The crystal and molecular structure of 8-methyladenosine 3'-monophosphate dihydrate.

8-Methyladenosine 3'-monophosphate dihydrate was synthesized and crystallized in the monoclinic space group P21 with the unit cell dimensions: a = 9.095(2) A, b = 16.750(3) A, c = 5.405(2) A and beta = 97.61(3) degrees. The structure was determined by the application of the heavy atom method and refined to give a final R factor of 0.047. The pertinent conformations are as follows: the syn conformation about the glycosyl bond (chiCN = 216.8 degrees), the C(2')-endo sugar puckering with the displacement of 0.55 A; and the gauche-gauche conformation about the C(4')-C(5') bond capable of forming an intramolecular hydrogen bonding between N(3) of adenine base and O(5') of the hydroxymethylene group on the ribose. The molecule exists in the zwitterionic form with the N(1) of the adenine base protonated by a phosphate proton and is stabilized by three-dimensional networks of hydrogen bonding through the crystalline water molecules or directly between the adjacent nucleotide molecules; no base stacking was observed.     

Crystal structure and conformation of 8-(2-hydroxyethylamino) and 8-(pyrrolidin-1-yl) adenosines

In the course of investigation of 8-alkylamino substituted adenosines, the title compounds were synthesized as potential partial agonists for adenosine receptors. The structure determination of these compounds was carried out with the X-ray crystallography study. Crystals of 8-(2-hydroxyethylamino)adenosine are monoclinic, space group P 2(1); a = 7.0422(2), b = 11.2635(3), c = 8.9215(2) A, beta = 92.261(1) degrees, V = 707.10(3) A3, Z = 2; R-factor is 0.0339. The nucleoside is characterized by the anti conformation; the ribose ring has the C(2')-endo conformation and gauche-gauche form across C(4')-C(5') bond. The molecular structure is stabilized by intramolecular hydrogen bond of N-HO type. Crystals of 8-(pyrrolidin-1-yl)adenosine are monoclinic, space group C 2; a = 19.271(1), b = 7.3572(4), c = 11.0465(7) A, beta = 103.254(2), V = 1524.4(2) degrees A3, Z = 4; R-factor is 0.0498. In this compound, there is syn conformation of the nucleoside; the ribose has the C(2')-endo conformation and gauche -gauche form across C(4')- C(5') bond. The molecular structure is stabilized by intramolecular hydrogen bond of O-HN type. For both compounds, the branching net of intermolecular hydrogen bonds occur in the crystal structures.     

The crystal and molecular structure of 2'-deoxy-2'-fluoroinosine monohydrate.

The structure of the hydrate of 2'-deoxy-2'-fluoroinosine has been determined by single-crystal x-ray diffraction. The nucleoside crystallizes in space group P2(1)2(1)2(1) with unit cell dimensions, a = 33.291, b = 10. 871, c = 6.897A. There are two nucleosides and two water molecules in the asymmetric unit. The structure was solved by direct methods and refined to a residual R = 0.095. The two independent nucleosides in the asymmetric unit show different conformations about the glycosidic bond, while other structural details are similar. The base orientation to the sugar is syn in molecule A, whereas anti in molecule B. The exocyclic C(4')-C(5') bond conformation defined with respect to C(3')-C(4')-C(5')-O(5') is gauche+ in both molecules A and B. The sugar ring pucker defined by the pseudorotation phase angle P is a twisted conformation in both, C(3')-endo-C(4')-exo with P = 29 degrees in molecule A and C(4')-exo-C(3')-endo with P = 41 degrees in molecule B. It is shown by comparison with x-ray results of other 2'-fluoronucleosides and unmodified nucleosides including inosines that, in addition to a strong preference of the C(3')-endo type pucker, twisted conformations involving C(4')-exo puckering may be one of characteristic features of 2'-fluoronucleosides.     

The crystal structure of the octamer [r(guauaca)dC]2 with six Watson-Crick base-pairs and two 3' overhang residues

The crystal structure of an alternating RNA octamer, r(guauaca)dC (RNA bases are in lower case while the only DNA base is in upper case), with two 3' overhang residues one of them a terminal deoxycytosine and the other a ribose adenine, has been determined at 2.2 A resolution. The refined structure has an Rwork 18.6% and Rfree 26.8%. There are two independent duplexes (molecules I and II) in the asymmetric unit cell, a = 24.95, b = 45.25 and c = 73.67 A, with space group P2(1)2(1)2(1). Instead of forming a blunt end duplex with two a+.c mispairs and six Watson-Crick base-pairs, the strands in the duplex slide towards the 3' direction forming a two-base overhang (radC) and a six Watson-Crick base-paired duplex. The duplexes are bent (molecule I, 20 degrees; molecule II, 25 degrees) and stack head-to-head to form a right-handed superhelix. The overhang residues are looped out and the penultimate adenines of the two residues at the top end (A15) are anti and at the bottom (A7) end are syn. The syn adenine bases form minor groove A*(G.C) base triples with C8-H...N2 hydrogen bonds. The anti adenine in molecule II also forms a triple and a different C2-H...N3 hydrogen bond, while the other anti adenine in molecule I does not, it stacks on the looped out overhang base dC. The 3' terminal deoxycytosines form two stacked hemiprotonated trans d(C.C)+ base-pairs and the pseudo dyad related molecules form four consecutive deoxyribose and ribose zipper hydrogen bonds in the minor groove.     

Structural studies of O6-methyldeoxyguanosine and related compounds: a promutagenic DNA lesion by methylating carcinogens.

O6-Methylation of guanine residues in DNA can induce mutations by formation of base mispairing due to the deprotonation of N(1). The electronic, geometric and conformational properties of three N(9)-Substituted O6-methylguanine derivatives, O6-methyldeoxyguanosine (O6mdGuo), O6-methylguanosine (O6mGuo) and O6, 9-dimethylguanine (O6mdGua), were investigated by X-ray and/or NMR studies. O6mdGuo crystallizes in the monoclinic space group P2(1) with cell parameters a = 5.267(1), b = 19.109(2), c = 12.330(2) A, beta = 92.45(1) degrees, V = 1239.8(3) A3, z = 4 (two nucleosides per asymmetric unit), and O6mGua in the monoclinic space group P2(1)/n with cell parameters a = 10.729(2), b = 7.640(1) c = 10.216(1) A, beta = 92.17(2) degrees, V = 836.7(2) A3, z = 4. The geometry and conformation of O6-methylguanine moieties observed in both crystals and are very similar. Furthermore, the molecular dimensions of the O6methylguanine residue resemble more closely those of adenine than those of guanine. The methoxy group is coplanar with the purine ring, the methyl group being cis to N(1). The conformation of O6-methylguanine nucleosides is variable. The glycosidic conformation of O6mdGuo is anti for molecule (a) and high-anti for molecule (b) in the crystal, while that of O6mGuo is syn [Parthasarathy, R & Fridey, S. M. (1986) Carcinogenesis 7, 221-227]. The sugar ring pucker of O6mdGuo is C(2')-endo for molecule (a) and C(1')-exo for molecule (b). The C(4')-C(5') exocyclic bond conformation in O6mdGuo is gauche- for molecule (a) but trans for molecule (b), in contrast with gauche+ for O6mGuo. The hydrogen bonds exhibited by O6-methylguanine derivatives differ from those in guanine derivatives; the amino N(2) and ring N(3) and N(7) atoms of O6-methylguanine residues are involved in hydrogen bonding. 1H-NMR data for O6mdGuo and O6mdGuo reveal the predominance of a C(2')-endo type sugar puckering. In O6mdGuo, however, a contribution of a C(1')-exo sugar puckering is significant. The NOE data also indicate that O6mdGuo molecules exist with nearly equal population for anti (including high anti) and syn glycosidic conformations. These observations and their biological implications are discussed.     

Theoretical studies of cis-Pt(II)-diammine binding to duplex DNA

The binding of cis-Pt(II) diammine (cis-DP) to double-stranded DNA was studied with several kinked conformations that can accommodate the formation of a square planar complex. Molecular mechanics (MM) calculations were performed to optimize the molecular fit. These results were combined with quantum mechanical (QM) calculations to ascertain the relative energetics of ligand binding through water vs direct binding of the phosphate to the ammine and platinum, and to guide the selection of DNA conformations to model complex formation. Based on QM and MM calculations, models are proposed that may be characterized by several general features. A structure involving hydrogen bonding between each ammine and distinct adjacent phosphate groups, referred to as closed conformation (CC), has already been reported. This is also found in the crystal structure of small dimers. We report alternative conformations that may be important in platination of duplex DNA. They are characterized by an intermediate conformation (IC), involving hydrogen bonding between one ammine and phosphate group, and an open conformation (OC), without ammine phosphate hydrogen bonding. The IC and OC can be stabilized by water bridges in the space between the ammine and the phosphate groups. Sugar puckers alternate from the type C(2')-endo or C(1')-exo (S), to the type C(3')-endo or C(2')-exo (N), with intermediate types near O(1')-endo (O). In general, the sugar puckers alternate from S to N to S through the platinated region (3'-TpG*pG*p-5'), with the complexed strand exhibiting, (3')-S*-N*-S-(5') alternation, while the complementary strand shows either (3')-S*-N*-S-(5') or (3')-S*-N*-O-(5') alternation. In both the OC and IC, a hydrogen bond is found between the ammine and O4(T) on thymine (T) at the (3') end, adjacent to the complex site. There is a continuous range of backbone conformations through the platinated region which relate the OC to the IC. The models presented suggest that the dynamics of the binding of the cis-Pt(II)-diammines to adjacent N7(G) in double-stranded DNA may encompass several conformational possibilities, and that water bridges may play a roll in supporting open and intermediate conformations. Proton-proton distances are reported to assist in the experimental determination of conformations.     

8-Chloroguanosine: solid-state and solution conformations and their biological implications

The three-dimensional structure of 8-chloroguanosine dihydrate was determined by X-ray crystallography. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), and the cell dimensions are a = 4.871 (1) A, b = 12.040 (1) A, and c = 24.506 (1) A. The structure was determined by direct methods, and least-squares refinement, which included all hydrogen atoms, converged at R = 0.031 for 1599 observed reflections. The conformation about the glycosidic bond is syn with chi CN = -131.1 degrees. The ribose ring has a C(2')-endo/C-(1')-exo (2T1) pucker, and the gauche+ conformation of the -CH2OH side chain is stabilized by an intramolecular O-(5')-H...N(3) hydrogen bond. Conformational analysis by means of 1H NMR spectroscopy showed that, in dimethyl sulfoxide, the sugar ring exhibits a marked preference for the C(2')-endo conformation (approximately 70%) and a conformation about the glycosidic bond predominantly syn (approximately 90%), hence similar to that in the solid state. However, the conformation of the exocyclic 5'-CH2OH group exhibits only a moderate preference for the gauche+ rotamer (approximately 40%), presumably due to the inability to form the intramolecular hydrogen bond to N(3) in a polar medium. The conformational features are examined in relation to the behavior of 8-substituted purine nucleosides in several enzymatic systems, with due account taken of the steric bulk and electronegativities of the 8-substituents.     

Absolute configuration of Rp-uridine 3'',5''-cyclic phosphorothioate.

Triethylammonium uridine-3',5'-cyclic phosphorothioate crystallizes in space group P2(1)2(1)2(1), a = 7.177(1), b = 13.155(6), c = 21.114(7) A, C15H26N3O7PS, MW 423.4, Z = 4, dx = 1.41g/cm3. The crystal structure was solved by direct methods on the basis of 1493 counter X-ray diffraction data (CuK alpha) and refined to R = 5.1%. The configuration of the thiophosphate group is Rp; conformational parameters are: glycosyl torsion angle anti, -151.9(5) degrees, sugar pucker C(3')-endo with P = 27.3 degrees, vmax = 45.5 degrees, six-membered cycle in chair form. The bond distances in the non-esterified P-S and P-O suggest that the negative charge is distributed between the groups. As illustrated in this and other studies, P-O has a much higher affinity for hydrogen bonds than P-S, indicated here by interactions with triethyl-ammonium N-H and O(2')-H as donors. One additional hydrogen bond N(3)-H---0(4) ties the bases which form a ribbon-like structure. 0(2) and S are not engaged in hydrogen bonds. The triethylammonium ion is two-fold disordered.     

Crystal structure of formycin 5'-phosphate: an explanation for its tight binding to AMP nucleosidase

Formycin 5'-monophosphate (FMP) is a strong competitive inhibitor of AMP nucleosidase with Km/Kis from 1200 to 2600 depending on the source of the enzyme. The crystal structure of FMP has been determined in order to understand the basis for its high affinity for AMP nucleosidases and other biological properties. The key structural features of FMP are (1) the base is the N(7)-H tautomer, (2) the N(3) of the base forms an intramolecular hydrogen bond to the phosphate oxygen O(1), (3) the glycosyl torsion angle is syn with O(4')-C(1') relative to C(9)-C(4) being -6.43 degrees, and (4) the furanose ring pucker is C(3')-endo, with a pseudorotation angle of 20.3 degrees. The major difference between the AMP and FMP structures is that the glycosyl torsion angles differ by 190 degrees. The computed conformational energy necessary to distort AMP so that it has the same glycosyl torsion angle as FMP is 4.6 kcal/mol. This corresponds to a 2100-fold difference in binding energy, in good agreement with the observed interaction between AMP nucleosidase and FMP.     

Molecular mechanics calculations on a triple stranded DNA involving C+.G-T and T.A+-C mismatched base triples

We have carried out molecular modeling of a triple stranded pyrimidine(Y). purine(R): pyrimidine(Y) (where ':' refers to Watson-Crick and '.' to Hoogsteen bonding) DNA, formed by a homopurine (d-TGAGGAAAGAAGGT) and homo-pyrimidine (d-CTCCTTTCTTCC). Molecular mechanics calculations using NMR constraints have provided a detailed three dimensional structure of the triplex. The entire stretches of purine and the pyrimidine nucleotides have a conformation close to B-DNA. The three strands are held by the canonical C+.G:C and T.A:T hydrogen bonds. The structure also contains two mismatch C+.G-T and T.A+-C base triples which have been characterized for the first time. In the A+-C base-pair of the T.A+-C triple, both hydrogen donors are situated on the purine (A+(1N) and A+(6N)). We observe a unique hydrogen bonding interaction scheme in case of C+.G-T where one acceptor, G(60), is bonded to three donors (C+(3NH), C+(4NH2) and T(3NH)). Though the C+.G-T base triple is less stable than C+.G:C, it is significantly more stable than T.A:T. On the other hand, T.A+-C is as stable as the T.A:T base triad.     

Self base pairing in a complementary deoxydinucleoside monophosphate duplex: crystal and molecular structure of deoxycytidylyl-(3'-5')-deoxyguanosine

The molecular structure of ammonium deoxycytidylyl-(3'-5')-deoxyguanosine, crystallized from aqueous acetone near pH 4, was determined for X-ray diffraction data. The crystals were tetragonal, space group P43212 with a = b = 11.078 (1) A and c = 45.826 (4) A. The structure was solved by tangent expansion of phases based on a derived phosphorus position and refined to R = 0.060 by full matrix least squares. Molecules related by a 2-fold symmetry axis are connected by hydrogen bonds between the bases and form parallel right-handed duplexes. Pairs of cytosines share a proton at N(3) and are joined by three hydrogen bonds: N(4)-H...O(2)...H-N(4), and N(3)-H...N(3). Guanines are joined by two hydrogen bonds: N(2)-H...N(3) and N(3)...H-N(2). Base-stacking interactions within the duplex are weak with the cytosine and guanine ring planes inclined at 24 degrees to each other in each monomer. Despite the unusual arrangement of the molecules, the sugar phosphate backbone has the g-g- conformation normally associated with right-handed double helical structures. Conformational parameters of the nucleosides are also typical with both sugars C(2')-endo and glycosidic torsion angles 55 degrees for cytidine and 94 degrees for guanosine. The bonding geometry of the bases is influenced by hydrogen bonding and charge-transfer networks in the crystal lattice. The solvent molecules interact with the dimer in three fused circular hydrogen bonding domains with a single disordered ammonium cation per d(CpG) dimer. Parallels with the formation of self base pairs and their implications in molecular biology are discussed.     

Investigations on the dynamic structures of adenine- and thymine-containing DNA.

The structures of poly(dA-dT), poly(dA-dBr5U) and of poly(dA).poly(dT) have been investigated in solution and in fibers, by Raman spectroscopy. Both the alternating poly(dA-dT), poly(dA-dBr5U) and non-alternating poly(dA).poly(dT) exhibit, in the region of sugar phosphate backbone vibrations, two bands of almost equal intensity at about 841 cm-1 and 817 cm-1. The analysis of the characteristic bands of thymine residues that are sensitive to sugar puckers gives indication of a significant displacement from the C(2')-endo conformer suggesting the adoption of alternative conformers such as O(4')-endo. In contrast, the diagnostic Raman bands for the sugar pucker of adenine residues suggest, instead, predominant adoption of C(2')-endo conformations. These Raman results are compatible with rapid dynamic changes of sugar puckers between C(2')-endo and O(4')-endo for the thymidine (and uridine) residues, whereas in adenine residues the sugar puckers fluctuate around the C(2')-endo pucker in all synthetic DNA molecules studied. Molecular dynamics simulations, performed on six different starting models using two distance-dependent dielectric functions epsilon(r) = 4 r and a sigmoidal dependence), all gave similar dynamic behavior in agreement with these Raman data and their interpretation. The mean calculated pseudorotation phases of the adenine residues are systematically higher (around C(2')-endo) than those of the thymine residues (close to O(4')-endo-C(1')-exo). Besides, the mean lifetimes of the thymine residues are 1.5 to 2.0-fold higher in the O(4')-endo than in the C(2')-endo domain, while those of the adenine residues are two to threefold higher in the C(2')-endo than in the O(4')-endo domain. In the Raman spectra of the alternating poly(dA-dBr5U), the splitting of a band into two components arising from the two contributions of ApBr5U and Br5UpA provides strong evidence for a repeating dinucleotide structure in solution. The calculated twist values averaged over the simulation runs are also systematically higher in the 5'T-A3' step (39 degrees) than in the 5'A-T3' step (33 degrees). Simultaneously, the calculated roll values are positive in the 5'T-A3' step (6 degrees) and negative in the 5'A-T3' step (-9 degrees), while the propeller twist values are about the same (-11 degrees to -16 degrees). On the other hand, in the homopolymer, the average twist value is close to 36 degrees with the roll angle close to 0 degrees and large propeller twist values (-20 degrees).     

Molecular orbital studies on the structure of nucleoside analogs. I. Conformation of 8-azapurine nucleosides

PCILO (perturbative configuration interaction using localized orbitals) computations have been carried out for the conformational properties of 8-azapurine nucleosides. The results indicate an anti conformation for Xcn and a gg conformation for phiC(4')-C(5') for C(2')-endo 8-aza analogs compared to the syn and gg conformation for the corresponding purine nucleosides. For C(3')-endo sugar puckering, both molecules prefer the syn conformation due to intramolecular hydrogen bonding between O(5')-H of the sugar and N(3) of the base, the preference being more profound in 8-aza analogs. The crystallographic conformation 8-azaadenosine has been attributed to crystal forces. The available NMR data on 8-azapurine nucleosides are in agreement with the PCILO predictions.     

Conformation of the common purine (beta) ribosides in solution: further evidence for a correlation between N-S state of the ribose moiety and syn-anti equilibrium.

The solution conformations of adenosine, guanosine and inosine in liquid ND3 have been determined by NMR. Comparison of the Karplus analysis of the proton HR spectra of the ribose moiety obtained in this solvent with the data from aqueous solutions of A and I proves that the conformations of the nucleosides are very similar in both liquids. From the analysis of the vicinal coupling constants of the ring protons it has been deduced that the S state C(2')-endo is slightly preferred. The mole fraction in S approximates 0.6 for all three nucleosides. C-13 relaxation measurements have been applied in the determination of the correlation times for rotational diffusion. Only at temperatures below - 40 degrees C is the pseudo-rotation of the furanoside ring slowed down sufficiently for it not to contribute to the measured relaxation rates. From NOE studies and T1 measurements on the individual protons it is derived that the N, C(3')-endo, form of the ribose is correlated with an anti conformation of the base (Y approximately 210 degrees to 220 degrees) and the S, C(2')-endo, form of the ribose with a syn conformation of the base (Y approximately 30 degrees to 50 degrees). The glycosyl torsion angles derived for the two conformations of A, G, and I are equal within the limits of accuracy.     

Recognition of hairpin-containing single-stranded DNA by oligonucleotides containing internal acridine derivatives.

Oligodeoxynucleotides with an internal intercalating agent have been targeted to single-stranded sequences containing hairpin structures. The oligonucleotide binds to nonadjacent single-stranded sequences on both sides of the hairpin structure in such a way as to form a three-way junction. The acridine derivative is inserted at a position that allows it to interact with the three-way junction. The melting temperature (Tm) of complexes formed between the hairpin-containing target and oligonucleotides containing one internal acridine derivative was higher than that obtained with the same target and an unmodified oligonucleotide (DeltaTm = +13 degrees C). The internal acridine provided the oligonucleotide with a higher affinity than covalent attachment to the 5' end. Oligonucleotides could also be designed to recognize a hairpin-containing single-stranded nucleic acid by formation of Watson-Crick hydrogen bonds with a single-stranded part and Hoogsteen hydrogen bonds with the stem of the hairpin. An internal acridine derivative was introduced at the junction between the two domains, the double helix domain with Watson-Crick base pairs and the triple helix domain involving Hoogsteen base triplets in the major groove of the hairpin stem. Oligonucleotides with an internal acridine or an acridine at their 5' end have similar binding affinities for the stem-loop-containing target. The bis-modified oligonucleotide containing two acridines, one at the 5' end and one at an internal site, did not exhibit a higher affinity than the oligonucleotides with only one intercalating agent. The design of oligonucleotides with an internal intercalating agent might be of interest to control gene expression through recognition of secondary structures in single-stranded targets.     

Solution structure of a trans-opened (10S)-dA adduct of (+)-(7S,8R,9S,10R)-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene in a fully complementary DNA duplex: evidence for a major syn conformation

Two-dimensional NMR was used to determine the solution structure of an undecanucleotide duplex, d(CGGTCACGAGG).d(CCTCGTGACCG), in which (+)-(7S,8R,9S,10R)-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene is covalently bonded to the exocyclic N(6)() amino group of the central deoxyadenosine, dA(6), through trans addition at C10 of the epoxide (to give a 10S adduct). The present study represents the first NMR structure of a benzo[a]pyrene (10S)-dA adduct in DNA with a complementary T opposite the modified dA. Exchangeable and nonexchangeable protons of the modified duplex were assigned by the use of TOCSY (in D(2)O) and NOESY spectra (in H(2)O and D(2)O). Sequential NOEs expected for a B-type DNA conformation with typical Watson-Crick base pairing are observed along the duplex, except at the lesion site. We observed a strong intraresidue NOE cross-peak between H1' and H8 of the modified dA(6). The sugar H2' and H2' ' of dC(5) lacked NOE cross-peaks with H8 of dA(6) but showed weak interactions with H2 of dA(6) instead. In addition, the chemical shift of the H8 proton (7.51 ppm) of dA(6) appears at a higher field than that of H2 (8.48 ppm). These NOE and chemical shift data for the dA(6) base protons are typical of a syn glycosidic bond at the modified base. Restrained molecular dynamics/energy minimization calculations show that the hydrocarbon is intercalated from the major groove on the 3'-side of the modified base between base pairs A(6)-T(17) and C(7)-G(16) and confirm the syn glycosidic angle (58 degrees ) of the modified dA(6). In the syn structure, a weak A-T hydrogen bond is possible between the N3-H proton of T(17) and N7 of dA(6) (at a distance of 3.11 A), whereas N1, the usual hydrogen bonding partner for N3-H of T when dA is in the anti conformation, is 6.31 A away from this proton. The 10(S)-dA modified DNA duplex remains in a right-handed helix, which bends in the direction of the aliphatic ring of BaP at about 42 degrees from the helical axis. ROESY experiments provided evidence for interconversion between the major, syn conformer and a minor, possibly anti, conformer.     

An 15N NMR study of adenine-uracil base pair in a non-aqueous solvent

[1,3,7,9,10-15N]-2',3',5'-Tri-O-acetyl adenosine (A) and its 8-D and 8-Br derivatives (AD and ABr) were prepared from 95% 15N enriched adenosine obtained from microbial fermentation. The chemical shifts and nuclear Overhauser effects of 15N resonances were measured as a function of the concentration of the mixed 1-cyclohexyluracil. The limiting shift of each 15N resonance was calculated and the structure of the A-U dimer was estimated. From the shifts of 15N-1 and 15N-7 signals it is determined that ABr-U dimers prefer the Watson-Crick type hydrogen bond while the Hoogsteen type pairs are predominant in the A-U dimers.     

Molecular orbital studies on nucleoside analogs. II. Conformation of 6-azapyrimidine nucleosides.

PCILO (Perturbative Configuration Interaction using Localised Orbitals) computations have been carried out for three 6-azapyrimidine nucleosides, 6-azauridine, 6-azacytidine and 6-azathymidine, for both C(2')-endo and C(3')-endo pucker of the sugar ring. The results indicate a syn (chiCN=180 degrees) conformation followed by chiCN=90 degrees and gg conformation for C(3')-endo 6-aza analogs as compareed to the anti (chiCN=0 degrees) and gg conformation preferred by the corresponding pyrimidine nucleosides. For C(2')-endo sugar geometry, 6-azauridine and 6-azacytidine prefer, respectively, chiCN=0 degrees (anti) and phi C(4')-C(5')=60 degrees C (gg) and chiCN-240 degrees (syn) and phi C(4')-C(5')=120 degrees. The corresponding nucleosides, uridine and cytidine, show a preference for syn (chiCN=240 degrees) and gg and anti(chiCN=0 degrees) and gg , respectively. The X-ray crystallographic conformations of 6-azauridine and 6-azacytidine have been attributed to intermolecular hydrogen bonding and crystal packing forces. The results of PMR, CD and ORD studies on 6-azauridine and 6-azacytidine in aqueous solutions are in agreement with the PCILO predictions.     

The first example of a Hoogsteen base-paired DNA duplex in dynamic equilibrium with a Watson-Crick base-paired duplex--a structural (NMR), kinetic and thermodynamic study.

A single-point substitution of the O4' oxygen by a CH2 group at the sugar residue of A6 (i.e. 2'-deoxyaristeromycin moiety) in a self-complementary DNA duplex, 5'-d(C1G2C3G4A5A6T7T8C9G10C11G12)2(-3), has been shown to steer the fully Watson-Crick basepaired DNA duplex (1A), akin to the native counterpart, to a doubly A6:T7 Hoogsteen basepaired (1B) B-type DNA duplex, resulting in a dynamic equilibrium of (1A)<==>(1B): Keq = k1/k(-1) = 0.56+/-0.08. The dynamic conversion of the fully Watson-Crick basepaired (1A) to the partly Hoogsteen basepaired (1B) structure is marginally kinetically and thermodynamically disfavoured [k1 (298K) = 3.9 0.8 sec(-1); deltaHdegrees++ = 164+/-14 kJ/mol; -TdeltaS degrees++ (298K) = -92 kJ/mol giving a deltaG degrees++ 298 of 72 kJ/mol. Ea (k1) = 167 14 kJ/mol] compared to the reverse conversion of the Hoogsteen (1B) to the Watson-Crick (1A) structure [k-1 (298K) = 7.0 0.6 sec-1, deltaH degrees++ = 153 13 kJ/mol; -TdeltaSdegrees++ (298K) = -82 kJ/mol giving a deltaGdegrees++(298) of 71 kJ/mol. Ea (k-1) = 155 13 kJ/mol]. Acomparison of deltaGdegrees++(298) of the forward (k1) and backward (k-1) conversions, (1A)<==>(1B), shows that there is ca 1 kJ/mol preference for the Watson-Crick (1A) over the double Hoogsteen basepaired (1B) DNA duplex, thus giving an equilibrium ratio of almost 2:1 in favour of the fully Watson-Crick basepaired duplex. The chemical environments of the two interconverting DNA duplexes are very different as evident from their widely separated sets of chemical shifts connected by temperature-dependent exchange peaks in the NOESY and ROESY spectra. The fully Watson-Crick basepaired structure (1A) is based on a total of 127 intra, 97 inter and 17 cross-strand distance constraints per strand, whereas the double A6:T7 Hoogsteen basepaired (1B) structure is based on 114 intra, 92 inter and 15 cross-strand distance constraints, giving an average of 22 and 20 NOE distance constraints per residue and strand, respectively. In addition, 55 NMR-derived backbone dihedral constraints per strand were used for both structures. The main effect of the Hoogsteen basepairs in (1B) on the overall structure is a narrowing of the minor groove and a corresponding widening of the major groove. The Hoogsteen basepairing at the central A6:T7 basepairs in (1B) has enforced a syn conformation on the glycosyl torsion of the 2'-deoxyaristeromycin moiety, A6, as a result of substitution of the endocyclic 4'-oxygen in the natural sugar with a methylene group in A6. A comparison of the Watson-Crick basepaired duplex (1A) to the Hoogsteen basepaired duplex (1B) shows that only a few changes, mainly in alpha, sigma and gamma torsions, in the sugar-phosphate backbone seem to be necessary to accommodate the Hoogsteen basepair.