Nuclear magnetic resonance structural refinement

Nuclear Magnetic Resonance Structure Calculation and Refinement 1531... 

The use of computer simulations to study internal motions and thermodynamic properties is receiving increased attention. One important use of the method is to provide a more fundamental understanding of the molecular information contained in various kinds of experiments on these complex systems. In the first part of this paper we review recent work in our laboratory concerned with the use of computer simulations for the interpretation of experimental probes of molecular structure and dynamics of proteins and nucleic acids. The interplay between computer simulations and three experimental techniques is emphasized (1) nuclear magnetic resonance relaxation spectroscopy, (2) refinement of macro-molecular x-ray structures, and (3) vibrational spectroscopy. The treatment of solvent effects in biopolymer simulations is a difficult problem. It is not possible to study systematically the effect of solvent conditions, e.g. added salt concentration, on biopolymer properties by means of simulations alone. In the last part of the paper we review a more analytical approach we have developed to study polyelectrolyte properties of solvated biopolymers. The results are compared with computer simulations. 

A review is given of the application of Molecular Dynamics (MD) computer simulation to complex molecular systems. Three topics are treated in particular the computation of free energy from simulations, applied to the prediction of the binding constant of an inhibitor to the enzyme dihydrofolate reductase the use of MD simulations in structural refinements based on two-dimensional high-resolution nuclear magnetic resonance data, applied to the lac repressor headpiece the simulation of a hydrated lipid bilayer in atomic detail. The latter shows a rather diffuse structure of the hydrophilic head group layer with considerable local compensation of charge density. 

The refinement of other analytical methods, such as electrophoresis [34,36], the various techniques of optical spectroscopy [103-105], and nuclear magnetic resonance [201], is supplemented by the recent advances in real-time affinity measurements [152,202], contributing to the understanding of biomolecular reactivity. Taken together, the improvement of analytical methods will eventually allow a comprehensive characterization of the structure, topology, and properties of the nucleic acid-based supramolecular components under consideration for distinctive applications in nanobiotechnology. 

In the case of heterogeneous polymers the experimental methods need to be refined. In order to analyze those polymers it is necessary to determine a set of functions / (M), which describe the distribution for each kind of heterogeneity i This could be the mass distributions of the blocks in a diblock copolymer. The standard SEC methods fail here and one needs to refine the method, e.g., by performing liquid chromatography at the critical point of adsorption [59] or combine SEC with methods, which are, for instance, sensitive to the chemical structure, e.g., high-pressure liquid chromatography (HPLC), infrared (IR), or nuclear magnetic resonance spectroscopy (NMR) [57],... 

The refinement or creation of new approaches may result in the elimination of existing activities. For example, the structure confirmation of newly synthesized lead compounds traditionally involved an extensive use of nuclear magnetic resonance (NMR). Once reliable LC/MS methodologies became available and their performance was benchmarked, they were soon accepted as an exclusive method for the rapid structure confirmation of lead compounds at an earlier stage of the lead identification process. 

R. B. Altman and O. Jardetzky, in Methods in Enzymology, Vol. 177, N. J. Oppenheimer and T. L. James, Eds., Academic Press, San Diego, Calif., 1989, pp. 218-246. Heuristic Refinement Method for Determination of Solution Structure of Proteins from Nuclear Magnetic Resonance Data. 

Jourdan, M., Garda, J., Defrancq, E., Kotera, M., and Lhomme, J. (1999) 2 -Deoxyribonolactone lesion in DNA refined solution structure determined hy nuclear magnetic resonance and molecular modeling. Biochemistry, 38, 3985-3995. 

The key developments of the subsequent decade were in the realm of bonded-phase technology. The efforts of many researchers gave us a superior understanding of the retention mechanisms, and the structure and preparation of bonded phases, including the pretreatment of the silica surface itself. Important tools were refined chromatographic characterization techniques and the use of sophisticated instruments such as solid-state nuclear magnetic resonance (NMR). /... 

Molecular dynamics attempts to solve the dynamically evolving ensemble of molecules given the interactions between molecules. The form of the forces between molecules or atoms, the number of interactions (i.e., two- or three-body interactions), and the number of molecules that can be tackled by the program determine the success of the model. Molecular dynamics simulations can predict the internal energy, heat capacity, viscosity, and infrared spectrum of the studied compound and form an integral part in the determination and refinement of structures from X-ray crystallography or nuclear magnetic resonance (NMR) experiments. 

Additionally, in part because of the pervasive intrusion (or timely arrival) of microprocessors, tools for structure determination are constantly being refined and enlarged in scope. Examples include multidimensional nuclear magnetic resonance (NMR), which is now commonly used for large molecules (the first commercial NMR spectrometers were introduced in the late 1950s) and, even more recently, near-field microscopy, which uses a lensless technique for VIS spectroscopy and thus sidesteps the normal problems of resolution by accumulating structural features a little at a time, is being developed (the first compound microscope became available in 1610). ... 

Besides the classical techniques for structural determination of proteins, namely X-ray diffraction or nuclear magnetic resonance, molecular modelling has become a complementary approach, providing refined structural details [4—7]. This view on the atomic scale paves the way to a comprehensive smdy of the correlations between protein structure and function, but a realistic description relies strongly on the performance of the theoretical tools. Nowadays, a full size protein is treated by force fields models [7-10], and smaller motifs, such as an active site of an enzyme, by multiscale approaches involving both quantum chemistry methods for local description, and molecular mechanics for its environment [11]. However, none of these methods are ab initio force fields require a parameterisation based on experimental data of model systems DPT quantum methods need to be assessed by comparison against high level ab initio calculations on small systems. 

For the former, i.e., diffraction, structural refinement techniques that wholly address both the crystalline and diffuse portions of the scattering profiles would be a tremendous advance. The latter component will entail heavier reliance on a variety of powerful probes of local structure including nuclear magnetic resonance, scanning tunneling microscopy, anomalous scattering methods, and radial distribution function analysis [21 j. As the number of chemical/structural architectures available continues to increase, determining the fundamental nature of the structure-property relationships will become an ever more important issue. 

---