Base Pair Sites Research Articles

CeO2-catalyzed Transformations of Nitriles and Amides

Abstract Recently, cerium oxides have been extensively used as heterogeneous catalysts for green organic reactions. This review describes recent progress in such reactions, with a focus on our studies on pure CeO2-catalyzed reactions of nitriles and amides with nucleophiles (H2O, alcohols, and amines). Hydration of nitriles to amides in water, one-pot ester synthesis from nitriles and alcohols, one-pot synthesis of N-alkylamides from nitriles with amines, and synthesis of N-alkylamides from amides and amines (transamidation) are selectively catalyzed by CeO2 under aqueous or solvent-free conditions. Mechanistic studies suggest that a cooperation of Lewis acid site (Ce4+ cation) and acid–base (Ce–O) pair site plays essential roles in the catalytic reactions.

A Partner Evokes Latent Differences between Hox Proteins

Hox transcription factors bind highly related DNA sequences in vitro, yet they regulate different genes and play distinct roles in anterior-posterior patterning in animals. Slattery et al. report that a common cofactor, Exd, accentuates latent sequence specificities of all eight Hox proteins and directs binding to relevant sites across the genome.

Identification of beta-1 adrenergic receptor polymorphisms in cats

In human beings, genetic polymorphisms within the beta-1 adrenergic receptor (ADRB1) gene have been associated with variable pharmacologic responses to beta blocker therapy. Beta-blockers are commonly given to cats with heart disease, particularly hypertrophic cardiomyopathy, a common cause of feline heart disease. We hypothesized that polymorphisms are present in the feline ADRB1 gene, which could result in an altered pharmacologic response to beta-blocker therapy. We sequenced the feline ADRB1 gene in 42 cats of five breeds. We identified three polymorphisms within the ADRB1 gene. Two polymorphisms did not change the amino acid produced and are unlikely to be clinically significant. A third polymorphism identified was an AA/CC substitution at the 830–831 base pair sites. This alteration changed the amino acid produced from proline to glutamine at position 277 and computer modeling predicts an altered protein structure. Further study is warranted to determine if this polymorphism alters response to beta blocker therapy.

Selective hydrogenation of nitrocyclohexane to cyclohexanone oxime by alumina-supported gold cluster catalysts

Metal oxides (Al 2O 3, SiO 2, MgO)-supported Au cluster catalysts prepared by colloid deposition method and well established Au/TiO 2 prepared by deposition–precipitation method were tested for the selective reduction of nitrocyclohexane into cyclohexanone oxime. The activity and selectivity depended strongly on the size of Au and support material. Au/Al 2O 3 with smaller Au particle size (2.5 nm) was the most effective catalyst, exhibiting a high cyclohexanone oxime yield (86%) under mild condition (0.6 MPa H 2, 100 °C). To study the origin of support and size effects, in situ FTIR experiments for OH/D 2 exchange reaction and nitrobenzene adsorption combined with poisoning experiments using pyridine and acetic acid were carried out. It is suggested that cooperation of coordinatively unsaturated Au atoms and the acid–base pair site (Al δ+–O δ− site) plays an important role in H 2 dissociation step. Al δ+–O δ− site also acts as the adsorption site of the nitro group.

ChemInform Abstract: Design of a Nonnatural Deoxyribonucleoside for Recognition of GC Base Pairs by Oligonucleotide-Directed Triple Helix Formation.

The deoxyribonucleoside 1-(2-deoxy-beta-D-ribofuranosyl)-3-methyl-5-amino-1H-pyrazolo4,3-d]py rimidin-7-one (P1) was designed such that two specific hydrogen bonds would form with guanine (G) of a Watson-Crick guanine-cytosine (GC) base pair in the major groove of double-helical DNA. One edge of the P1 heterocycle mimics N3-protonated cytosine, which would circumvent the pH dependence observed for the formation of triple helices containing C + GC base triplets. P1 was synthesized in five steps and incorporated by automated methods in pyrimidine oligodeoxyribonucleotides. From affinity cleaving analyses, the stabilities of base triplets decrease in the order P1.GC >> P1.CG >> P1.AT approximately P1.TA (pH 7.4, 35-degrees-C). P1 binds GC base pairs within a pyrimidinte triple-helix motif as selectively and strongly as C but over an extended pH range. Oligodeoxyribonucleotides containing P1 residues were shown to bind within plasmid DNA a single 15 base pair site containing five GC base pairs at pH 7.8 and a single 16 base pair site containing six contiguous GC base pairs at pH 7.4.

NFATc4 is negatively regulated in miR-133a-mediated cardiomyocyte hypertrophic repression

Activation of NFAT (nuclear factor of activated T cells)-mediated hypertrophic signaling is a major regulatory response to hypertrophic stimuli. A recent study unveiled potential regulatory roles for microRNA-133a (miR-133a) in cardiac hypertrophy. To date, however, no connection has been made between miR-133a and NFAT signaling. In this study, we determined that NFATc4, a hypertrophy-associated mediator, is negatively regulated by miR-133a. Two conserved base-pairing sites between the NFATc4 3'-untranslated region (UTR) and miR-133a were verified. Mutation of these sites in the NFATc4 3'-UTR completely blocked the negative effect of miR-133a on NFATc4, suggesting that NFATc4 is a direct target for miR-133a regulation. Using a gain-of-function approach, we demonstrate that miR-133 significantly reduces the endogenous level of, as well as the hypertrophic stimulus-mediated increase in, NFATc4 gene expression. This latter effect of miR-133a on NFATc4 gene expression was coincided with an attenuated cardiomyocyte hypertrophy induced by an alpha-adrenergic receptor agonist. Conversely, cells treated with miR-133a inhibitor resulted in an increase in NFATc4 expression level. Application of miR-133a had no apparent effect on NFATc4 nuclear localization. We conclude that the negative regulation of NFATc4 expression contributes to miR-133a-mediated hypertrophic repression.

Size- and support-dependent silver cluster catalysis for chemoselective hydrogenation of nitroaromatics

Silver clusters on θ-Al 2O 3 support catalyze highly chemoselective reduction of a nitro group for the reduction of substituted nitroaromatics such as nitrostyrene. These catalysts show higher selectivity than conventional platinum-group metal-based heterogeneous catalysts. Systematic studies on the influence of the metal particle size and support oxides show that the intrinsic activity increases with decrease in the silver particle size, and acid–base bifunctional supports such as Al 2O 3 give higher activity than acidic or basic supports. Kinetic and in situ infrared studies provide a reaction mechanism which explains fundamental reasons of these tendencies. Cooperation of the acid–base pair site on Al 2O 3 and the coordinatively unsaturated Ag sites on the silver cluster is responsible for the rate-limiting H 2 dissociation to yield a H +/H − pair at metal/support interface, while the basic site on oxides acts as an adsorption site of nitroaromatics. High chemoselectivity could be attributed to a preferential transfer of the H +/H − pair to the polar bonds in the nitro group.

Influence of solitons on the conductance properties of double-stranded deoxyribonucleic acid

A numerical study is presented to investigate the role of solitons in the electronic states of double-stranded DNA (dsDNA) molecule in the metal/DNA/metal system. Based on tight-binding Hamiltonian model and within the framework of a generalized Green’s function technique, we consider a ladder model for poly(dG)-poly(dC) DNA molecule containing M cells with four sites (two base pair sites and two backbone sites) in each cell. In the presence of a sublattice of solitons, our results show that the homogeneous soliton distributions induce the electronic states in the band gap of DNA molecule. In addition, the room temperature current-voltage characteristic of the system shows a linear and ohmic-like behaviour.

Necessity of Quantum Coherence to Account for the Spectrum of Time-Dependent Mutations Exhibited by Bacteriophage T4

Transcriptase measurements of quantum expectations due to time-dependent coherent states populating informational DNA base-pair sites, designated by G-C --> *G-*C, G-C --> G'-C', and A-T --> *A-*T, provide a model for transcription and replication of time-dependent DNA lesions exhibited by bacteriophage T4. Coherent states are introduced as consequences of hydrogen bond arrangement, keto-amino --> enol-imine, where product protons are shared between two sets of indistinguishable electron lone-pairs and thus participate in coupled quantum oscillations at frequencies of ~10(13) s(-1). The transcriptase deciphers and executes genetic specificity instructions by implementing measurements on superposition proton states at *G-*C, G'-C', and *A-*T sites in an interval Δt << 10(-13) s. Decohered states participate in Topal-Fresco replication, which introduces substitutions *C --> T, *G --> A, G' --> T, and G' --> C, but superposition *A-*T states are deleted. These results imply an evolutionary shift favoring A-T richness.

Chemoselective Hydrogenation of Nitroaromatics by Supported Gold Catalysts: Mechanistic Reasons of Size- and Support-Dependent Activity and Selectivity

Supported Au nanoparticles (NPs) prepared by colloid deposition method were well characterized, and their catalytic performance was tested for chemoselective reduction of a nitro group of substituted nitroaromatics by H2. Systematic studies on the effects of NPs size and support show small size of Au NPs, and acid-base sites of supports are required for high activity. The Au/Al2O3 catalyst with Au particle size of 2.5 nm selectively hydrogenates a nitro group in the presence of various other reducible functional groups, and it shows higher intrinsic activity than the state-of-the-art catalyst (Au NPs on TiO2). In situ FTIR studies provide a reaction mechanism, which explains fundamental reasons of the observed structure−activity relationship. Cooperation of the acid−base pair site on Al2O3 and the coordinatively unsaturated Au atoms on the Au NPs are responsible for the H2 dissociation to yield a H+/H− pair at the metal/support interface. High chemoselectivity could be attributed to a preferential transfer of the H+/H− pair to the polar bonds in the nitro group as well as a preferential adsorption of nitroaromatics on the catalyst through the nitro group.

Transport properties of poly(GACT)-poly(CTGA) deoxyribonucleic acid: A ladder model approach

In this paper, based on the tight-binding Hamiltonian model and within the framework of a generalized Green’s function technique, the electronic conduction through the poly(GACT)-poly(CTGA) DNA molecule in SWNT/DNA/SWNT structure has been numerically investigated. In a ladder model, we consider DNA as a planar molecule containing M cells and four further sites (two base pair sites and two backbone sites) in each cell, sandwiched between two semi-infinite single-walled carbon nanotubes (SWNT) as the electrodes. Having relied on Landauer formalism, we focussed on studying the current-voltage characteristics of DNA, the effect of the coupling strength of SWNT/DNA interface and the role of tube radius of nanotube contacts on the electronic transmission through the foregoing structure. Finally, a characteristic time was calculated for the electron transmission, which measures the delay caused by the tunnelling through the SWNT/DNA interface. The results clearly show that the calculated characteristic time and also the conductance of the system are sensitive to the coupling strength between DNA molecule and nanotube contacts.

RNA Chaperones Stimulate Formation and Yield of the U3 snoRNA–Pre-rRNA Duplexes Needed for Eukaryotic Ribosome Biogenesis

Short duplexes between the U3 small nucleolar RNA and the precursor ribosomal RNA must form quickly and with high yield to satisfy the high demand for ribosome synthesis in rapidly growing eukaryotic cells. These interactions, designated the U3–ETS (external transcribed spacer) and U3–18S duplexes, are essential to initiate the processing of small subunit ribosomal RNA. Previously, we showed that duplexes corresponding to those in Saccharomyces cerevisiae are only observed in vitro after addition of one of two proteins: Imp3p or Imp4p. Here, we used fluorescence-based and other in vitro assays to determine whether these proteins possess RNA chaperone activities and to assess whether these activities are sufficient to satisfy the duplex yield and rate requirements expected in vivo. Assembly of both proteins with the U3 small nucleolar RNA into a chaperone complex destabilizes a U3 stem structure, apparently to expose its 18S base-pairing site. As a result, the chaperone complex accelerates formation of the U3–18S duplex from an undetectable rate to one comparable with the intrinsic rate observed for hybridizing short duplexes. The chaperone complex also stabilizes the U3–ETS duplex by 2.7 kcal/mol. These chaperone activities provide high U3–ETS duplex yield and rapid U3–18S duplex formation over a broad concentration range to help ensure that the U3–precursor ribosomal RNA interactions limit neither ribosome biogenesis nor rapid cell growth. The thermodynamic and kinetic framework used is general and thus suitable for investigating the mechanism of action of other RNA chaperones.

Evidence for transcriptase quantum processing implies entanglement and decoherence of superposition proton states

Evidence requiring transcriptase quantum processing is identified and elementary quantum methods are used to qualitatively describe origins and consequences of time-dependent coherent proton states populating informational DNA base pair sites in T4 phage, designated by G–C → G′–C′, G–C → *G–*C and AT → *A–*T. Coherent states at these ‘point’ DNA lesions are introduced as consequences of hydrogen bond arrangement, keto– amino → enol– imine, where product protons are shared between two sets of indistinguishable electron lone-pairs, and thus, participate in coupled quantum oscillations at frequencies of ∼10 13 s −1. This quantum mixing of proton energy states introduces stability enhancements of ∼0.25–7 kcal/mole. Transcriptase genetic specificity is determined by hydrogen bond components contributing to the formation of complementary interstrand hydrogen bonds which, in these cases, is variable due to coupled quantum oscillations of coherent enol–imine protons. The transcriptase deciphers and executes genetic specificity instructions by implementing measurements on superposition proton states at G′–C′, *G–*C and *A–*T sites in an interval Δ t ≪ 10 −13 s. After initiation of transcriptase measurement, model calculations indicate proton decoherence time, τ D, satisfies the relation Δ t < τ D < 10 −13 s. Decohered states participate in Topal–Fresco replication to introduce substitutions G′ → T, G′ → C, *C → T and *G → A. Measurements of 37 °C lifetimes of the keto–amino DNA hydrogen bond indicate a range of ∼3200–68,000 yrs. Arguments are presented that quantum uncertainty limits on amino protons may drive the keto– amino → enol– imine arrangement. Data imply that natural selection at the quantum level has generated effective schemes (a) for introducing superposition proton states – at rates appropriate for DNA evolution – in decoherence-free subspaces and (b) for creating entanglement states that augment (i) transcriptase quantum processing and (ii) efficient decoherence for accurate Topal–Fresco replication.

Fabrication, photoemission studies, and sensor of Hg nanoparticles templated on plasmid DNA

Fabrication of Hg nanoparticles (NPs) templated on plasmid DNA has been investigated here. The Hg NPs get embedded inside the DNA scaffold through local melting of double helix due to the strong and exclusive interaction of the NPs with the nitrogen of the nucleic acid bases. The interaction of the Hg NPs with the guanine-cytosine base pair sites is responsible for the formation of two Hg metal-base complexes that can find application as the signature of ion-DNA interactions. The modifications in the transport properties of metal conjugated DNA can be utilized as sensor for mercury contamination.

Minor Groove Binding Compounds That Jump a GC Base Pair and Bind to Adjacent AT Base Pair Sites

Most A/T specific heterocyclic diamidine derivatives need at least four A/T base pairs for tight binding to the DNA minor groove. Addition of a GC base pair to A/T sequences typically causes a large decrease in binding constant. The ability to target biologically important sequences of DNA could be significantly increased if compounds that could recognize A/T sites with an intervening GC base pair could be designed. The kinetoplast DNA sequence of parasitic microorganisms, for example, contains numerous three A/T binding sites that are separated by a single G. A series of compounds were prepared to target the AAAGTTT sequence as a model system for discovery of "G-jumpers". The new synthetic compounds have two aromatic-amidine groups for A/T recognition, and these are connected through an oxy-methylene linker to cross the GC. CD experiments indicated a minor groove binding mode, as expected, for these compounds. T(max), surface plasmon resonance, and isothermal titration calorimetry experiments revealed 1:1 binding to the AAAGTTT sequence with an affinity that depends on compound structure. Benzimidazole derivatives gave the strongest binding and had generally good solution properties. The binding affinities to the classical AATT sequence were similar to that for AAAGTTT for these extended compounds, but binding was weaker to the AAAGCTTT sequence with two intervening GC base pairs. Binding to both AAAGTTT and AATT was enthalpy driven for strong binding benzimidazole derivatives.

Minor Groove Binding Compounds That Jump a GC Base Pair and Bind to Adjacent AT Base Pair Sites †

Minor Groove Binding Compounds That Jump a GC Base Pair and Bind to Adjacent AT Base Pair Sites †

An unnatural base pair system for efficient PCR amplification and functionalization of DNA molecules

Toward the expansion of the genetic alphabet, we present an unnatural base pair system for efficient PCR amplification, enabling the site-specific incorporation of extra functional components into DNA. This system can be applied to conventional PCR protocols employing DNA templates containing unnatural bases, natural and unnatural base triphosphates, and a 3′→5′ exonuclease-proficient DNA polymerase. For highly faithful and efficient PCR amplification involving the unnatural base pairing, we identified the natural-base sequences surrounding the unnatural bases in DNA templates by an in vitro selection technique, using a DNA library containing the unnatural base. The system facilitates the site-specific incorporation of a variety of modified unnatural bases, linked with functional groups of interest, into amplified DNA. DNA fragments (0.15 amol) containing the unnatural base pair can be amplified 107-fold by 30 cycles of PCR, with <1% total mutation rate of the unnatural base pair site. Using the system, we demonstrated efficient PCR amplification and functionalization of DNA fragments for the extremely sensitive detection of zeptomol-scale target DNA molecules from mixtures with excess amounts (pmol scale) of foreign DNA species. This unnatural base pair system will be applicable to a wide range of DNA/RNA-based technologies.

Solution Structure of a DNA Duplex Containing a Guanine−Difluorotoluene Pair: A Wobble Pair without Hydrogen Bonding?

The incorporation of synthetic nucleoside analogues into DNA duplexes provides a unique opportunity to probe both structure and function of nucleic acids. We used 1H and 19F NMR and molecular dynamics calculations to determine the solution structures of two similar DNA decamer duplexes, one containing a central G-T mismatched or "wobble" base pair, and one in which the thymine in this base pair is replaced by difluorotoluene (a thymine isostere) creating a G-F pair. Here, we show that the non-hydrogen-bonding G-F pair stacks relatively well into the helix and that the distortions caused by each non-Watson-Crick G-T or G-F base pair are quite localized to a three base pair site around the mismatch. A detailed structural analysis reveals that the absence of hydrogen bonding introduces more dynamic motion into the G-F pair relative to G-T and permits the G-F pair to exhibit stacking and conformational features characteristic of both a Watson-Crick base pair (on the guanine containing strand) and a wobble base pair (on the strand containing the difluorotoluene). We used these results to posit a rationale for recognition and repair of mismatch sites in DNA.

The Regulatory Element in the 3′-Untranslated Region of Human Papillomavirus 16 Inhibits Expression by Binding CUG-binding Protein 1

The 3'-untranslated regions (UTRs) of human papillomavirus 16 (HPV16) and bovine papillomavirus 1 (BPV1) contain a negative regulatory element (NRE) that inhibits viral late gene expression. The BPV1 NRE consists of a single 9-nucleotide (nt) U1 small nuclear ribonucleoprotein (snRNP) base pairing site (herein called a U1 binding site) that via U1 snRNP binding leads to inhibition of the late poly(A) site. The 79-nt HPV16 NRE is far more complicated, consisting of 4 overlapping very weak U1 binding sites followed by a poorly understood GU-rich element (GRE). We undertook a molecular dissection of the HPV16 GRE and identify via UV cross-linking, RNA affinity chromatography, and mass spectrometry that is bound by the CUG-binding protein 1 (CUGBP1). Reporter assays coupled with knocking down CUGBP1 levels by small interfering RNA and Dox-regulated shRNA, demonstrate CUGBP1 is inhibitory in vivo. CUGBP1 is the first GRE-binding protein to have RNA interfering knockdown evidence in support of its role in vivo. Several fine-scale GRE mutations that inactivate GRE activity in vivo and GRE binding to CUGBP1 in vitro are identified. The CUGBP1.GRE complex has no activity on its own but specifically synergizes with weak U1 binding sites to inhibit expression in vivo. No synergy is seen if the U1 binding sites are made weaker by a 1-nt down-mutation or made stronger by a 1-nt up-mutation, underscoring that the GRE operates only on weak sites. Interestingly, inhibition occurs at multiple levels, in particular at the level of poly(A) site activity, nuclear-cytoplasmic export, and translation of the mRNA. Implications for understanding the HPV16 life cycle are discussed.

Recognition of longer duplex DNA by cooperative strand invasion

Peptide nucleic acid is a synthetic DNA mimic in which the sugar-phosphate backbone has been replaced by a peptide backbone. A remarkable feature of PNA is its ability to recognize sequences within duplex DNA by strand invasion. We have previously demonstrated that a PNA targeting six bases within duplex DNA cooperatively binds to 12 base-pair site by strand invasion. We here report an successful extension of the target site size to 18 base pairs without the expense of specificity.