In order to obtain information about the conformational features in a polyribonucleotide at the nearest neighbor level, detailed nuclear magnetic resonance studies of the dinucleoside monophosphates ApA, ApG, GpA, UpU, CpC, UpC, and CpU were undertaken. Proton spectra were recorded at 100, 220, 270, or 300 MHz for D2O solutions, 0.01-0.03 M, pD 7.4 at 20+/-2 degrees C. Spectra of ApA, ApG, UpU, and UpC were also recorded in the temperature range of 70-90 degrees C. Unambiguous signal assignments of all proton resonances were made with the aid of selectively deuterated dimers. Complete, accurate sets of nuclear magnetic resonance (NMR) parameters were derived for each nucleotidyl unit by simulation-iteration methods. A complete set of chemical shift and coupling constant data was also obtained for all the constituent monomeric units at a concentration and ionization state comparable to that of the dimers. Conformational properties were evaluated quantitatively for most of the bonds in the dinucleoside monophosphates using procedures developed in earlier studies. All of the dimers have a flexible conformational framework in aqueous solution. While flexibility is allowed and alternate conformations are accessible, these molecules nevertheless attempt to achieve conformational identity by showing preferences--sometimes overwhelming preferences--for certain orientations. Thus the ribose rings exist as equilibrium mixtures of C2'-endo in equilibrium C3'-endo conformers with a bias for the C3'-endo pucker in most cases. The C4'-C5' bonds of both nucleotidyl units show significant preference (70-85%) for a gg conformation. Similarly, the dominant conformer (80-90%) about C5'-O5' is g'g'. Even though an unambiguous determination of the orientation about C3'-O3' cannot be made, there is suggestive evidence that the orientation of the 3' phosphate group is coupled to the ribose conformational equilibrium and it is likely that a 3Eg- in equilibrium 2Eg+ equilibrium exists with a bias for the 3Eg- coupled conformation in which the H3'-C3'-O3'-P dihedral angle is about 34-38 degrees. The individual nucleotidyl units in the dimers differ in several key ways from corresponding monomer conformations. Specifically, the ribose equilibrium C2'-endo in equilibrium C3'-endo shifts in favor of C3'-endo upon dimerization, the only exception being UpU. The C4'-C5' and C5'-O5' bonding network in the dimer forms a stable conformational unit and no correlation exists in the dimers between the conformational preference of this fragment and ribose conformer population. The temperature data for the dimers and dimerization data clearly indicate that the transition C2'-endo leads to C3'-endo is directly related to XCN changes brought about by dimerization and stacking...
The detailed conformational features and dynamics of heterodinucleoside monophosphates ApU, ApC, GpU, GpC, UpA, CpA, UpG, and CpG have been studied in aqueous solution by high field nuclear magnetic resonance (NMR) spectroscopy. Analysis of the resultant NMR parameters leads to a number of discernible trends throughout the series. Thus the ribose rings of the dimers exist as equilibrium mixtures of C(2')-endo(2E) in equilibrium C(3')-endo(3E) conformers with a proclivity for the 3E pucker in most cases; the C(4')-C(5') bonds of both nucleotidyl units show significant preference (74-96%) for a gg conformation and the dominant conformer (85-89%) about C(5')-O(5') is g'g'. Orientation about the C(3')-O(3') bond is coupled to the ribose conformational equilibrium and the system exists with a bias for the 3Eg- coupled conformation in which the H(3')-C(3')-O(3')-P dihedral angle occupies the narrow range of 33-35 degrees. Dimerization, on the average, causes about 10% increase in gg and g'g' populations and the g-domain becomes increasingly populated about the C(3')-O(3') bond. The ribose equilibrium 2E in equilibrium 3E shifts in favor of 3E upon dimerization, the effect being very conspicuous for the pu-py series (similar to 40 yields 60%) and less noticeable for the py-pu systems (similar to 47 yields 58%), clearly suggesting a correlation between sequence and ribose conformational equilibrium. The temperature and dimerization data for the heterodinucleoside monophosphates show that the transition 2E yields 3E is directly related to XCN changes induced by dimerization and stacking. Analysis of the ribose coupling data shows that the percentage populations of stacked species vary from dimer to dimer with GpC displaying a maximum of 45% stacked population and UpG about 10%. However, in general, the pu-py dimers show a higher preference (27-45%) for stacked conformations than py-pu dimers (10-25%). It is proposed that the pronounced deshielding of H(5') of the 5'-nucleotidyl units upon dimerization is associated with the presence of right-handed stacks (g-g-), whereas the chemical shift trends of H(5') and H(5') of 3'-nucleotidyl units are due to the presence of left-handed stacks (g+g+) in all the dimers. In pu-py dimers, the population of the g-g- species is found to be greater than that of g+g+. Also the population of g-g- stacks in pu-py dimers is generally greater than in their corresponding matched py-pu dimers. Thus the base sequence has not only an explicit effect on the overall populations of stacked species, but also on the handedness of the stacks. The present results further confirm the interdependence of conformational bonds throughout the nucleotidyl framework.
The importance of O-acetyl groups to the immunogenicity of Neisseria meningitidis serogroup A polysaccharide (PS) was examined in studies using human sera and mouse immunization.
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