6-Pyrazolylpurine and its deaza derivatives as nucleobases for silver(I)-mediated base pairing with pyrimidines.

The artificial nucleobase 6-pyrazolylpurine (6PP) and its deaza derivatives 1-deaza-6-pyrazolylpurine (1D6PP), 7-deaza-6-pyrazolylpurine (7D6PP), and 1,7-dideaza-6-pyrazolylpurine (1,7D6PP) were investigated with respect to their ability to differentiate between the canonical nucleobases cytosine and thymine by means of silver(I)-mediated base pairing. As shown by temperature-dependent UV spectroscopy and by circular dichroism spectroscopy, 6PP and (to a lesser extent) 7D6PP form stable silver(I)-mediated base pairs with cytosine, but not with thymine. 1D6PP and 1,7D6PP do not engage in the formation of stabilizing silver(I)-mediated base pairs with cytosine or thymine. The different behavior of 1D6PP, 7D6PP, and 1,7D6PP indicates that silver(I) binding occurs via the N1 position of the purine derivative, i.e. via the Watson-Crick face. The data show that 6PP is capable of differentiating between cytosine and thymine, which is potentially relevant in the context of detecting single-nucleotide polymorphisms.

Chiral-at-Metal Silver-Mediated Base Pairs: Metal-Centred Chirality versus DNA Helical Chirality.

When covalently incorporating ligands capable of forming chiral metal complexes into a DNA oligonucleotide duplex, an enantiospecific formation of metal-mediated base pairs is possible. We have been investigating the chirality of the silver-mediated base pair P-AgI -P (P, 1H-imidazo[4,5-f][1,10]phenanthroline) depending on the number of consecutive P : P pairs within a series of duplexes. Towards this end, both enantiomers of the nucleoside analogue 3-(1H-imidazo[4,5-f][1,10]phenanthrolin-1-yl)propane-1,2-diol comprising an acyclic backbone were introduced into DNA duplexes, resulting in diastereomeric metal-mediated base pairs. The same chiral-at-metal complex is formed inside the duplex for up to five neighbouring P-AgI -P pairs, irrespective of whether (S)-P or (R)-P is used. With six silver-mediated base pairs, the chirality of the metal complex is inverted for (S)-P but not for (R)-P. This indicates an intricate balance of what determines the configuration of the metal complex, the intrinsically preferred metal-centred chirality or the DNA helical chirality.

Single-Stranded DNA as Supramolecular Template for One-Dimensional Palladium(II) Arrays.

Atomic-level control over the position and growth of a single and continuous metal chain is an ambitious goal that often requires complex and costly processes. Herein, we demonstrate that 1Pd-DNA molecules, comprising a continuous single chain of PdII ions, can be prepared by a simple self-assembly reaction between the complex [Pd(Cheld)(CH3 CN)] (1Pd_CH3 CN) (Cheld=chelidamic acid) and single-stranded DNA homopolymers (ss-DNA) containing adenine (A) or 7-deazaadenine (X) bases. The single PdII -base pairs [1Pd(N1-A)] and [1Pd(N1-X)] were synthesized and characterized in solution and solid-state (X-ray diffraction) revealing an arrangement similar to that of natural Watson-Crick base pairs. Subsequently, 1Pd-DNA hybrids were prepared, characterized, and their structures studied by small-angle X-ray scattering (SAXS) and ab-initio calculations. The results indicate that the 1Pd-DNA structures resemble that of double-stranded DNA, with one strand being replaced by a supramolecular stack of continuous PdII complexes.

Stable Hg(II)-mediated base pairs with a phenanthroline-derived nucleobase surrogate in antiparallel-stranded DNA.

Metal-mediated base pairs involving artificial nucleobases have emerged as a promising means for the site-specific functionalization of nucleic acids with metal ions. In this context, a GNA-appended (GNA: glycol nucleic acid) nucleoside analogue containing the artificial nucleobase 1H-imidazo[4,5-f][1,10]phenanthroline (P) has already been applied successfully in a variety of homo- and heteroleptic metal-mediated base pairs, mainly involving Ag(I) ions. Herein, we report a thorough investigation of the Hg(II)-binding properties of P when incorporated into antiparallel-stranded DNA duplexes. The artificial nucleobase P is able to form Hg(II)-mediated homoleptic base pairs of the type P-Hg(II)-P with a [2 + 2] coordination environment. In addition, the heteroleptic P-Hg(II)-T pair was investigated. The addition of a stoichiometric amount of Hg(II) to a duplex comprising either a P:P pair or a P:T pair stabilizes the DNA duplex by 4.3 °C and 14.5 °C, respectively. The P-Hg(II)-T base pair, hence, represents the most stabilizing non-organometallic Hg(II)-mediated base pair reported to date. The formation of the Hg(II)-mediated base pairs was investigated by means of temperature-dependent UV spectroscopy and CD spectroscopy.

Silver(I) Coordination in Silver(I)-Mediated Homo Base Pairs of 6-Pyrazolylpurine in DNA Duplexes Involves the Watson-Crick Edge.

DNA duplexes comprising 6-(1H-pyrazol-1-yl)-9H-purine (6PP), 1-deaza-6PP (1D 6PP), 7-deaza-6PP (7D 6PP) and 1,7-dideaza-6PP (1,7D 6PP) 2'-deoxyribonucleosides, respectively, were investigated towards their ability to form metal-mediated base pairs in the presence of AgI . In 6PP and 7D 6PP, the AgI ion can coordinate to the nucleobase via the endocyclic N1 nitrogen atom, that is, via the Watson-Crick edge. In contrast, this nitrogen atom is not available in 1D 6PP and 1,7D 6PP, so that in 1D 6PP an AgI coordination is only possible via the Hoogsteen edge (N7). Reference duplexes with either adenine:adenine mispairs or canonical adenine:thymine base pairs were used to investigate the impact of the pyrazolyl moiety on the AgI -binding properties. To determine the thermal and structural duplex stabilities in the absence or presence of AgI , all duplexes were examined by UV and circular dichroism spectroscopic studies. These investigations shed light on the question of whether N1- or N7-coordination is preferred in purine-based metal-mediated base pairs.

Silver(I)-mediated base pairing in parallel-stranded DNA involving the luminescent cytosine analog 1,3-diaza-2-oxophenoxazine.

1,3-Diaza-2-oxophenoxazine (X) has been introduced as a ligand in silver(I)-mediated base pairing in a parallel DNA duplex. This fluorescent cytosine analog is capable of forming stabilizing X-Ag(I)-X and X-Ag(I)-C base pairs in DNA duplexes, as confirmed by temperature-dependent UV spectroscopy and luminescence spectroscopy. DFT calculations of the silver(I)-mediated base pairs suggest the presence of a synergistic hydrogen bond. Molecular dynamics (MD) simulations of entire DNA duplexes nicely underline the geometrical flexibility of these base pairs, with the synergistic hydrogen bond facing either the major or the minor groove. Upon silver(I) binding to the X:X or X:C base pairs, the luminescence emission maximum experiences a red shift from 448 to 460 nm upon excitation at 370 nm. Importantly, the luminescence of the 1,3-diaza-2-oxophenoxazine ligand is not quenched significantly upon binding a silver(I) ion. In fact, the luminescence intensity even increases upon formation of a X-Ag(I)-C base pair, which is expected to be beneficial for the development of biosensors. As a consequence, the silver(I)-mediated phenoxazinone base pairs represent the first strongly fluorescent metal-mediated base pairs.

Excess Electron Transfer through DNA Duplexes Comprising a Metal-Mediated Base Pair.

Excess electron transfer through one set of DNA duplexes comprising either one or two metal-mediated thymine-HgII -thymine base pairs was studied. Towards this end, the metal-mediated base pair(s) were introduced between an artificial nucleoside bearing a N,N,N',N'-tetramethyl-1,5-diaminonaphthalene derivative (dN, acting as a photoinducible electron donor) and 5-bromo-2'-deoxyuridine (dB, acting as an electron acceptor). Upon one-electron reduction, dB loses a bromide ion. The amount of unreacted dB remaining after irradiation-determined by LC/ICP-MS-was used to evaluate the charge-transfer efficiency across the metal-mediated base pair(s). Reference measurements with canonical adenine:thymine base pairs prove the applicability of this approach for the detection of charge transfer in DNA. The data indicate that, for the set of DNA duplexes under investigation, excess electron transfer across a thymine-HgII -thymine base pair proceeds with low efficiency, comparable to the transfer across a thymine:thymine mispair. Two contiguous thymine-HgII -thymine base pairs effectively shut down excess electron transfer.

Metal-Mediated Base Pairs: From Characterization to Application.

The investigation of metal-mediated base pairs and the development of their applications represent a prominent area of research at the border of bioinorganic chemistry and supramolecular coordination chemistry. In metal-mediated base pairs, the complementary nucleobases in a nucleic acid duplex are connected by coordinate bonds to an embedded metal ion rather than by hydrogen bonds. Because metal-mediated base pairs facilitate a site-specific introduction of metal-based functionality into nucleic acids, they are ideally suited for use in DNA nanotechnology. This minireview gives an overview of the general requirements that need to be considered when devising a new metal-mediated base pair, both from a conceptual and from an experimental point of view. In addition, it presents selected recent applications of metal-modified nucleic acids to indicate the scope of metal-mediated base pairing.

6-Pyrazolylpurine as an Artificial Nucleobase for Metal-Mediated Base Pairing in DNA Duplexes.

The artificial nucleobase 6-pyrazol-1-yl-purine (6PP) has been investigated with respect to its usability in metal-mediated base pairing. As was shown by temperature-dependent UV spectroscopy, 6PP may form weakly stabilizing 6PP-Ag(I)-6PP homo base pairs. Interestingly, 6PP can be used to selectively recognize a complementary pyrimidine nucleobase. The addition of Ag(I) to a DNA duplex comprising a central 6PP:C mispair (C = cytosine) leads to a slight destabilization of the duplex. In contrast, a stabilizing 6PP-Ag(I)-T base pair is formed with a complementary thymine (T) residue. It is interesting to note that 6PP is capable of differentiating between the pyrimidine moieties despite the fact that it is not as sterically crowded as 6-(3,5-dimethylpyrazol-1-yl)purine, an artificial nucleobase that had previously been suggested for the recognition of nucleic acid sequences via the formation of a metal-mediated base pair. Hence, the additional methyl groups of 6-(3,5-dimethylpyrazol-1-yl)purine may not be required for the specific recognition of the complementary nucleobase.

The Effects of Magnesium Ions on the Enzymatic Synthesis of Ligand-Bearing Artificial DNA by Template-Independent Polymerase.

A metal-mediated base pair, composed of two ligand-bearing nucleotides and a bridging metal ion, is one of the most promising components for developing DNA-based functional molecules. We have recently reported an enzymatic method to synthesize hydroxypyridone (H)-type ligand-bearing artificial DNA strands. Terminal deoxynucleotidyl transferase (TdT), a template-independent DNA polymerase, was found to oligomerize H nucleotides to afford ligand-bearing DNAs, which were subsequently hybridized through copper-mediated base pairing (H-Cu(II)-H). In this study, we investigated the effects of a metal cofactor, Mg(II) ion, on the TdT-catalyzed polymerization of H nucleotides. At a high Mg(II) concentration (10 mM), the reaction was halted after several H nucleotides were appended. In contrast, at lower Mg(II) concentrations, H nucleotides were further appended to the H-tailed product to afford longer ligand-bearing DNA strands. An electrophoresis mobility shift assay revealed that the binding affinity of TdT to the H-tailed DNAs depends on the Mg(II) concentration. In the presence of excess Mg(II) ions, TdT did not bind to the H-tailed strands; thus, further elongation was impeded. This is possibly because the interaction with Mg(II) ions caused folding of the H-tailed strands into unfavorable secondary structures. This finding provides an insight into the enzymatic synthesis of longer ligand-bearing DNA strands.

A Dinuclear Mercury(II)-Mediated Base Pair in DNA.

The first dinuclear metal-mediated base pair containing divalent metal ions has been prepared. A combination of the neutral bis(monodentate) purine derivative 1,N6 -ethenoadenine (ϵA), which preferentially binds two metal ions with a parallel alignment of the N-M bonds, and the canonical nucleobase thymine (T), which readily deprotonates in the presence of HgII and thereby partially compensates the charge accumulation due to the two closely spaced divalent metal ions, yields the dinuclear T-HgII2 -ϵA base pair. This metal-mediated base pair stabilizes the DNA oligonucleotide duplex as shown by an increase of 8 °C in its melting temperature. Formation of the base pair was demonstrated by temperature-dependent UV spectroscopy as well as by titration experiments monitored by UV and CD spectroscopy.

Bifacial Base-Pairing Behaviors of 5-Hydroxyuracil DNA Bases through Hydrogen Bonding and Metal Coordination.

A novel bifacial ligand-bearing nucleobase, 5-hydroxyuracil (U(OH) ), which forms both a hydrogen-bonded base pair (U(OH) -A) and a metal-mediated base pair (U(OH) -M-U(OH) ) has been developed. The U(OH) -M-U(OH) base pairs were quantitatively formed in the presence of lanthanide ions such as Gd(III) when U(OH) -U(OH) pairs were consecutively incorporated into DNA duplexes. This result established metal-assisted duplex stabilization as well as DNA-templated assembly of lanthanide ions. Notably, a duplex possessing U(OH) -A base pairs was destabilized by addition of Gd(III) ions. This observation suggests that the hybridization behaviors of the U(OH) -containing DNA strands are altered by metal complexation. Thus, the U(OH) nucleobase with a bifacial base-pairing property holds great promise as a component for metal-responsive DNA materials.

A highly stabilizing silver(I)-mediated base pair in parallel-stranded DNA.

The first parallel-stranded DNA duplex with Hoogsteen base pairing that readily incorporates an Ag(+) ion into an internal mispair to form a metal-mediated base pair has been created. Towards this end, the highly stabilizing 6 FP-Ag(+)-6 FP base pair comprising the artificial nucleobase 6-furylpurine (6 FP) was devised. A combination of temperature-dependent UV spectroscopy, CD spectroscopy, and DFT calculations was used to confirm the formation of this base pair. The nucleobase 6 FP is capable of forming metal-mediated base pairs both by the Watson-Crick edge (i.e. in regular antiparallel-stranded DNA) and by the Hoogsteen edge (i.e. in parallel-stranded DNA), depending on the oligonucleotide sequence and the experimental conditions. The 6 FP-Ag(+)-6 FP base pair within parallel-stranded DNA is the most strongly stabilizing Ag(+)-mediated base pair reported to date for any type of nucleic acid, with an increase in melting temperature of almost 15 °C upon the binding of one Ag(+) ion.

Uridine triphosphate hybrid catalyst for carbon­carbon bond formation reactions with enhanced enantioselectivity by mercury(II) ions.

DNA hybrid catalysts are constructed by embedding active metal species into the chiral scaffolds of DNA, which have been successfully applied to some important aqueous-phase enantioselective transformations. Owing to simple components and inherent chirality, nucleotide hybrid catalysts are emerging in response to soving the unclear locations of catalytic centers and the plausible catalytic mechanisms in DNA-based asymmetric catalysis. However, the tertiary structure of nucleotides lacks tunability, severely impeding further design of nucleotide hybrid catalysts for potential applications. To this end, a design strategy for tunable nucleotide hybrid catalysts is put forward by introducing metal-mediated base pairs. Herein, we found that the formation of uracil­mercury(II)-uracil (U-Hg2+-U) base pairs could enhance the enantioselectivity in uracil-containing nucleotide-based asymmetric reactions. Compared with uracil triphosphate (UTP) complexing with Cu2+ ions (UTP∙Cu2+), the presence of Hg2+ ions gave rise to an increased enantiomeric excess (ee) of 38 % in Diels-Alder reactions and 22 % ee in Michael reactions. The Hg2+-tuning behaviors of UTP hybrid catalyst have been demonstrated to largely depend on nucleotides, Hg2+ concentrations, metal cofactors, additives and reaction types. Based on ultraviolet-visible, circular dichroism and nuclear magnetic resonance spectroscopic techniques, the chiral enhancement of Hg2+-containing UTP hybrid catalyst is proved to largely depend on the formation of U-Hg2+-U base pairs and the plausible cross-linked structure of UTP-Hg2+-UTP/Cu2+ assembly. This work provides a tunable strategy based on the concept of metal-mediated base pairs, allowing further design of potent oligonucleotide-based catalysts for other enantioselective reactions.

Synthesis of a novel nucleotide unit containing cytosine analog bearing phosphodiester function and formation of silver(I)-mediated DNA duplex.

Novel modified cytosine analogs bearing phosphodiester/thiophosphodiester functionality were synthesized. The interactions between different metal ions and modified cytosine-cytosine base-pairs in DNA duplexes were investigated by UV-melting experiments. The thiophosphodiester modification binds to the Ag(I) ions strongly compared to the phosphodiester counterpart as examined in ESI-MS spectra as well.

Silver(I)-mediated base pairing involving an S-glycosidic GNA nucleoside analogue.

The 4S-Ag(I)-C base pair (4S, 3-((2-(methylthio)pyrimidin-4-yl)thio)propane-1,2-diol; C, deoxycytidine) represents the first metal-mediated base pair comprising an S-glycosidic nucleoside analogue. We report here the synthesis of the phosphoramidite suitable for the automated solid-phase synthesis of DNA oligonucleotides containing 4S and its silver(I)-binding ability. The DNA duplexes comprising a 4S:C mispair exhibit a large thermal stabilization upon the addition of one equivalent of silver ions, giving rise to the formation of the above-mentioned silver(I)-mediated base pair. By formally replacing the sulfur atom in the glycosidic bond by an oxygen atom, i.e., by applying 3-((2-(methylthio)pyrimidin-4-yl)oxy)propane-1,2-diol (4 O) as the artificial nucleoside analogue, the participation of this atom as a donor atom in silver(I)-mediated base pairing is shown to be neglectable.Supplemental data for this article is available online at.

Silver(I)-mediated base pairing in DNA involving the artificial nucleobase 7,8-dihydro-8-oxo-1,N6-ethenoadenine.

The artificial nucleobase 7,8-dihydro-8-oxo-1,N6-ethenoadenine (X) was investigated with respect to its ability to engage in Ag(I)-mediated base pairing in DNA. Spectroscopic data indicate the formation of dinuclear X-Ag(I)2-X homo base pairs and mononuclear X-Ag(I)-C base pairs (C, cytosine). Density functional theory calculations and molecular dynamics simulations indicate that the nucleobase changes from its lactam tautomeric form prior to the formation of the Ag(I)-mediated base pair to the lactim form after the incorporation of the Ag(I) ions. Fluorescence spectroscopy indicates that the two Ag(I) ions of the homo base pair are incorporated sequentially. Isothermal titration calorimetry confirms that the affinity of one of the Ag(I) ions is about tenfold higher than that of the other Ag(I) ion. The computational analysis by means of density functional theory confirms a much larger reaction energy for the incorporation of the first Ag(I) ion. The thermal stabilization upon the formation of the dinuclear Ag(I)-mediated homo base pair exceeds the one previously observed for the closely related nucleobase 1,N6-ethenoadenine by far, despite very similar structures. This additional stabilization may stem from the presence of water molecules engaged in hydrogen bonding with the additional oxygen atom of the artificial nucleobase X. The highly stabilizing Ag(I)-mediated base pair is a valuable addition to established dinuclear metal-mediated base pairs.

Effects of metal ions on thermal stabilities of DNA duplexes containing homo- and heterochiral mismatched base pairs: comparison of internal and terminal substitutions.

The effects of metal ions on the stabilities of duplexes containing a D-homochiral and heterochiral mismatched base pairs were studied. In some duplexes containing an internal mismatched base pair, significant stabilization by HgII and AgI ions was observed. While, in duplexes containing a terminal mismatched base pair, only the duplexes containing T-T and LT-T mispairs were significantly stabilized by HgII ions, and the stabilities of the duplexes containing T-T and LT-T mispairs exceeded those of the corresponding homochiral matched duplex. The results suggest that the formation of homo- and heterochiral T-HgII-T base pairs at duplex termini would be useful for the thermal and enzymatic stabilization of DNA-based nanodevice.

Metal-mediated base pairing in DNA involving the artificial nucleobase imidazole-4-carboxylate.

The use of imidazole-4-carboxylate (X) as an artificial nucleobase in metal-mediated base pairing is reported. Towards this end, the corresponding deoxyribonucleoside was synthesized and structurally characterized as its sodium salt (sodium 1,2-dideoxy-1-(4-carboxyimidazol-1-yl)-d-ribofuranose). The deoxyribonucleoside was incorporated into different DNA duplexes (parallel-stranded and antiparallel-stranded), and their Cu(II)- and Ag(I)-binding behavior was investigated. It was shown that both X-Cu(II)-X and X-Ag(I)-X base pairs can be formed, with the former being more stabilizing than the latter. The formation of an X-Cu(II)-X base pair is accompanied by an increase in the duplex melting temperature of approximately 20 °C for antiparallel-stranded duplexes and of 12 °C for the parallel-stranded duplex under investigation. Imidazole-4-carboxylate represents the first imidazole-based nucleoside for Cu(II)-mediated base pairing. Moreover, it is the smallest nucleoside known to form stable Cu(II)-mediated base pairs. Structures of the X-Cu(II)-X and X-Ag(I)-X base pairs are proposed, too, based on molecular structures obtained using the model nucleobase 1-benzyl-1H-imidazole-4-carboxylate.

A stable zinc(II)-mediated base pair in a parallel-stranded DNA duplex.

A stable zinc(II)-mediated base pair was formed within a parallel-stranded DNA duplex comprising a GNA (glycol nucleic acid) functionalized nucleoside analog containing the artificial nucleobase 1H-imidazo[4,5-f][1,10]phenanthroline (P). The formation of the metal-mediated base pair was confirmed by UV and CD spectroscopic analyses. In the Zn(II)-mediated homo base pairs of the type P-Zn(II)-P, the metal ion adopts a [2 + 2] coordination environment. CD spectroscopic data suggest that the chiral metal complex is formed enantiospecifically, likely induced by the helical chirality of the surrounding DNA duplex. The Zn(II)-mediated base pair stabilizes the DNA oligonucleotide duplex by 9 °C. This stable Zn(II)-mediated base pair within a parallel-stranded duplex extends the scope of site-specific functionalization of nucleic acids by virtue of metal-mediated base pairing.