Relaxation Studies of Nucleic Acids

For the above reasons, suitable methods are only starting to emerge. In the following paragraphs we will discuss the application of the relaxation measurements to dynamics studies and structure determination separately. 

Unlike imino nitrogen, protonated carbon nuclei are abundant throughout the molecules of nucleic acids providing many sites as possible probes into the molecular motions. How- 

Oy and Oz are the angles between the corresponding tensor component and the C-H bond. 

The nature of the chemical shift tensor is a potential source of complications in relaxation studies. For sugar carbons, the CSAs are around 40 ppm and their contribution to relaxation of protonated carbons is nearly negligible. On the other hand, CSA values of the protonated carbons of the bases are between 120 and 180 ppm, the tensors deviate quite significantly from axial symmetry and none of their principal components is colli-near with the C-H bond. This makes interpretation of the relaxation rates in terms of molecular dynamics prohibitively complicated or, if neglected, introduces an error whose magnitude has not yet been evaluated. 

In nucleic acids, the cross-correlation studies were applied to investigation of the sugar conformation [101-103] of the phosphodiester backbone [65] as well as to some more spe- 

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The study of nucleic acid bases by NMR has been reported in a number of monographs (/), but very little data is available on the, 3C and, 5N NMR chemical shift tensors in these compounds. The low sensitivity of NMR spectroscopy and the long relaxation times exhibited by many of these compounds have posed the main impediments for these studies. The use of sample doping with free radical relaxation reagents, to reduce the relaxation times facilitating 2D multiple pulse experiment (2, 3), enables one to measure and analyze the principal values of the chemical shift tensors in natural abundance samples. In previous papers from this laboratory we have presented, 5N NMR chemical shift principal values for adenine, guanine, cytosine, thymine and uracil (4, 5). 

J. Fagon, J. Boisbouvier, P. Schanda, A. Pardi, J.P. Simotre, B. Bmtscher, Longitudinal-relaxation-enhanced NMR experiments for the study of nucleic acids in solution, J. Am. Chem. Soc. 131 (2009) 8571—8577. 

In principle, paramagnetic ions also may be used to induce hyperfine shifts in nucleic acids to aid detection of binding sites. Ions with high relative magnetic anisotropy and short unpaired electron relaxation times (i.e., Co Fe and trivalent lanthanide ions except for Gd ) are candidates for such studies. Indeed, Tb and Eu ions have been used as fluorescent probes of nucleic acid structures it is expected that NMR studies also would be informative. " ... 

Chi-Wan Chen and Jack S. Cohen introduce the application of - P NMR to DNA and RNA conformations. High-resolution P NMR of transfer ribonucleic acids and P studies on drug-nucleic acid complexes (prepared by the editor with Evelyn M. Goldfield) are then presented. Phillip A. Hart introduces phosphorus relaxation methods, which are shown to provide important information on the conformation and dynamics of nucleic acids and phosphoproteins. Thomas L. James delves further into relaxation behavior of solution-state nucleic acids, and Heisaburo Shindo completes this section by describing solid-state " P NMR of nucleic acids. 

Although other studies have examined certain aspects of nucleic acid structure predominantly using NMR chemical shifts, the following discussion is only concerned with dynamics information in nucleic acids that has been derived from NMR relaxation experiments. 

Quinacrine concentrates in the scolex of the parasite and causes the muscles needed for holding onto the intestinal wall to relax. The worms are stained yellow and pass from the body, still aUve. Quinacrine can intercalate with DNA and inhibit nucleic acid synthesis. It creates fluorescent bands in deoxyadenylate—deoxythmidylate-rich regions of DNA and has been used as a stain in the study of human genetics. 

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. 

New techniques for data analysis and improvements in instrumentation have now made it possible to carry out stmctural and conformational studies of biopolymers including proteins, polysaccharides, and nucleic acids. NMR, which may be done on noncrystalline materials in solution, provides a technique complementary to X-ray diffraction, which requires crystals for analysis. One-dimensional NMR, as described to this point, can offer structural data for smaller molecules. But proteins and other biopolymers with large numbers of protons will yield a very crowded spectrum with many overlapping lines. In multidimensional NMR (2-D, 3-D, 4-D), peaks are spread out through two or more axes to improve resolution. The techniques of correlation spectroscopy (COSY), nuclear Overhausser effect spectroscopy (NOESY), and transverse relaxation-optimized spectroscopy (TROSY) depend on the observation that nonequivalent protons interact with each other. By using multiple-pulse techniques, it is possible to perturb one nucleus and observe the effect on the spin states of other nuclei. The availability of powerful computers and Fourier transform (FT) calculations makes it possible to elucidate structures of proteins up to 40,000 daltons in molecular mass and there is future promise for studies on proteins over 100,000... 

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