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Results for “Base Pairing”

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At least 19 recordsLinked to original sources

PubMed1990

Substitution of 2 base pairs (1 base pair per DNA half-site) within the Escherichia coli lac promoter DNA site for catabolite gene activator protein places the lac promoter in the FNR regulon.

The consensus DNA site for Escherichia coli catabolite gene activator protein (CAP) is 5'-AAATGTGATCTAGATCACATTT-3'. The proposed consensus DNA site for E. coli FNR is 5'-AAATTTGATATATATCAAATTT-3'. In this report, we show that substitution of 2 base pairs (1 base pair per DNA half-site) within the E. coli lac DNA site for CAP suffices to remove the lac promoter from the CAP regulon and to place the lac promoter in the FNR regulon. FNR stimulates transcription of the derivative of the lac promoter having G:C----T:A substitutions at base pair 5 each DNA half-site (13-fold stimulation). FNR does not stimulate transcription of the wild-type lac promoter, or of derivatives of the lac promoter having G:C----A:T or G:C----C:G substitutions at base pair 5 of each DNA half-site. Stimulation of transcription is strictly dependent on anaerobiosis. FNR-stimulated transcription initiates at the same base pair as does CAP-dependent transcription of the wild-type lac promoter.

Aerobiosis↗

PubMed2001

Yeast U1 snRNP-pre-mRNA complex formation without U1snRNA-pre-mRNA base pairing.

Base pairing between the 5' end of U1 snRNA and the conserved 5' splice site of pre-mRNA is important for commitment complex formation in vitro. However, the biochemical mechanisms by which pre-mRNA is initially recognized by the splicing machinery is not well understood. To evaluate the role of this base pairing interaction, we truncated U1 snRNA to eliminate the RNA-RNA interaction and surprisingly found that U1 snRNP can still form a nearly normal RNA-protein complex and maintain sequence specificity. We propose that some feature of U1 snRNP, perhaps one or more protein factors, is more important than the base pairing for initial 5' splice site recognition. In addition, at least five sets of interactions contribute to complex formation or stability. Only one of these is base pairing between the 5' splice site and the 5' end of U1 snRNA, without which the U1 snRNP-pre-mRNA complex is less stable and has a somewhat altered conformation.

5' Untranslated Regions↗

PubMed2001

Stretching single-stranded DNA: interplay of electrostatic, base-pairing, and base-pair stacking interactions.

Recent single-macromolecule observations revealed that the force/extension characteristics of single-stranded DNA (ssDNA) are closely related to solution ionic concentration and DNA sequence composition. To understand this, we studied the elastic property of ssDNA through the Monte Carlo implementation of a modified freely jointed chain (FJC), with electrostatic, base-pairing, and base-pair stacking interactions all incorporated. The simulated force-extension profiles for both random and designed sequences have attained quantitative agreements with the experimental data. In low-salt solution, electrostatic interaction dominates, and at low forces, the molecule can be more easily aligned than an unmodified FJC. In high-salt solution, secondary hairpin structure appears in ssDNA by the formation of base pairs between complementary bases, and external stretching causes a hairpin-coil structural transition, which is continuous for ssDNA made of random sequences. In designed sequences such as poly(dA-dT) and poly(dG-dC), the stacking potential between base pairs encourages the aggregation of base pairs into bulk hairpins and makes the hairpin-coil transition a discontinuous (first-order) process. The sensitivity of elongation to the base-pairing rule is also investigated. The comparison of modeling calculations and the experimental data suggests that the base pairing of single-stranded polynucleotide molecules tends to form a nested and independent planar hairpin structure rather than a random intersecting pattern.

Base Pairing↗

PubMed1971

The influence of neighboring base pairs upon base-pair substitution mutation rates.

The 2-aminopurine-induced transition, A.T --> G.C, was studied at particular sites in bacteriophage T4 as a function of the nearby base-pair composition of the DNA. Changing a base pair changed the transition rate at the adjacent base pair up to 23-fold, and at the next base pair by a lesser amount. Destabilization was achieved by replacing an A.T base pair by a G.C base pair.

Adenine↗

PubMed2003

Motifs in nucleic acids: molecular mechanics restraints for base pairing and base stacking.

In building and refining nucleic acid structures, it is often desirable to enforce particular base pairing and/or base stacking interactions. Energy-based modeling programs with classical molecular mechanics force fields do not lend themselves to the easy imposition of penalty terms corresponding to such restraints, because the requirement that two bases lie in or near the same plane (pairing) or that they lie in parallel planes (stacking) cannot be easily expressed in terms of traditional interactions involving two atoms (bonds), three atoms (angles), or four atoms (torsions). Here we derive expressions that define a collection of pseudobonds and pseudoangles through which molecular mechanics restraints for base pairing and stacking can be imposed. We have implemented these restraints into the JUMNA package for modeling DNA and RNA structures. JUMNA scripts can specify base pairing with a variety of standard geometries (Watson-Crick, Hoogsteen, wobble, etc.), or with user-defined geometries; they can also specify stacking arrangements. We have also implemented "soft-core" functions to modify van der Waals and electrostatic interactions to avoid steric conflicts in particularly difficult refinements where two backbones need to pass through one another. Test cases are presented to show the utility of the method. The restraints could be adapted for implementation in other molecular mechanics packages.

Base Pairing↗

PubMed1987

Genetic conversion of G.C base-pairs to A.U base-pairs in a transfer RNA.

The cloverleaf stem segments of the suppressor gene of bacteriophage T4 tRNA(Gln) contain ten G.C and ten A.U base-pairs. To gain a better appreciation of the G.C base-pair requirement, we isolated multiple mutants of this suppressor gene in which base-pairs of G.C were replaced by A.U. One active suppressor gene contained only A.U base-pairs on the anticodon stem, indicating that G.C base-pairs in this region of tRNA(Gln) are not essential for function. In contrast, replacement was not possible at two base-pairs on the D stem and at one base-pair on the T stem.

Base Composition↗

PubMed1997

Natural 30 base pair and 69 base pair deletion variants of the LMP1 oncogene do stimulate NF-kappaB-mediated transcription.

An increasing number of reports shows a link between the Epstein-Barr virus (EBV) and lymphoid neoplasia. The latent membrane protein 1 (LMP1) is likely to play a determinant role in this process since this EBV encoded protein has oncogenic properties and is usually expressed in EBV-associated lymphoproliferative diseases (LPD), except Burkitt's lymphoma. We previously identified in LPD patients mutational hot spots and a 30 bp or 69 bp deletion in the LMP1 gene region coding for the C-terminal domain. These deletions are located in an area shown to be important for the activation of the transcription factor NF-kappaB. These findings lead us to test whether these natural deletion variants may have a functional effect. We measured the stimulation of their activity using a luciferase reporter plasmid containing NF-kappaB responsive elements. We tested the NF-kappaB inducing activity of four naturally occurring LMP1 deletion variants. Our results show that these deletion variants activate NF-kappaB to the same level as the wild-type form, indicating that the crucial residues for NF-kappaB activation are conserved among the variants isolated and lie within the last 32 amino acids of the C-terminal domain of the LMP1 oncogene.

Base Composition↗

PubMed2005

An efficient unnatural base pair for a base-pair-expanded transcription system.

For the site-specific incorporation of artificial components into RNA by transcription, an efficient, unnatural base pair between 2-amino-6-(2-thiazolyl)purine (denoted as v) and 2-oxo(1H)pyridine (denoted as y) was developed. The substrates of y and 5-substituted y were site-specifically incorporated into RNA by T7 RNA polymerase opposite v in templates. The efficiency and fidelity of the v-y pairing in transcription were as high as those of the natural A-T(U) and G-C pairings. Furthermore, RNAs containing two adjacent y bases were also transcribed from DNA templates containing two v bases. This specific transcription allows the large-scale preparation of artificial RNAs and can be combined with other systems to simultaneously incorporate several different components into a transcript.

Base Pairing↗

PubMed1984

Circular dichroism measurements show that C.C+ base pairs can coexist with A.T base pairs between antiparallel strands of an oligodeoxynucleotide double-helix.

We have studied the coil-to-helix transition of the DNA oligomer d(C4A4T4C4), using circular dichroism measurements to monitor the formation of A.T base pairs within the central self-complementary A4T4 region and the formation of protonated C.C+ base pairs at the ends of the oligomer. We found that both A.T and C.C+ base pairs formed in a coordinated fashion as the temperature and pH were lowered. The CD data of the helix form of the oligomer were consistent with the presence of paired oligomers, but not with hairpin loops. The pKa for formation of C.C+ base pairs between the C4 ends of the oligomer was higher than the pKa for formation of C.C+ base pairs in d(C8), indicating that the formation of C.C+ base pairs in the oligomer was influenced by the presence of a paired A4T4 region. We conclude that A.T and C.C+ base pairs coexist in the self-complex of the oligomer and, therefore, that C.C+ base pairs can form between antiparallel DNA strands.

Base Sequence↗

PubMed1980

Molecular cloning of bovine thyroglobulin complementary DNA. Characterization of 2500-base-pair and 1900-base-pair fragments.

Double-stranded thyroglobulin complementary DNA (cDNA) was synthesized from purified 33-S bovine thyroglobulin mRNA. This synthetic structural gene has previously been shown to contain three sites for the restriction endonuclease HindIII, yielding two internal fragments of 1900 and 2500 base pairs respectively. Recombinant molecules were prepared by ligating the HindIII-restricted cDNA to the plasmid pBR322 which had been linearized by the same enzyme. When Escherichia coli was transformed with this mixture, it yielded two kinds of colonies each harboring recombinant plasmids containing one of the two cDNA fragments. Both recombinant molecules hybridized specifically to translatable thyroglobulin mRNA. Sequence homology between the two cloned DNAs could not be detected by cross-hybridization experiments; this argues against the existence of internal structural repetition in thyroglobulin subunits. Together, the two cloned DNA fragments represent 55% of the 8000-base-pair double-stranded thyroglobulin DNA.

Animals↗

PubMed2003

Structure and the energy of base pairing in non-natural bases of nucleic acids: the azaguanine-cytosine and azaadenine-thymine base pairs.

Watson-Crick optimized geometries and the energies of base pairing for the natural pairs of nucleic bases: adenine-thymine (AT) and guanine-cytosine (GC) have been recalculated by ab initio methods in order to compare results to those found for the non-natural azaadenine-thymine (AAT) and azaguanine-cytosine (AGC) pairs. Geometry optimizations carried out at the HF/6-31G** level and energies obtained at MP2/6-31G**, show that AAT and AGC have hydrogen bonding patterns similar to the natural AT and GC and that the interaction energies (DeltaH0int) for the former are ca. 7 kcal/mol more stable than the latter. Accordingly, the pairs based on azapurines would be favored with respect to the natural pairs. Some possible explanations why nature does not use extensively the azabases in base pairing are given.

Azaguanine↗

PubMed2007

Hydrogen-bonded nucleic acid base pairs containing unusual base tautomers: complete basis set calculations at the MP2 and CCSD(T) levels.

The total interaction energies of altogether 15 hydrogen-bonded nucleic acid base pairs containing unusual base tautomers were calculated. The geometry properties of all selected adenine-thymine and guanine-cytosine hydrogen-bonded base pairs enable their incorporation into DNA. Unusual base pairing patterns were compared with Watson-Crick H-bonded structures of the adenine-thymine and guanine-cytosine pairs. The complete basis set (CBS) limit of the MP2 interaction energy and the CCSD(T) correction term, determined as the difference between the CCSD(T) and MP2 interaction energies, was evaluated. Extrapolation to the MP2 CBS limit was done using the aug-cc-pVDZ and aug-cc-pVTZ results, and the CCSD(T) correction term was determined with the 6-31G*(0.25) basis set. Final interaction energies were corrected while taking into account both tautomeric penalization determined at the CBS level and solvation/desolvation free energies. The situation for the adenine-thymine pairs is straightforward, and tautomeric pairs are significantly less stable than the Watson-Crick pair consisting of the canonical forms. In the case of the guanine-cytosine pair, the Watson-Crick structure made by canonical forms is again the most stable. The other two structures are, however, energetically rather similar (by 5 and 6 kcal/mol), which provides a very small but non-negligible chance of detecting these structures in the DNA double helix (1:5000). Due to the fact that DNA bases and base pairs incorporated into DNA are solvated less favorably than in isolated systems, this probability represents the very upper limit. The results clearly show how precisely the canonical building blocks of DNA molecules were chosen and how well their stability is maintained.

Chemical Phenomena↗

PubMed1976

Chemical evidence that chromatin DNA exists as 160 base pair beads interspersed with 40 base pair bridges.

Digestion of rat liver nuclei by an endogenous endonuclease generates double-stranded DNA fragments which are initially about 205 base pairs long, as reported previously by Hewish and Burgoyne. As digestion proceeds, the average size of these fragments is reduced to about 160 base pairs. Electrophoresis under denaturing conditions shows that these DNA fragments contain single strand nicks at ten base intervals. Fifteen bands, 10-150 bases, are clearly resolvable. DNA Fragments of 160 to 200 nucleotides are not resolved as distinct species. The results suggest that the chromosomal subunit contains both a 160 base-pair DNA segment, in a conformation susceptible to single strand nicking at ten base intervals, and a forty base-pair DNA segment in a conformation more uniformly susceptible to endogenous endonuclease activity. This chemical evidence agrees with morphological observations suggesting that chromatin has a "bead and bridge" structure.

Animals↗

PubMed1998

High apparent rate of simultaneous compensatory base-pair substitutions in ribosomal RNA.

We present a model for the evolution of paired bases in RNA sequences. The new model allows for the instantaneous rate of substitution of both members of a base pair in a compensatory substitution (e.g., A-U-->G-C) and expands our previous work by allowing for unpaired bases or noncanonical pairs. We implemented the model with distance and maximum likelihood methods to estimate the rates of simultaneous substitution of both bases, alphad, vs. rates of substitution of individual bases, alphas in rRNA. In the rapidly evolving D2 expansion segments of Drosophila large subunit rRNA, we estimate a low ratio of alphad/alphas, indicating that most compensatory substitutions involve a G-U intermediate. In contrast, we find a surprisingly high ratio of alphad/alphas in the core small subunit rRNA, indicating that the evolution of the slowly evolving rRNA sequences is modeled much more accurately if simultaneous substitution of both members of a base pair is allowed to occur approximately as often as substitution of individual bases. Using simulations, we have ruled out several potential sources of error in the estimation of alphad/alphas. We conclude that in the core rRNA sequences compensatory substitutions can be fixed so rapidly as to appear to be instantaneous.

Animals↗

PubMed2001

At nonzero temperatures, stacked structures of methylated nucleic acid base pairs and microhydrated nonmethylated nucleic acid base pairs are favored over planar hydrogen-bonded structures: a molecular dynamics simulations study.

The dynamic structure of all ten possible nucleic acid (NA) base pairs and methylated NA base pairs hydrated by a small number of water molecules (from 1 to 16) was determined by using molecular dynamics simulations in the NVE microcanonical and NVT canonical ensembles with the Cornell force field (W. D. Cornell, P. Cieplak, C. I. Bayly, I. R. Gould, K. M. Merz, D. M. Ferguson, D. C. Spellmeyer, T. Fox, J. E. Caldwell, P. Kollman, J. Am. Chem. Soc. 1995, 117, 5179). The presence of one water molecule does not affect the structure of any hydrogen-bonded (H-bonded) nonmethylated base pair. An equal population of H-bonded and stacked structures of adenine...adenine, adenine...guanine and adenine... thymine pairs is reached if as few as two water molecules are present, while obtaining equal populations of these structures in the case of adenine...cytosine, cytosine...thymine, guanine... guanine and guanine...thymine required the presence of four water molecules, and in the case of guanine...cytosine, six. A comparable population of planar, H-bonded and stacked structures for cytosine...cytosine and thymine... thymine base pairs was only obtained if at least eight water molecules hydrated a pair. Methylation of bases changed the situation dramatically and stacked structures were favoured over H-bonded ones even in the absence of water molecules in most cases. Only in the case of methyl cytosine...methyl cytosine, methyl guanine...methyl guanine and methyl guanine...methyl cytosine pairs were two, two or six water molecules, respectively, needed in order to obtain a comparable population of planar, H-bonded and stacked structures. We believe that these results give clear evidence that the preferred stacked structure of NA base pairs in the microhydrated environment, and also apparently in a regular solvent, is due to the hydrophilic interaction of a small number of water molecules. In the case of methylated bases, it is also due to the fact that the hydrogen atoms most suitable for the formation of H-bonds have been replaced by a methyl group. A preferred stacked structure is, thus, not due to a hydrophobic interaction between a large bulk of water molecules and the base pair, as believed.

Base Pairing↗

PubMed1984

O6-Methylguanine-DNA transmethylase converts O6-methylguanine thymine base pairs to guanine thymine base pairs in DNA.

O6-Methylguanine lesions in natural and synthetic DNAs were studied as substrates for the O6-methylguanine-DNA methyltransferase in vitro. The results indicate that O6-methylguanine is repaired by this protein when base paired to T as well as to C in double-stranded DNA. The results also indicate that O6-methylguanine is less subject to repair than previously reported when it is in single-stranded DNA and suggest that O6-methylguanine may not be repaired when it is at the 3' terminus.

Base Composition↗

PubMed1987

B-Z junctions in supercoiled pRW751 DNA contain unpaired bases or non-Watson-Crick base pairs.

Structural distortions on the boundary between right-handed and left-handed DNA segments in negatively supercoiled plasmid pRW751 (a derivative of pBR322 containing (dC-dG)13 and (dC-dG)16 segments) were studied by means of osmium tetroxide, pyridine and glyoxal. These two probes react preferentially with single-stranded DNA, but only the latter requires non-paired bases for the reaction. Nuclease S1 and testing of the inhibition of BamHI cleavage (whose recognition sequences GGATCC lie on the "outer" boundaries between the (dC-dG)n and the pBR322 nucleotide sequence) were used to detect the site-specific chemical modification in pRW751. As a result of glyoxal treatment BamHI cleavage was strongly inhibited in topoisomeric samples whose superhelical density was sufficiently negative to stabilize the (dC-dG)n segments in the left-handed form. Osmium tetroxide, pyridine modification resulted in a similar inhibition of BamHI cleavage and in a formation of nuclease S1 sensitive sites. The results suggest that the "outer" B-Z junctions in pRW751 contain one or few non-paired bases or non-Watson-Crick base pairs.

Base Composition↗

PubMed1999

High base pair opening rates in tracts of GC base pairs.

Sequence-dependent structural features of the DNA double helix have a strong influence on the base pair opening dynamics. Here we report a detailed study of the kinetics of base pair breathing in tracts of GC base pairs in DNA duplexes derived from 1H NMR measurements of the imino proton exchange rates upon titration with the exchange catalyst ammonia. In the limit of infinite exchange catalyst concentration, the exchange times of the guanine imino protons of the GC tracts extrapolate to much shorter base pair lifetimes than commonly observed for isolated GC base pairs. The base pair lifetimes in the GC tracts are below 5 ms for almost all of the base pairs. The unusually rapid base pair opening dynamics of GC tracts are in striking contrast to the behavior of AT tracts, where very long base pair lifetimes are observed. The implication of these findings for the structural principles governing spontaneous helix opening as well as the DNA-binding specificity of the cytosine-5-methyltransferases, where flipping of the cytosine base has been observed, are discussed.

Base Pairing↗