Molecular conformations of DNA and RNA subunits

Issued as Report of expenditures, Project no. G-41-64

Influence of ribose 2′-O-methylation on GpC conformation by classical potential energy calculations

Potential energy calculations were employed to examine the effect of ribose 2′-O-methylation on the conformation of GpC. Minimum energy conformations and allowed conformational regions were calculated for 2′MeGpC and Gp2′MeC. The two lowest energy conformations of 2′MeGpC and Gp2′MeC are similar to those of GpC itself. The helical RNA conformation (sugar pucker-C(3′)-endo, ω′ and ω,g−g−, bases-anti) is the global minimum, and a helix-reversing conformation with ω′, ω in the vicinity of 20°, 80° is next in energy. However, subtle differences between the three molecules are noted. When the substitution is on the 5′ ribose (Gp2′MeC), the energy of the helical conformation is less than that of GpC, due to favorable interactions of the added methyl group. When the substitution is at the 3′ ribose (2′MeGpC) these stabilizing interactions are outweighed by steric restrictions, and the helical conformation is of higher energy than for GpC. Furthermore, the statistical weight of the 2′MeGpC g− g− helical region is substantially less than the corresponding weight for Gp2′MeC. In addition, 2′MeGpC′s methoxy group is conformationally restricted to a narrow range centered at 76°. This group has a broadly allowed region between 50 and 175° in Gp2′MeC. These differences occur because the appended methyl group in 2′MeGpC is located in the interior of the helix cylinder, as it would be in polynucleotide, while it hangs unimpeded in Gp2′MeC. These findings suggest that 2′-O-methylation has both stabilizing and destabilizing influences on the helical conformation of RNA. For 2′MeGpC the destabilizing steric hindrance imposed by the nature of the guanine base dominates

Conformational stability in dinucleoside phosphate crystals. Semiempirical potential energy calculations for uridylyl-3'-5'-adenosine monophosphate (UpA) and guanylyl-3',5'-cytidine monophosphate (GpC)

Classical potential energy calculations were performed for the dinucleoside phosphates UpA and GpC. Two widely accessible low-energy regions of conformation space were found for the w', w pair. That of lowest energy contains conformations similar to helical RNA, with w' and w in the vicinity of 300° and 280°, respectively. All five experimental observations of crystalline GpC, two of ApU, and the helical fragment of ApApA fall in this range. The second lowest region has w' and w at about 20° and 80°, respectively, which is in the general region of one experimentally observed crystalline conformer of UpA, and the nonhelical region of ApApA. It is concluded that GpC and ApU, which were crystallized as either sodium or calcium salts, are shielded from each other in the crystal by the water of hydration and are therefore free to adopt their predicted in vacuo minimum energy helical conformations. By contrast, crystalline UpA had only 1/2 water per molecule, and was forced into higher energy conformations in order to maximize intermolecular hydrogen bonding

The crystal and molecular structure of a calcium salt of guanylyl-3',5'-cytidine (GpC)

The calcium salt, Ca(C_19H_24N_8O_I2P)_2.18H_20, of guanylyl-3',5'-cytidine (GpC) has been refined to an R of 8·2 % for 2918 observed reflections (11% for 4237 reflections, including unobserved). The molecule crystallized in space group P2_1 with a=21·224, b=34·207, c=9·327 Å, β=90·527°, Z=4. The asymmetric unit contains four GpC, 36 waters and two Ca^2+ ions, for a total of 198 non-hydrogen atoms. The four GpC occur as two dimers related by a pseudo C-face-centering. Each dimer consists of two crystallographically independent GpC as Watson-Crick base-pairs, and possesses a pseudo twofold axis broken by a Ca^2+ ion and associated solvent. The structure was solved by an unusual series of steps including semi-empirical potential-energy methods, packing analysis, rigid-body refinement, least-squares and difference Fourier techniques, and direct-methods tangent-formula phase refinement. The four GpC have conformational angles in the range of helical RNA, but are not identical. The different crystallographic environments perturb the GpC from exact symmetry and demonstrate the range of the basic helical conformations. All eight bases are anti, sugars are all C(3’) endo, the C(4')-C(5') bond rotations are gauche-gauche, and the ω', ω angle pair about the O-P bonds is gauche—gauche-

Structure of guanylyl-3′,5′-cytidine monophosphate. II. Description of the molecular and crystal structure of the calcium derivative in space group P2₁

The structural features of calcium guanosine-3′,5′-cytidine monophosphate (GpC) have been elucidated by X-ray diffraction analysis. The molecule was crystallized in space group P2₁ with cell constants of a = 21.224 Å, b = 34.207 Å, c = 9.327 Å, and β = 90.527°, Z = 8. The hydration of the crystal is 21% by weight with 72 water molecules in the unit cell. The four GpC molecules in the asymmetric unit occur as two Watson-Crick hydrogen-bonded dimers related by a pseudo-C face centering. Each dimer consists of two independent GpC molecules whose bases are hydrogen bonded to each other in the traditional Watson-Crick fashion. Each dimer possesses a pseudo twofold axis broken by a calcium ion and associated solvent. The four molecules are conformationally similar to helical RNA, but are not identical to it or to each other. Instead, values of conformational angles reflect the intrinsic flexibility of the molecule within the range of basic helical conformations. All eight bases are anti, sugars are all C3′-endo, and the C4′-C5′ bond rotations are gauche-gauche. The R factor is 12.6% for 2918 observed reflections at 1.2-Å resolution

Classical potential energy calculations for ApA, CpC, GpG, and UpU. The influence of the bases on RNA subunit conformations

Classical potential energy calculations have been made for the ribodinucleoside monophosphates ApA, CpC, GpG, and UpU. Van der Waal's, electrostatic, and torsional contributions to the energy were calculated, and the energy was minimized with the seven backbone conformational angles as simultaneously variable parameters. At the global minimum, ApA and CpC have conformations like double helical RNA: the angles ω′ and ω are g−g−, the sugar pucker is C3′-endo, and the bases are anti. GpG and UpU, on the other hand, have the ω′,ω angle pair g−t at the global minimum, and for GpG the bases are syn. Energy contour maps for ω′ and ω show two broad, low energy regions for ApA, CpC, and UpU: one is g−g−, and the second encompasses g−t and g+g+ within a single low energy contour. The two regions are connected by a path at 10–13 kcal./mole. For GpG, with bases syn, however, only a small low-energy region at g−t is found. The helical ‘A’ RNA conformation is 8.5 kcal/mole higher for this molecule. Thus, the base composition is shown to influence the conformations adopted by dinucleoside phosphates. Comparison of calculations with experimental data, where available, show good agreement

DNA Backbone Conformation in Cis-Syn Pyrimidine[ ]Pyrimidine Cyclobutane Dimers

A theoretical calculation involving minimization of energy functions was made to determine the conformation of a deoxydinucleoside monophosphate pyrimidine[ ]pyrimidine cis-syn cyclobutane dimer. Such single-stranded moieties are useful models for the in uiuo situation because single strands are more easily damaged, and the damage inhibits the normal function of single-stranded replicating DNA. The calculated minimum-energy conformer was incorporated in a mode l of B-DNA to assess the influence of this covalent lesion on the normal form of DNA

Solution conformation of the major adduct between the carcinogen (+)-anti-benzo[ɑ]pyrene diol epoxide and DNA

We have synthesized, separated, and purified ≈10 mg of a deoxyundecanucleotide duplex containing a single centrally positioned covalent adduct between (+)-anti-benzo[a]pyrene (BP) diol epoxide and the exocyclic amino group of guanosine. Excellent proton NMR spectra are observed for the (+)-trans-anti-BP diol epoxide-N^2-dG adduct positioned opposite dC and flanked by G.C pairs in the d[C1-C2-A3-T4-C5-(BP)G6-C7-T8-A9-C10-C11].d[12- G13-T14-A15-G16-C17-G18-A19-T20-G 21-G22] duplex +ADdesignated (BP)G.C 11-mer+BD. We have determined the solution structure centered about the BP covalent adduct site in the (BP)G.C 11-mer duplex by incorporating intramolecular and intermolecular proton-proton distance bounds deduced from the NMR data sets as constraints in energy minimization computations. The BP ring is positioned in the minor groove and directed toward the 5' end of the modified strand. One face of the BP ring of (BP)G6 is stacked over the G18 and A19 sugar-phosphate backbone on the partner strand and the other face is exposed to solvent. A minimally perturbed B-DNA helix is observed for the d[T4-C5-(BP)G6-C7-T8].d[A15-G16-C17-G18-A19] segment centered about the adduct site with Watson-Crick alignment for both the (BP)G6.C17 pair and flanking G.C pairs. A widening of the minor groove at the adduct site is detected that accommodates the BP ring whose long axis makes an angle of ≈45° with the average direction of the DNA helix axis. Our study holds future promise for the characterization of other steroisomerically pure adducts of BP diol epoxides with DNA to elucidate the molecular basis of structure-activity relationships associated with the stereoisomer-dependent spectrum of mutational and carcinogenic activities

Conformation of a rare nucleoside in the anti-codon loop of tRNAs: Potential energy calculations for 2′-O-methyl cytidine

The classical potential energy of 2′-O-methyl cytidine was calculated using contributions from van der Waals', electrostatic, and torsional terms. All five conformational angles, namely χ, Ψ, the methoxy angles m₁ and m₂ which are unique to O′-methylated nucleosides, and the sugar pucker P were varied simultaneously, and the energy was minimized with respect to these parameters. An extensive search of conformation space was made, particularly with respect to the sugar pucker. At the predicted global minimum, χ and Ψ are anti-gg, P is C(2′)-endo-C(3′)-exo, and the methyl group is staggered with respect to the sugar. This calculated minimum agrees very well with the recently determined crystal structure of 2′-O-methyl cytidine. Thus, 2′-O-methylation still permits the conformational regions of the common nucleosides to be adopted. However, we infer that the predicted C(2′)-endo sugar pucker results from the added methyl group, since cytidine is C(3′)-endo in the crystal, and the common ribopyrimidine nucleosides generally favor C(3′)-endo