Ab initio conformational analysis of nucleic acid components: intrinsic energetic contributions to nucleic acid structure and dynamics

In recent years, the use of high-level ab initio calculations has allowed for the intrinsic conformational properties of nucleic acid building blocks to be revisited. This has provided new insights into the intrinsic conformational energetics of these compounds and its relationship to nucleic acids structure and dynamics. In this article we review recent developments and present new results. New data include comparison of various levels of theory on conformational properties of nucleic acid building blocks, calculations on the abasic sugar, known to occur in vivo in DNA, on the TA conformation of DNA observed in the complex with the TATA box binding protein, and on inosine. Tests of the Hartree-Fock (HF), second-order M?ller-Plesset (MP2), and Density Functional Theory/Becke3, Lee, Yang and Par (DFT/B3LYP) levels of theory show the overall shape of backbone torsional energy profiles (for gamma, epsilon, and chi) to be similar for the different levels, though some systematic differences are identified between the MP2 and DFT/B3LYP profiles. The east pseudorotation energy barrier in deoxyribonucleosides is also sensitive to the level of theory, with the HF and DFT/B3LYP east barriers being significantly lower (approximately 2.5 kcal/mol) than the MP2 counterpart (approximately 4.0 kcal/mol). Additional calculations at various levels of theory suggest that the east barrier in deoxyribonucleosides is between 3.0 and 4.0 kcal/mol. In the abasic sugar, the west pseudorotation energy barrier is found to be slightly lower than the east barrier and the south pucker is favored more than in standard nucleosides. Results on the TA conformation suggest that, at the nucleoside level, this conformation is significantly destabilized relative to the global energy minimum, or relative to the A- and B-DNA conformations. Deoxyribocytosine would destabilize the TA conformation more than other bases relative to the A-DNA conformation, but not relative to the B-DNA conformation.     

Intrinsic conformational properties of deoxyribonucleosides: implicated role for cytosine in the equilibrium among the A, B, and Z forms of DNA.

Structural properties of biomolecules are dictated by their intrinsic conformational energetics in combination with environmental contributions. Calculations using high-level ab initio methods on the deoxyribonucleosides have been performed to investigate the influence of base on the intrinsic conformational energetics of nucleosides. Energy minima in the north and south ranges of the deoxyribose pseudorotation surfaces have been located, allowing characterization of the influence of base on the structures and energy differences between those minima. With all bases, chi values associated with the south energy minimum are lower than in canonical B-DNA, while chi values associated with the north energy minimum are close to those in canonical A-DNA. In deoxycytidine, chi adopts an A-DNA conformation in both the north and south energy minima. Energy differences between the A and B conformations of the nucleosides are <0.5 kcal/mol in the present calculations, except with deoxycytidine, where the A form is favored by 2.3 kcal/mol, leading the intrinsic conformational energetics of GC basepairs to favor the A form of DNA by 1.5 kcal/mol as compared with AT pairs. This indicates that the intrinsic conformational properties of cytosine at the nucleoside level contribute to the A form of DNA containing predominately GC-rich sequences. In the context of a B versus Z DNA equilibrium, deoxycytidine favors the Z form over the B form by 1.6 kcal/mol as compared with deoxythymidine, suggesting that the intrinsic conformational properties of cytosine also contribute to GC-rich sequences occurring in Z DNA with a higher frequency than AT-rich sequences. Results show that the east pseudorotation energy barrier involves a decrease in the furanose amplitude and is systematically lower than the inversion barrier, with the energy differences influenced by the base. Energy barriers going from the south (B form) sugar pucker to the east pseudorotation barrier are lower in pyrimidines as compared with purines, indicating that the intrinsic conformational properties associated with base may also influence the sugar pseudorotational population distribution seen in DNA crystal structures and the kinetics of B to A transitions. The present work provides evidence that base composition, in addition to base sequence, can influence DNA conformation.     

Stereochemical studies on nucleic acid analogues. I. Conformations of alpha-nucleosides and alpha-nucleotides: interconversion of sugar puckers via O4'-exo.

The preferred conformations of ribo and deoxyribo alpha-nucleosides and alpha-nucleotides, the stereoisomers of the naturally occurring beta-isomers, are worked out by minimizing the conformational energy as a function of all the major parameters including the sugar ring conformations along the pseudorotation path. The results of the studies bring out certain distinct conformational features that are at variance with their beta counterparts. The range of glycosyl conformations are found to be not only quite restricted here but favor predominantly the anti conformation. The syn glycosyl conformation for the entire region of P values are found to be energetically less favorable, with the barrier to anti in equilibrium with syn interconversion being higher especially in alpha-ribonucleosides. The energetically preferred range of pseudorotation phase angles (P) is also considerably restricted and P values corresponding to the C1'-exo range of sugars are highly unfavorable for alpha-nucleosides, in sharp contrast to the broad range of sugar ring conformations favored by beta-isomers. While both trans congruent to 180 degrees and skew congruent to 270 degrees conformations around the C3'-O3' (phi') bond are favored for alpha-3'-nucleotides with deoxyribose sugars, ribose sugars seem to favor only the skew values of phi'. Most interestingly and in sharp contrast to beta-stereoisomers, an energy barrier is encountered at P values corresponding to O4'-endo sugars. This suggests that the possible sugar pucker interconversion between C2'-endo/C3'-exo and C3'-endo/C2'-exo in alpha-anomers could take place only through the O4'-exo region. Likewise the possible path of anti in equilibrium with syn interconversion in alpha-nucleosides is not via high anti, in sharp contrast to beta-nucleosides. These observations should be borne in mind while assigning the sugar ring conformers in alpha-nucleosides and those containing them from nmr investigations. Comparison of the results with beta-anomers seem to suggest on the whole a lack of conformational variability or the restricted nature of alpha-stereoisomers. This could be one of the reasons for its nonselection in the naturally occurring nucleic acids.     

The occurrence of the syn-C3′ endo conformation and the distorted backbone conformations for C4′-C5′ and P-O5′ in oligo and polynucleotides

Abstract

The nucleoside constituents of nucleic acids prefer the anti conformation (1). When the sugar pucker is taken into account the nucleosides prefer the C2′endo-anti conformation. Of the nearly 300 nucleosides known, about 250 are in the anti conformation and 50 are in the syn-conformation, i.e., anti to syn conformation is 5:1. The nucleotide building blocks of nucleic acids show the same trend as nucleosides. Both the deoxy-guanosine and ribo- guanosine residues in nucleosides and nucleotides prefer the syn-C2′endo conformation with an intra-molecular hydrogen bond (for nucleosides) between the O5′- H and the N3 of the base and, a few syn-C3′endo conformations are also observed. Evidence is presented for the occurrence of the C3′endo-syn conformation for guanines in mis-paired double helical right-handed structures with the distorted sugar phosphate C4′-C5′ and P-O5′ bonds respectively, from g+ (gg) and g- to trans. Evidence is also provided for guanosine nucleotides in left-handed double-helical (Z-DNA) oligo and polynucleotides which has the same syn-C3′endo conformation and the distorted backbone sugar-phosphate bonds (C4′-C5′ and P- O5′) as in the earlier right-handed case.     

The occurence of the syn-C3' endo conformation and the distorted backbone conformations for C4'-C5' and P-O5' in oligo and polynucleotides.

The nucleoside constituents of nucleic acids prefer the anti conformation (1). When the sugar pucker is taken into account the nucleosides prefer the C2'endo-anti conformation. Of the nearly 300 nucleosides known, about 250 are in the anti conformation and 50 are in the syn-conformation, i.e., anti to syn conformation is 5:1. The nucleotide building blocks of nucleic acids show the same trend as nucleosides. Both the deoxy-guanosine and riboguanosine residues in nucleosides and nucleotides prefer the syn-C2'endo conformation with an intra-molecular hydrogen bond (for nucleosides) between the O5'-H and the N3 of the base and, a few syn-C3'endo conformations are also observed. Evidence is presented for the occurrence of the C3'endo-syn conformation for guanines in mis-paired double helical right-handed structures with the distorted sugar phosphate C4'-C5' and P-O5' bonds respectively, from g+ (gg) and g- to trans. Evidence is also provided for guanosine nucleotides in left-handed double-helical (Z-DNA) oligo and polynucleotides which has the same syn-C3'endo conformation and the distorted backbone sugar-phosphate bonds (C4'-C5' and P-O5') as in the earlier right-handed case.     

Z-RNA: the solution NMR structure of r(CGCGCG).

Left-handed double-helical Z-RNA has been studied using the ribohexanucleotide pentaphosphate r(CpGpCpGpCpG). One-dimensional and two-dimensional proton nmr experiments were used to probe the structural details of the left-handed helix in concentrated sodium perchlorate solution. In 1M NaClO4 the RNA adopts the normal A-form double helix, and in 6M NaClO4 it is nearly all in the Z form. In 4M NaClO4 it exists as nearly equal parts of A form and Z form. Resonances corresponding to both A and Z form appear in the nmr spectrum, indicating that the duplex exchanges slowly between forms. Spin-spin coupling constants between protons in the ribose rings were used to determine the sugar-pucker conformations of the individual nucleotides. Quantitative nuclear Overhauser experiments were used to determine proton-proton distances within the nucleoside, and from these distances values for the glycosidic torsion angle were determined. The results show that the cytidines adopt C2'-endo sugar puckers (S type) with pseudo-rotation phase values (P) of approximately 165 degrees. The bases are in the anti conformation, with chi values of approximately -140 degrees. The internal guanosines adopt C3'-endo sugar puckers (N type) with P approximately 18 degrees, while the 3'-terminal guanosine ribose exists in an equilibrium between S- and N-type conformations. All three guanosine bases adopt the syn conformation, with chi approximately 70 degrees. The results indicate that the solution structure of Z-RNA is very similar to that of Z-DNA.     

Nuclear overhauser effect studies on the conformation of magnesium adenosine 5'-triphosphate bound to rabbit muscle creatine kinase

Nuclear Overhauser effects were used to determine interproton distances on MgATP bound to rabbit muscle creatine kinase. The internuclear distances were used in a distance geometry program that objectively determines both the conformation of the bound MgATP and its uniqueness. Two classes of structures were found that satisfied the measured interproton distances. Both classes had the same anti glycosidic torsional angle (chi = 78 +/- 10 degrees) but differed in their ribose ring puckers (O1'-endo or C4'-exo). The uniqueness of the glycosidic torsional angle is consistent with the preference of creatine kinase for adenine nucleotides. One of these conformations of MgATP bound to creatine kinase is indistinguishable from the conformation found for Co(NH3)4ATP bound to the catalytic subunit of protein kinase, which also has a high specificity for adenine nucleotides [chi = 78 +/- 10 degrees, O1'-endo; Rosevear, P.R., Bramson, H.N., O'Brian, C., Kaiser, E.T., & Mildvan, A.S. (1983) Biochemistry 22, 3439]. Distance geometry calculations also suggest that upper limit distances, when low enough (less than or equal to 3.4 A), can be used instead of measured distances to define, within experimental error, the glycosidic torsional angle of bound nucleotides. However, this approach does not permit an evaluation of the ribose ring pucker.     

Charges of phosphate groups. A role in stabilization of 2'-deoxyribonucleotides. A DFT investigation

We have analyzed the relative stabilities and Gibbs tautomeric free energy for tautomeric transitions of neutral 2'-deoxyribonucleotides and its mono- and di-protonated forms. Geometry optimizations of these nucleic acid constituents have been performed at the DFT/B3LYP level using the standard 6-31G(d) basis set. The prediction of relative stabilities, Gibbs tautomeric free energy has been made at the B3LYP/6-311++G(d,p)//B3LYP/6-31G(d) level of theory. For each nucleoside four major conformers, i.e., north/anti, north/syn, south/anti, and south/syn have been taken into consideration. We have found the substantial effect of the uncompensated charge on the relative stability of 2'-deoxyribonucleotides. In particular, when the charge of 2'-deoxyribonucleotide anions is completely compensated by protons, the syn conformations have been found to be the global minima due to stabilization provided by intramolecular hydrogen bonds. However, the negative charge that appears due to the successive removal of the protons from the phosphate group destabilizes these syn conformations and stabilizes preferably the south/anti conformations (except of 2'-deoxyguanosine phosphate). Only 2'-deoxyribonucleotides, possessing south/anti and north/anti orientations, containing guanine and cytosine can contribute significantly to the rate of spontaneous point mutations due to the formation of biologically relevant amounts of 'rare' tautomers. However, we found strong influence of uncompensated negative charge for 2'-deoxyribonucleotides which possess syn conformations. Finally we have found that the proton transfer could result in the spontaneous change of 2'-deoxyribonucleotides conformations. We conclude that this phenomenon could be considered as a new way for the stabilization of 'rare' isomers for such DNA bases as cytosine and thymine.     

Zipper-like Watson-Crick base-pairs

A series of DNA heptadecamers containing the DNA analogues of RNA E-like 5'-d(GXA)/(AYG)-5' motifs (X/Y is complementary T/A, A/T, C/G, or G/C pair) were studied using nuclear magnetic resonance (NMR) methodology and distance geometry (DG)/molecular dynamics (MD) approaches. Such oligomers reveal excellent resolution in NMR spectra and exhibit many unusual nuclear Overhauser effects (NOEs) that allow for good characterization of an unusual zipper-like conformation with zipper-like Watson-Crick base-pairs; the potential canonical X.Y H-bonding is not present, and the central X/Y pairs are transformed instead into inter-strand stacks that are bracketed by sheared G.A base-pairs. Such phenomenal structural change is brought about mainly through two backbone torsional angle adjustments, i.e. delta from C2'-endo to C3'-endo for the sugar puckers of unpaired residues and gamma from gauche(+) to trans for the following 3'-adenosine residues. Such motifs are analogous to the previously studied (GGA)(2) motif presumably present in the human centromeric (TGGAA)(n) tandem repeat sequence. The novel zipper-like motifs are only 4-7 deg. C less stable than the (GGA)(2) motif, suggesting that inter-strand base stacking plays an important role in stabilizing unusual nucleic acid structures. The discovery that canonical Watson-Crick G.C or A.T hydrogen-bonded pairs can be transformed into stacking pairs greatly increases the repertoire for unusual nucleic acid structural motifs.     

NMR study of stacking interactions and conformational adjustments in the dinucleotide-carcinogen adduct 2'-deoxycytidylyl-(3----5)-2'-deoxy-8-(N-fluoren-2-ylac etamido)guanosi ne

The conformation and dynamics of the dinucleotide d-CpG modified at the C(8) position of the guanine ring by the carcinogen 2-(acetylamino)fluorene has been investigated by high-field 1H NMR spectroscopy. A two-state analysis of chemical shift data has enabled estimation of the extent of intramolecular stacking in aqueous solution as a function of temperature. The stacking, which is mostly fluorene-cytosine, is virtually complete in the low-temperature range. The 500-MHz 1H NMR spectrum consists of two subspectra near ambient temperatures due to a 14.3 +/- 0.3 kcal/mol barrier to internal rotation about the amide bond in the stacked form. A large barrier to internal rotation about the guanyl-nitrogen bond at C(8) has also been ascertained, but separate NMR subspectra were not detected due to the predominance of one of the torsional diastereomers (alpha' = 90 degrees) in the fully stacked state. Problems of self-association and chemical exchange were identified and overcome to enable analysis of the sugar-phosphate backbone conformation utilizing coupling constants. For the exocyclic C(4')-C(5') bond of the deoxyguanosine moiety, there is a high gauche+ (gamma = 60 degrees) conformer population, which is uncommon for a purine nucleotide with a syn orientation about the glycosyl bond. The gauche- conformation (gamma = 300 degrees), which is normally present in syn purine nucleotides in solution, was not detected. The exocyclic C(5')-O(5') torsion of the deoxy-guanosine moiety remains near the classical energy minimum (beta = 180 degrees) in the major stacked conformations. The sugar ring of the deoxycytidine moiety is predominantly in the C2'-endo conformation, while the deoxyguanosine ring is a mixture of conformations, one of which appears to be unusually puckered. The results support intercalation models of modified DNA and suggest a looped-out structure, with the modified guanine being the first base in the loop. Such structures could explain the relatively rapid rate of repair and the frame-shift mutations of this type of adduct.     

NMR studies of conformations and interactions of substrates and ribonucleotide templates bound to the large fragment of DNA polymerase I

The large fragment of DNA polymerase I (Pol I) effectively uses oligoribouridylates and oligoriboadenylates as templates, with kinetic properties similar to those of poly(U) and poly(A), respectively, and has little or no activity in degrading them. In the presence of such oligoribonucleotide templates, nuclear Overhauser effects (NOE's) were used to determine interproton distances within and conformations of substrates bound to the large fragment of Pol I, as well as conformations and interactions of the enzyme-bound templates. In the enzyme-oligo(rU)54 +/- 11-Mg2+dATP complex, the substrate dATP has a high anti-glycosidic torsional angle (chi = 62 +/- 10 degrees) and an O1'-endo/C3'-endo sugar pucker (delta = 90 +/- 10 degrees) differing only slightly from those previously found for enzyme-bound dATP in the absence of template [Ferrin, L.J., & Mildvan, A.S. (1985) Biochemistry 24, 4680-4694]. Both conformations are similar to those of deoxynucleotidyl units of B DNA but differ greatly from those of A or Z DNA. The conformation of the enzyme-bound substrate analogue AMPCPP (chi = 50 +/- 10 degrees, delta = 90 +/- 10 degrees) is very similar to that of enzyme-bound dATP and is unaltered by the binding of the template oligo(rU)54 +/- 11 or by the subsequent binding of the primer (Ap)9A. In the enzyme-oligo(rA)50-Mg2+TTP complex, the substrate TTP has an anti-glycosidic torsional angle (chi = 40 +/- 10 degrees) and an O1'-endo sugar pucker (delta = 100 +/- 10 degrees), indistinguishable from those found in the absence of template and compatible with those of B DNA but not with those of A or Z DNA. In the absence of templates, the interproton distances on enzyme-bound dGTP cannot be fit by a single conformation but require a 40% contribution from a syn structure (chi = 222 degrees) and a 60% contribution from one or more anti structures. The presence of the template oligo(rU)43 +/- 9 simplifies the conformation of enzyme-bound dGTP to a single structure with an anti-glycosyl angle (chi = 32 +/- 10 degrees) and an O1'-endo/C3'-endo sugar pucker (delta = 90 +/- 10 degrees), compatible with those of B DNA, possibly due to the formation of a G-U wobble base pair. However, no significant misincorporation of guanine deoxynucleotides by the enzyme is detected with oligo(rU) as template.(ABSTRACT TRUNCATED AT 400 WORDS)     

Nuclear Overhauser effect studies of the conformations of MgATP bound to the active and secondary sites of muscle pyruvate kinase

MgATP binds both at the active site (site 1) and at a secondary site (site 2) on each monomer of muscle pyruvate kinase as previously found by binding studies and by X-ray analysis. Interproton distances on MgATP bound at each site have been measured by the time-dependent nuclear Overhauser effect in the absence and presence of phosphoenolpyruvate (P-enolpyruvate), which blocks ATP binding at site 1. Interproton distances at site 2 are consistent with a single conformation of bound ATP with a high antiglycosidic torsional angle (chi = 68 +/- 10 degrees) and a C3'-endo ribose pucker (delta = 90 +/- 10 degrees). Interproton distances at site 1, determined in the absence of P-enolpyruvate by assuming the averaging of distances at both sites, cannot be fit by a single adenine-ribose conformation but require the contribution of at least three low-energy structures: 62 +/- 10% low anti (chi = 30 degrees), C3'-endo; 20 +/- 8% high anti (chi = 55 degrees), O1'-endo; and 18 +/- 8% syn (chi = 217 degrees), C2'-endo. Although a different set of ATP conformations might also have fit the interproton distances, the mixture of conformations used also fits previously determined distances from Mn2+ to the protons of ATP bound at site 1 [Sloan, D. L., & Mildvan, A. S. (1976) J. Biol. Chem. 251, 2412] and is similar to the adenine-ribose portion of free Co(NH3)4ATP, which consists of 35% low anti, 51% high anti, and 14% syn [Rosevear, P. R., Bramson, H. N., O'Brian, C., Kaiser, E. T., & Mildvan, A. S. (1983) Biochemistry 22, 3439].(ABSTRACT TRUNCATED AT 250 WORDS)     

Conformational analysis of canonical 2-deoxyribonucleotides. 1. Pyrimidine nucleotides

The molecular structure and relative stability of north and south conformers of 2'-deoxyribonucleotides containing pyrimidine nucleic acid bases ( 2'-deoxythymidilic (pdT), 2'-deoxycytidilic (pdC) acids and their mono- and dianions) have been obtained and analyzed at the DFT/B3LYP level using the standard 6-31G(d) basis set. We have revealed that, when the nucleobase moiety is incorporated into the nucleotides, it maintains a nonplanar and nonrigid conformation due to out-of-plane deformation of the amino group and pyrimidine ring. It has been demonstrated that an increase of negative charge of the phosphate group results in increase of amino group pyramidalization, discrimination between conformers with syn and anti orientation of base with respect to sugar, strengthening of intramolecular C-H.O hydrogen bonds leading to deformation and fixation of geometry of nucleotides, and weakening of phosphodiester bond. These results allow to make suggestions about sources of twist and buckle deformations of base pairs, mechanisms of repaire of DNA via change of base orientation, and conditions for breakage of the P-O bonds during hydrolysis.     

Anti-Syn Conformational Range of Pyrimidines with Deoxyribofuranose

Abstract

The ability of pyrimidine bases to adopt the syn conformation in DNA has been investigated. The distances between atoms on the sugar and base and the resulting steric energies have been calculated as a function of glycosidic torsion angle for the principal sugar puckers of the deoxyribose of cytosine. The results indicate that pyrimidines can assume both the anti and syn conformations for the ³E, ₄E, ₁E, ²E, ₃E sugar puckers and syn for the ₂E sugar pucker. For these sugar puckers the difference between the minimum energies of the anti and syn conformations is in the range of 0.1–2.0 kcal/mole, with the minimum syn energy being lower in the case of the ₄E, ₁E and ²E sugar puckers. It is particularly significant that cytosine can assume the syn conformation for the ³E sugar pucker commonly observed for the syn nucleotides in Z-DNA with both alternating pyrimidine/purine (APP) and non-APP sequences. The results of this investigation confirm that steric interactions resulting from putting a pyrimidine nucleotide in the syn conformation are not a major factor in the preference for APP base sequences in Z-DNA.     

Conformational analysis of arabinonucleosides and nucleotides. A comparison with the ribonucleosides and nucleotides.

Conformations of arabino nucleosides and nucleotides have been analyzed by semiempirical energy calculations. It is found that the change in the configuration of the O(2')-hydroxyl group in arabinoses compared to riboses exerts significant influence on the conformational priorities of the glycosyl and the exocyclic C(4')-C(5') bond torsions. While the anti conformations for the bases are preferred, the anti in equilibrium or formed from syn interconversion is considerably hampered compared to ribosides due to large energy barrier. Further the preferred anti glycosyl torsions are shifted to higher values for C(3')-endo puckers and in ribosides. While the gauche+ conformation around the C(4')-C(5') bond is favored for C(3')-endo arabinosides, it is strongly stabilized for C(2')-endo arabinosides only in the presence of the intrasugar hydrogen bond O(2')-H ... O(5'). The net attractive electrostatic interactions between the phosphate and the base stabilizes the preferred conformations of 5'-arabinonucleotides also.     

Coupling of the guanosine glycosidic bond conformation and the ribonucleotide cleavage reaction: implications for barnase catalysis

To examine the possible relationship of guanine-dependent GpA conformations with ribonucleotide cleavage, two potential of mean force (PMF) calculations were performed in aqueous solution. In the first calculation, the guanosine glycosidic (Gchi) angle was used as the reaction coordinate, and computations were performed on two GpA ionic species: protonated (neutral) or deprotonated (negatively charged) guanosine ribose O2 '. Similar energetic profiles featuring two minima corresponding to the anti and syn Gchi regions were obtained for both ionic forms. For both forms the anti conformation was more stable than the syn, and barriers of approximately 4 kcal/mol were obtained for the anti --> syn transition. Structural analysis showed a remarkable sensitivity of the phosphate moiety to the conformation of the Gchi angle, suggesting a possible connection between this conformation and the mechanism of ribonucleotide cleavage. This hypothesis was confirmed by the second PMF calculations, for which the O2 '--P distance for the deprotonated GpA was used as reaction coordinate. The computations were performed from two selected starting points: the anti and syn minima determined in the first PMF study of the deprotonated guanosine ribose O2'. The simulations revealed that the O2 ' attack along the syn Gchi was more favorable than that along the anti Gchi: energetically, significantly lower barriers were obtained in the syn than in the anti conformation for the O--P bond formation; structurally, a lesser O2 '--P initial distance, and a better suited orientation for an in-line attack was observed in the syn relative to the anti conformation. These results are consistent with the catalytically competent conformation of barnase-ribonucleotide complex, which requires a guanine syn conformation of the substrate to enable abstraction of the ribose H2 ' proton by the general base Glu73, thereby suggesting a coupling between the reactive substrate conformation and enzyme structure and mechanism.     

Crystal structure and conformation of 5-fluorouridine: conformational preferences for 5-fluorinated pyranosides

Crystals of 5-fluorouridine (5FUrd) have unit cell dimensions a = 7.716(1), b = 5.861(2), c = 13.041(1)A, alpha = gamma = 90 degrees, beta = 96.70 degrees (1), space group P2(1), Z = 2, rho obs = 1.56 gm/c.c and rho calc = 1574 gm/c.c The crystal structure was determined with diffractometric data and refined to a final reliability index of 0.042 for the observed 2205 reflections (I > or = 3sigma). The nucleoside has the anti conformation [chi = 53.1(4) degrees] with the furanose ring in the favorite C2'-endo conformation. The conformation across the sugar exocyclic bond is g+, with values of 49.1(4) and -69.3(4) degrees for phi(theta c) and phi (infinity) respectively. The pseudorotational amplitude tau(m) is 34.5 (2) with a phase angle of 171.6(4) degrees. The crystal structure is stabilized by a network of N-H...O and O-H...O involving the N3 of the uracil base and the sugar 03' and 02' as donors and the 02 and 04 of the uracil base and 03' oxygen as acceptors respectively. Fluorine is neither involved in the hydrogen bonding nor in the stacking interactions. Our studies of several 5-fluorinated nucleosides show the following preferred conformational features: 1) the most favored anti conformation for the nucleoside [chi varies from -20 to + 60 degrees] 2) an inverse correlation between the glycosyl bond distance and the chi angle 3) a wide variation of conformations of the sugar ranging froni C2'-endo through C3'-endo to C4'-exo 4) the preferred g+ across the exocyclic C4'-C5' bond and 5) no role for the fluorine atom in the hydrogen bonding or base stacking interactions.