive molecular mechanics calculation and EPR simulation cycles. This method was successfully tested with two dinuclear complexes with known X-ray structures and applied to the determination of a copper(II) dimer with unknown structure (Fig. 9.5 and Table 9.9)[119]. 

NMR spectroscopy has developed during the last few years into a very powerful method to establish both the conformation of an oligosaccharide in solution and in the crystalline state. The solution conformation of a saccharide can be determined generally by the combination of 13C-NMR and H-NMR techniques. 2-d methods became available in the last few years and are prerequisites for the elucidation of the three dimensional structure of an oligosaccharide [7]. Both NOE measurements [21] as well as traditional determination of coupling constants and chemical shifts are important tools for the determination of preferred conformations. 2-dimensional methods have greatly improved the accessibility of these parameters from complex molecules. 

Several applications of these couplings to the conformational analysis of various mono- and oligosaccharides have also been described.138 140 143 149 150 These applications incorporate both theoretical and experimental approaches toward determination of the conformation of saccharides in solution, a question still controversial because of the time-averaged character of the NMR data obtained (NOE and couplings) and the occurrence of conformational equilibria.3 Variations of Vc H values for maltose according to the solvent used have been observed,46 and... 

However, one of the interesting aspects of NMR structural results is that they often suggest no discernible solution structure for small systems, such as a tri- or tetra-peptides. This is due to the flexibility and structural variance displayed by these sample systems. However, the lack of interactions between parts of the molecule, which normally are detected via Nuclear Overhauser enhancements and refined into molecular structures during NMR structural determinations, should not be interpreted as a lack of a solution structure. It is the slow time scale of NMR, coupled with the rapidly interconverting conformations, which weakens these effects to the point where they can no longer be detected with certainty, and structural techniques which operate on a much faster time scale (e.g., UV-CD spectroscopy, or forms of vibrational spectroscopy) demonstrate that there is a preferred class of solution conformers even in small peptide systems. 

T. F. Havel and K. Wiithrich, J. Mol Biol., 182, 281 (1985). An Evaluation of the Combined Use of Nuclear Magnetic Resonance and Distance Geometry for the Determination of Protein Conformations in Solution. 

A special field of application of these techniques is the determination of molecular conformations in cases where X-ray and neutron techniques are not applicable, in solution or in the liquid phase. It is known that conformers afford essentially the same spectra the intensities of some bands may vary. It is sufficient to investigate some bands in order to determine the conformation, and quantum chemical methods can be applied (Oelichmann et al., 1981a, b, 1982 Grunenberg and Bougeard, 1987). 

Coupling constants and NOEs are the main NMR parameters used in determining the solution conformations of drug leads. NOEs provide information about through-space proxim-... 

A new method for the quantitative determination of the conformational equilibrium of bisquinolizidine alkaloids in solution, by H and NMR, was developed by Wysocka and Brukwicki [199]. Using sparteine and 5,6-dehydromultiflorine. Fig. (30), as model compounds for a C ring in boat and chair conformation, respectively, the authors showed that the percentage of conformers with the C ring in boat conformation can be determined on the basis of the experimental chemical shifts of C-12 and C-14 and of the J coupling value of H-7/H-17P, by the formula Fb = (5-6c) /(6b-6c), where 5 is the experimentel value, and 5b and 5c represent the 5 or J coupling values of the model compoimds in boat and chair conformation, respectively. 

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