Pyrimidine Base Pairs Research Articles

Heterobimetallic Base Pair Programming in Designer 3D DNA Crystals.

Metal-mediated DNA (mmDNA) presents a pathway toward engineering bioinorganic and electronic behavior into DNA devices. Many chemical and biophysical forces drive the programmable chelation of metals between pyrimidine base pairs. Here, we developed a crystallographic method using the three-dimensional (3D) DNA tensegrity triangle motif to capture single- and multi-metal binding modes across granular changes to environmental pH using anomalous scattering. Leveraging this programmable crystal, we determined 28 biomolecular structures to capture mmDNA reactions. We found that silver(I) binds with increasing occupancy in T-T and U-U pairs at elevated pH levels, and we exploited this to capture silver(I) and mercury(II) within the same base pair and to isolate the titration points for homo- and heterometal base pair modes. We additionally determined the structure of a C-C pair with both silver(I) and mercury(II). Finally, we extend our paradigm to capture cadmium(II) in T-T pairs together with mercury(II) at high pH. The precision self-assembly of heterobimetallic DNA chemistry at the sub-nanometer scale will enable atomistic design frameworks for more elaborate mmDNA-based nanodevices and nanotechnologies.

Stem Region of tRNA Genes Favors Transition Substitution Towards Keto Bases in Bacteria.

Transversion and transition mutations have variable effects on the stability of RNA secondary structure considering that the former destabilizes the double helix geometry to a greater extent by introducing purine:purine (R:R) or pyrimidine:pyrimidine (Y:Y) base pairs. Therefore, transversion frequency is likely to be lower than that of transition in the secondary structure regions of RNA genes. Here, we performed an analysis of transition and transversion frequencies in tRNA genes defined well with secondary structure and compared with the intergenic regions in five bacterial species namely Escherichia coli, Klebsiella pneumoniae, Salmonella enterica, Staphylococcus aureus and Streptococcus pneumoniae using a large genome sequence data set. In general, the transversion frequency was observed to be lower than that of transition in both tRNA genes and intergenic regions. The transition to transversion ratio was observed to be greater in tRNA genes than that in the intergenic regions in all the five bacteria that we studied. Interestingly, the intraspecies base substitution analysis in tRNA genes revealed that non-compensatory substitutions were more frequent than compensatory substitutions in the stem region. Further, transition to transversion ratio in the loop region was observed to be significantly lesser than that among the non-compensatory substitutions in the stem region. This indicated that the transversion is more deleterious than transition in the stem regions. In addition, substitutions from amino bases (A/C) to keto bases (G/T) were also observed to be more than the reverse substitutions in the stem region. Substitution from amino bases to keto bases are likely to facilitate the stable G:U pairing unlike the reverse substitution that facilitates the unstable A:C pairing in the stem region of tRNA. This work provides additional support that the secondary structure of tRNA molecule is what drives the different substitutions in its gene sequence.

Isomorphic building blocks for information-bearing duplexes—part 2: pyrimidine base pairs with sugar phosphate backbones

Earlier searches using DFT had identified two isomorphic H-bonded pyrimidine base pairs B1:B2 and B3:B4 as possible repeat units for macromolecular H-bonded duplexes with a capacity to store information as in DNA. This DFT study seeks to design repeat units with suitable backbones which can hold a sequence of base pairs in desired order. A deoxyribose phosphate backbone and a deoxyallopyranose phosphate backbone are proposed here. These sets of backboned nucleotide pairs are examined for their stability and their isomorphism. The results yield stable H-bonded nucleotide pairs with pairing energies of the order of those for the Watson-Crick DNA base pairs. Isomorphism is examined in the central pyrimidine base pair moiety, in the backbone torsional angles and in the overall topology. Out of three different sets of nucleotide pairs, the anionic deoxyribonucleotide pairs DA1:DA2 and DA3:DA4 emerge as showing the maximal overall isomorphism, while the pyranonucleotide pairs PyA1:PyA2 and PyA3:PyA4 are predicted as the most stable.

Anatomy of Base Pairing in DNA by Interacting Quantum Atoms.

The formation of purine and pyrimidine base pairs (BPs), which contributes to shaping of the canonical and noncanonical 3D structures of nucleic acids, is one the most investigated phenomena in chemistry and life sciences. In this contribution, the anatomy of the bond energy (BDE) of the base-pairing interaction in 39 different arrangements found experimentally or predicted for DNA structures containing the four common nucleobases (A, C, G, T) in their neutral or protonated forms is described in light of the theory of interacting quantum atoms within the context of the quantum theory of atoms in molecules. The interplay of individual energy components involved in the three stages of the bond formation process (structural deformation, electron-density promotion, and intermolecular interaction) is studied. We recognized that for the neutral BPs, variations in the kinetic and electrostatic contributions to the BDE are rather negligible, leaving the exchange-correlation energy as the main stabilizing component. It is shown that the contribution of the exchange-correlation term can be recovered by including atoms that are formally assumed to be hydrogen bonded (primary interaction). In contrast, to recover the electrostatic component of interaction, one must consider both the primary and secondary (formally nonbonded atoms) interatomic interactions. The results of our study were employed to design new types of BPs with altered bonding anatomy. We demonstrate that improving the electrostatic characteristics of the BPs does not necessarily result in greater interaction energies if weak secondary hydrogen bonding is destroyed. However, the main tuning factor for systems with conserved interacting faces (primary interactions) is the electrostatic component of the interaction energy resulting from the secondary atom-atom electrostatics.

Structure of an RNA helix with pyrimidine mismatches and cross-strand stacking.

The structure of a 22-base-pair RNA helix with mismatched pyrimidine base pairs is reported. The helix contains two symmetry-related CUG sequences: a triplet-repeat motif implicated in myotonic dystrophy type 1. The CUG repeat contains a U-U mismatch sandwiched between Watson-Crick pairs. Additionally, the center of the helix contains a dimerized UUCG motif with tandem pyrimidine (U-C/C-U) mismatches flanked by U-G wobble pairs. This region of the structure is significantly different from previously observed structures that share the same sequence and neighboring base pairs. The tandem pyrimidine mismatches are unusual and display sheared, cross-strand stacking geometries that locally constrict the helical width, a type of stacking previously associated with purines in internal loops. Thus, pyrimidine-rich regions of RNA have a high degree of structural diversity.

Isomorphic repeat units for putative information-bearing H-bonded duplexes. Part I. Pyrimidine base pairs with polyamide backbone

Abstract Synthetic information-bearing macromolecular H-bonded duplexes built up from suitable sub-units are the aim of many attempts in design and synthesis. Repeat units for such duplexes had been earlier designed computationally, consisting of two isomorphic H-bonded pyrimidine base pairs with information-storing potential like the DNA base pairs. A backbone for this set of four pyrimidine bases is now designed to make complete repeat units suitable for incorporation into the putative polymeric duplex. Each strand of the duplex has the repetitive form (-CH2-CHB-CO-NH-)n with B as a pyrimidine base, so the backbone has a polyamide structure. Each backboned monomer is thus defined as NH2-CH2-CHB-COOH (2-amino-1-pyrimidylpropionic acid). The four monomeric units are paired through H-bonding to yield two backboned pairs, studied using density functional theory at the ωB97XD/6-311++G(d,p) level. Parallel and antiparallel backbone orientations are considered, each yielding two backboned H-bonded pairs. The results predict these H-bonded repeat unit pairs to be isomorphic and stable, suggesting a periodic structure for the putative H-bonded macromolecular duplex, which would possess the same information-bearing capacity as DNA.

The spontaneous replication error and the mismatch discrimination mechanisms of human DNA polymerase β.

To provide molecular-level insights into the spontaneous replication error and the mismatch discrimination mechanisms of human DNA polymerase β (polβ), we report four crystal structures of polβ complexed with dG•dTTP and dA•dCTP mismatches in the presence of Mg2+ or Mn2+. The Mg2+-bound ground-state structures show that the dA•dCTP-Mg2+ complex adopts an ‘intermediate’ protein conformation while the dG•dTTP-Mg2+ complex adopts an open protein conformation. The Mn2+-bound ‘pre-chemistry-state’ structures show that the dA•dCTP-Mn2+ complex is structurally very similar to the dA•dCTP-Mg2+ complex, whereas the dG•dTTP-Mn2+ complex undergoes a large-scale conformational change to adopt a Watson–Crick-like dG•dTTP base pair and a closed protein conformation. These structural differences, together with our molecular dynamics simulation studies, suggest that polβ increases replication fidelity via a two-stage mismatch discrimination mechanism, where one is in the ground state and the other in the closed conformation state. In the closed conformation state, polβ appears to allow only a Watson–Crick-like conformation for purine•pyrimidine base pairs, thereby discriminating the mismatched base pairs based on their ability to form the Watson–Crick-like conformation. Overall, the present studies provide new insights into the spontaneous replication error and the replication fidelity mechanisms of polβ.

ChemInform Abstract: Binding of Metal Ions by Pyrimidine Base Pairs in DNA Duplexes

AbstractReview: 86 refs.

Binding of metal ions by pyrimidine base pairs in DNA duplexes

Pyrimidine base pairs in DNA duplexes selectively capture metal ions to form metal ion-mediated base pairs, which can be evaluated by thermal denaturation, isothermal titration calorimetry, and nuclear magnetic resonance spectroscopy. In this critical review, we discuss the metal ion binding of pyrimidine bases (thymine, cytosine, 4-thiothymine, 2-thiothymine, 5-fluorouracil) in DNA duplexes. Thymine-thymine (T-T) and cytosine-cytosine (C-C) base pairs selectively capture Hg(II) and Ag(I) ions, respectively, and the metallo-base pairs, T-Hg(II)-T and C-Ag(I)-C, are formed in DNA duplexes. The metal ion binding properties of the pyrimidine-pyrimidine pairs can be changed by small chemical modifications. The binding selectivity of a metal ion to a 5-fluorouracil-5-fluorouracil pair in a DNA duplex can be switched by changing the pH of the solution. Two silver ions bind to each thiopyrimidine-thiopyrimidine pair in the duplexes, and the duplexes are largely stabilized. Oligonucleotides containing these bases are commercially available and can readily be applied in many scientific fields (86 references).

The Many Facets of Adenine: Coordination, Crystal Patterns, and Catalysis

Canonical purine-pyrimidine base pairs, the key to the complementary hydrogen bonding in nucleic acids, are fundamental molecular recognition motifs crucial for the formation and stability of double-helical DNA. Consequently, focused study and modeling of nucleobase hydrogen-bonding schemes have spawned a vast array of chemical and biophysical investigations. The Watson-Crick, reverse Watson-Crick, Hoogsteen, and reverse Hoogsteen hydrogen-bonding schemes stabilize various nucleic acid structures. As a result, numerous modified bases have been designed to maximize such interactions, addressing specific problems related to base pairing and giving rise to supramolecular ensembles in solution or in the solid state. It is also important to realize that suitably predisposed imino nitrogens and other functional groups present in heterocyclic nucleobases present a versatile molecular framework for the construction of coordination architectures, which may be harnessed to mimic base polyads and higher order nucleic acid structures. Adenine, a purine nucleobase, is an important naturally occurring nitrogen heterocycle present in nucleic acids. It is notable that the adenine unit is also frequently encountered as an inextricable part of enzyme cofactors and second messenger systems, such as NAD(+), FADH(2), and cAMP, which are essential for certain catalytic reactions and biochemical processes. In addition, a crucial catalytic role of the adenine moiety is also observed in group II intron catalysis and at the ribosomal peptidyltransferase center. Such versatile functional roles of the adenine framework serve as an inspiration for addressing research problems, ranging from classical coordination chemistry to the development of new materials. In this Account, we begin by describing the emerging use of adenine nucleobase for the design of metal-nucleobase frameworks. The coordination of metal ions affords a variety of oligomeric and polymeric species; we focus on silver- and copper-based structures and also discuss ferrocenylated adenine tetrads. We then consider the use of supramolecular adenine coordination complexes for transferring molecular properties onto surfaces. This technique is particularly useful for transferring noncovalent interactions, such as van der Waals forces, electrostatic interactions, and hydrogen bonding, to designed architectures in nanoscale applications. Finally, we explore the issue of adenine-based catalytic entities. Here, adenine moieties are first fixed in a polymeric matrix, followed by metalation of the matrix. These metalated adenine-containing polymers are then assayed for catalytic assistance in various chemical and biochemical reactions. Taken together, the versatile coordination abilities and hydrogen-bonding capacity of adenine offer a novel entry point for a natural ligand into materials synthesis.

Selective Recognition of Unnatural Imidazopyridopyrimidine:Naphthyridine Base Pairs Consisting of Four Hydrogen Bonds by the Klenow Fragment

In this work, we investigated how thermally stable ImO(N):NaN(O) and ImN(O):NaO(N) pairs are recognized by the Klenow fragment (KF). As a result, these complementary base pairs, especially the ImN(O):NaO(N) pair, were recognized selectively due to the four hydrogen bonds between the nucleobases and the shape complementarity of the Im:Na pair similar to the purine:pyrimidine base pair.

Preparation of DNA double helical structures containing both mercury and silver ions

DNA duplexes consisting of metal-mediated pyrimidine base pairs, T-Hg(II)-T and C-Ag(I)-C, were prepared by mixing short oligodeoxyribonucleotides (ODNs) in the presence of the metal ions. The metal ions were arrayed linearly in the prepared double helical structures. The choice of sequence controls the order of the metal ions.

Breaking pseudo-twofold symmetry in the poliovirus 3'-UTR Y-stem by restoring Watson-Crick base pairs

The previously described NMR structure of a 5'-CU-3'/5'-UU-3' motif, which is highly conserved within the 3'-UTR Y-stem of poliovirus-like enteroviruses, revealed striking regularities of the local helix geometry, thus retaining the pseudo-twofold symmetry of the RNA helix. A mutant virus with both pyrimidine base pairs changed into Watson-Crick replicated as wild type, indicating the functional importance of this symmetry relation in viral RNA replication. Here we investigated the effect of changing only one of the two pyrimidine base pairs to Watson-Crick. We determined the NMR structures of two Y-stem variants: one containing the 5'-CU-3'/5'-AU-3' motif, which has been found in wild-type virus isolates as well, and the other containing a 5'-CU-3'/5'-UG-3' motif, which is not present in any enterovirus sequenced to date. Both structures show single pyrimidine mismatches with intercalated bases. In the 5'-CU-3'/5'-AU-3' motif a C-U Watson-Crick-type base pair is formed that retains the pseudo-twofold symmetry, while in the 5'-CU-3'/5'-UG-3' motif a single asymmetric U-U mismatch breaks the twofold symmetry. Surprisingly, for the nonnatural variant no effect of the single base-pair replacement was observed on polioviral RNA replication using an in vitro replicon assay.

Theoretical Study on the Interaction of Platinum Compounds with DNA Base Pairs

The binding of a series of square planar Pt(Ⅱ) adducts on the Watson-Crick base pair AT and GC was investigated using advanced quantum chemical method. The result showed that the AT base pair was mainly influenced by electrostatic effect, the GC base pair was influenced by both electrostatic and polarization effect. The platinated purine and pyrimidine base pair was moderately strengthened, whereas interaction between the purine and pyrimidine base pair was slightly influenced compared with that of the isolated one. Further calculations on the Pd(Ⅱ) and Ni(Ⅱ) analogs showed a similar trend. The natural bonding orbital (NBO) analysis of the second-order perturbative energy ∆E2, well predicated strength of the H-bond in the base pair.

Raman Spectroscopic Analysis of the Effect of Ultraviolet Irradiation on Calf Thymus DNA

Raman spectroscopy was used for the first time to detect the effect of independent UVA (ultraviolet-A: 320-400 nm) and UVB (ultraviolet-B: 280-320 nm) irradiation on the calf thymus DNA in aqueous solution. After both UVA and UVB irradiation for 1 h or 3 h, the damage to the conformation of DNA was moderate, but the reduction of the B-form DNA component was obvious. Both UVA and UVB caused significant damage to the deoxyribose moiety and bases, among which the pyrimidine base pairs were more seriously affected. There appeared to be preferential damaging sites on DNA molecules caused by UVA and UVB irradiation. UVA irradiation caused more damage to the deoxyribose than UVB irradiation, while UVB irradiation caused more significant damage to the pyrimidine moiety than UVA irradiation. After UVB irradiation for 3 h, unstacking of the AT base pairs and the cytosine ring took place, severe damage to the thymine moiety occurred, and some base pairs were modified. Moreover, with either UVA or UVB irradiation for 3 h, the photoreactivation of DNA occurred. The damage to the DNA caused by UVB was immediate, while the damage caused by UVA was proportional to the irradiation duration. The experimental results partly indicate the formation of some cyclobutane pyrimidine dimers and (6-4) photoproducts.

Solution structure of a consensus stem-loop D RNA domain that plays important roles in regulating translation and replication in enteroviruses and rhinoviruses.

Stem-loop D from the cloverleaf RNA is a highly conserved domain within the 5'-UTR of enteroviruses and rhinoviruses. Interaction between the stem-loop D RNA and the viral 3C or 3CD proteins constitutes an essential feature of a ribonucleoprotein complex that plays a critical role in regulating viral translation and replication. Here we report the solution NMR structure of a 38-nucleotide RNA with a sequence that encompasses the entire stem-loop D domain and corresponds to the consensus sequence found in enteroviruses and rhinoviruses. Sequence variants corresponding to Poliovirus type 1 and Coxsackievirus B3 have virtually the same structure, based on small differences in chemical shifts. A substantial number (136) of (1)H-(13)C one-bond residual dipolar coupling (RDC) values were used in the structure determination in addition to conventional distance and torsion angle restraints. Inclusion of the RDC restraints was essential for achieving well-defined structures, both globally and locally. The structure of the consensus stem-loop D is an elongated A-type helical stem capped by a UACG tetraloop with a wobble UG closing base pair. Three consecutive pyrimidine base pairs (two UU and one CU pair) are present in the middle of the helical stem, creating distinctive local structural features such as a dramatically widened major groove. A dinucleotide bulge is located near the base of the stem. The bulge itself is flexible and not as well defined as the other parts of the molecule, but the flanking base pairs are intact. The peculiar spatial arrangement of the distinctive structural elements implies that they may work synergistically to achieve optimal binding affinity and specificity toward the viral 3C or 3CD proteins.

YNMG tetraloop formation by a dyskeratosis congenita mutation in human telomerase RNA.

Autosomal dominant dyskeratosis congenita (DKC) has been linked to mutations in the RNA component of telomerase, the ribonucleoprotein responsible for telomere maintenance. Recent studies have investigated the role of the GC (107-108) --> AG mutation in the conserved P3 helix in the pseudoknot domain of human telomerase RNA. The mutation was found to significantly destabilize the pseudoknot conformation, resulting in a shift in the thermodynamic equilibrium to favor formation of a P2b hairpin intermediate. In the wild-type sequence, the hairpin intermediate was found to form a novel sequence of pyrimidine base pairs in a continuous stem capped by a structured pentaloop. The DKC mutant hairpin was observed to be slightly more stable than the wild-type hairpin, further shifting the pseudoknot-hairpin equilibrium to favor the mutant P2b hairpin. Here we examined the solution structure of the DKC mutant hairpin to identify the reason for this additional stability. We found that the mutant hairpin forms the same stem structure as wild-type and that the additional stabilization observed using optical melting can be explained by the formation of a YNMG-type tetraloop structure, with the last nucleotide of the pentaloop bulged out into the major groove. Our results provide a structural explanation for the increased stability of the mutant hairpin and further our understanding of the effect of this mutation on the structure and stability of the dominant conformation of the pseudoknot domain in this type of DKC.

8-Aza-7-deazapurine–pyrimidine base pairs: the contribution of 2- and 7-substituents to the stability of duplex DNA

The 8-aza-7-deazapurin-2,6-diamine 2′-deoxyribonucleoside ( 2a )–dT base pair is more stable than the 2-amino-2′-deoxyadenosine ( 1 )–dT pair. Halogeno or propynyl substituents introduced in the 7-position of the 8-aza-7-deazapurine system ( 2b– d ) lead to an additional duplex stabilization. Pyrazolo[3,4- d]pyrimidine nucleosides related to 2′-deoxyadenosine or 2′-deoxyguanosine show a similar behavior. The base pair stabilization is comparably small when identical substituents are present in the 5-position of 2′-deoxyuridine. The extraordinary stability of the 2b– d –dT base pairs is caused by better proton donor properties of the two amino groups of the 8-aza-7-deazapurin-2,6-diamine moiety and the better base stacking induced by the 7-substituents compared to that of 2-aminoadenine.

NMR Studies of DNA Structures Containing Sheared Purine•Purine and Purine•Pyrimidine Base Pairs

Summary Sheared purine•purine or purine•pyrimidine base pairs are important motifs in nucleic acid structures. They can exist either as tandem base pairs or as a single base pair closing hairpin mini-loops. Presence of such stable motifs greatly increases the structural repertoire of nucleic acid molecules and may play important roles in the contemporary nucleic acid studies. Several RNA crystal structures containing adjacent cross-strand base stacking have been solved. In this review, I will discuss the NMR studies and structural features for DNA molecules containing sheared G•A, A•A, or A•C base pairs. Special attention will be focused on exchangeable proton data, which have turned out to be crucial to the determination of unusual nucleic acid structures. The special features of the partial HIV-1 RT 21-mer act as a good example for correlating the 31P chemical shifts to the ζ and α torsional angles of the backbone phosphate diesters.

Measurement of Local DNA Reorganization on the Picosecond and Nanosecond Time Scales

Picosecond and nanosecond dynamics in the interior of DNA are observed for the first time as a dynamic Stokes shift in the fluorescence of a specially designed base-pair analogue. A recently synthesized coumarin C-riboside has been incorporated into an oligonucleotide in place of a normal purine·pyrimidine base pair. The dynamic Stokes shift of this probe occurs with components near 300 ps and 13 ns. These times are too long for solvent relaxation or simple vibrational motion of the DNA framework. They imply that the interior of DNA is a unique dynamic environment unlike either a fluid or a molecular crystal.