MECHANISMS OF QUENCHING OF THE FLUORESCENCE OF A BENZO[a]PYRENE TETRAOL METABOLITE MODEL COMPOUND BY 2'-DEOXYNUCLEOSIDES

Abstract— The hydrophobic interactions of bulky polycyclic aromatic hydrocarbons with nucleic acid bases and the formation of noncovalent complexes with DNA are important in the expressions of the mutagenic and carcinogenic potentials of this class of compounds. The fluorescence of the polycyclic aromatic residues can be employed as a probe of these interactions. In this work, the interactions of the (+)-trans stereoisomer of the tetraol 7,8,9,10-tetrahydroxytetrahydrobenzo[a]pyrene (BPT), a hydrolysis product of a highly mutagenic and carcinogenic diol epoxide derivative of benzo[a]pyrene, were studied with 2′-deoxynucleosides in aqueous solution by fluorescence and UV spectroscopic techniques. Ground-state complexes between BPT and the purine derivatives 2′-deoxyguanosine (dG), 2′-deoxyadenosine (dA), and 2′-deoxyinosine (dI) are formed with association constants in the range of ～40–130 M^?1 Complex formation with the pyrimidine derivatives 2′-deoxythymidine (dT), 2′-deoxycytidine (dC), and 2′-deoxyuridine (dU) is significantly weaker. Whereas dG is a strong quencher of the fluorescence of BPT by both static and dynamic mechanisms (dynamic quenching rate constant k_dyn= [2.5 ± 0.41 × 10⁹M¹ s ¹, which is close to the estimated diffusion-controlled value of ～ 5 × 10⁹M^{? 1} s^?1), both dA and dI are weak quenchers and form fluorescenceemitting complexes with BPT. The pyrimidine derivatives dC, dU, and dT are efficient dynamic fluorescence quenchers (K_dyn～ [1.5–3.0] × 10⁹M^?1 s^?1), with a small static quenching component due to complex formation evident only in the case of dT. None of the four nucleosidcs dG, dA, dC and dT are dynamic quenchers of BPT in the triplet excited state; the observed lower yields of triplets are attributed to the quenching of single excited states of BPT by 2′-deoxynucleosides without passing through the triplet manifold of BPT. Possible fluorescence quenching mechanisms involving photoinduced electron transfer are discussed. The strong quenching of the fluorescence of BPT by dG, dC and dT accounts for the low fluorescence yields of BPT-native DNA and of pyrene-DNA complexes.     

Site‐Directed Spin‐Labeling of DNA by the Azide–Alkyne ‘Click’ Reaction: Nanometer Distance Measurements on 7‐Deaza‐2′‐deoxyadenosine and 2′‐Deoxyuridine Nitroxide Conjugates Spatially Separated or Linked to a ‘dA‐dT’ Base Pair

Nucleobase‐directed spin‐labeling by the azide‐alkyne ‘click’ (CuAAC) reaction has been performed for the first time with oligonucleotides. 7‐Deaza‐7‐ethynyl‐2′‐deoxyadenosine ( 1 ) and 5‐ethynyl‐2′‐deoxyuridine ( 2 ) were chosen to incorporate terminal triple bonds into DNA. Oligonucleotides containing 1 or 2 were synthesized on a solid phase and spin labeling with 4‐azido‐2,2,6,6‐tetramethylpiperidine 1‐oxyl (4‐azido‐TEMPO, 3 ) was performed by post‐modification in solution. Two spin labels ( 3 ) were incorporated with high efficiency into the DNA duplex at spatially separated positions or into a ‘dA‐dT’ base pair. Modification at the 5‐position of the pyrimidine base or at the 7‐position of the 7‐deazapurine residue gave steric freedom to the spin label in the major groove of duplex DNA. By applying cw and pulse EPR spectroscopy, very accurate distances between spin labels, within the range of 1–2 nm, were measured. The spin–spin distance was 1.8±0.2 nm for DNA duplex 17 ( dA*^7,10 ) ?11 containing two spin labels that are separated by two nucleotides within one individual strand. A distance of 1.4±0.2 nm was found for the spin‐labeled ‘dA‐dT’ base pair 15 ( dA*⁷ ) ?16 ( dT*⁶ ). The ‘click’ approach has the potential to be applied to all four constituents of DNA, which indicates the universal applicability of the method. New insights into the structural changes of canonical or modified DNA are expected to provide additional information on novel DNA structures, protein interaction, DNA architecture, and synthetic biology.     

7‐Halogenated 7‐Deaza‐2′‐deoxyxanthine 2′‐Deoxyribonucleosides

The synthesis of the 7‐halogenated derivatives 1b (7‐bromo) and 1c (7‐iodo) of 7‐deaza‐2′‐deoxyxanthosine ( 1a ) is described. A partial Br→I exchange was observed when the demethylation of 6‐methoxy precursor compound 4b was performed with Me₃SiCl/NaI. This reaction is circumvented by the nucleophilic displacement of the MeO group under strong alkaline conditions. The halogenated 7‐deaza‐2′‐deoxyxanthosine derivatives 1b , c show a decreased S‐conformer population of the sugar moiety compared to the nonhalogenated 1a . They are expected to form stronger triplexes when they replace 1a in the 1 ?dA?dT base triplet.     

Sequence-specific control of azobenzene assemblies by molecular recognition of DNA

We investigated the molecular recognition between the amphiphile AzoAde, which is composed of azobenzene in the hydrophobic and adenine in the hydrophilic portion of the molecule, and oligonucleotides having a homogeneous base (dA30, dT30, dG30, and dC30) at the air-water interface. On the basis of the complementary base-pairing of DNA in the duplex, orderly arrangement of AzoAde on templated dT30 was examined using pi-A isotherm, UV-vis RAS, FT-IR RAS, and XPS measurements. Although there was little interaction between AzoAde and mismatched oligonucleotides (dA30, dG30, and dC30), AzoAde prepared on a dT30 subphase stoichiometrically assembled and interacted with dT30, subsequently forming a J-form assembly at the air-water interface. AFM observation of the LB films revealed the nanostructure of the J-formed AzoAde monolayer on the dT30 subphase as well as the domain structures of the H-formed monolayers on the other oligonucleotide subphases. Therefore, dT30 has a potential application as a template for assembling AzoAde at the air-water interface.     

Base pairing and replicative processing of the formamidopyrimidine-dG DNA lesion

The 2,6-diamino-4-hydroxy-5-formamidopyrimidine of 2'-deoxyguanosine (FaPydG) is one of the major DNA lesions found after oxidative stress in cells. To clarify the base pairing and coding potential of this major DNA lesion with the aim to estimate its mutagenic effect, we prepared oligonucleotides containing a cyclopentane based analogue of the DNA lesion (cFaPydG). In addition, oligonucleotides containing the cyclopentane analogue of 2'-deoxyguanosine (cdG), and oligonucleotides containing 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) were synthesized. The thermodynamic stability of duplexes containing these building blocks and all canonical counterbases were determined by concentration dependent melting-point measurements (van't Hoff plots). The data reveal that cFaPydG greatly destabilizes a DNA duplex (DeltaDeltaG degrees (298K) approximately 2-4 kcal mol(-1)). The optimal base pairing partner for the cFaPydG lesion is dC. Investigation of duplexes containing dG and cdG shows that the effect of substituting the deoxyribose by a cyclopentane moiety is marginal. The data also provide strong evidence that the FaPydG lesion is unable to form a base pair with dA. Our computational studies indicate that the syn-conformation required for base pairing with dA is energetically unfavorable. This is in contrast to 8-oxodG for which the syn-conformation represents the energetic minimum. Kinetic primer extension studies using S. cerevisiae Pol eta reveal that cFaPydG is replicated in an error-free fashion. dC is inserted 2-3 orders of magnitude more efficiently than dT or dA, showing that FaPydG is a lesion which retains the coding potential of dG. This is also in contrast to 8-oxodG, for which base pairing with dC and dA was established.     

Naphthalene Imide Conjugates: Formation of Supramolecular Assemblies,and the Encapsulation and Release of Dyes through DNA‐Mediated Disassembly

We report the synthesis of two new amphiphilic conjugates 1 and 2 based on naphthalene di‐ and monoimide chromophores and the investigation of their photophysical, self‐assembly and DNA‐binding properties. These conjugates showed aqueous good solubility and exhibited strong interactions with DNA and polynucleotides such as poly(dG?dC)–poly(dG?dC) and poly(dA?dT)–poly(dA?dT). The interaction of these conjugates with DNA was evaluated by photo‐ and biophysical techniques. These studies revealed that the conjugates interact with DNA through intercalation with association constants in the order of 5–8×10⁴ M ^?1. Of these two conjugates, bolaamphiphile 1 exhibited a supramolecular assembly that formed vesicles with an approximate diameter of 220 nm in the aqueous medium at a critical aggregation concentration of 0.4 mM , which was confirmed by SEM and TEM. These vesicular structures showed a strong affinity for hydrophobic molecules such as Nile red through encapsulation. Uniquely, when exposed to DNA the vesicles disassembled, and therefore this transformation could be utilised for the encapsulation and release of hydrophobic molecules by employing DNA as a stimulus.     

Silver Ions in Non‐canonical DNA Base Pairs: Metal‐Mediated Mismatch Stabilization of 2′‐Deoxyadenosine and 7‐Deazapurine Derivatives with 2′‐Deoxycytidine and 2′‐Deoxyguanosine

Novel silver‐mediated dA?dC, dA*?dC, and dA*?dG base pairs were formed in a natural DNA double helix environment (dA* denotes 7‐deaza‐dA, 7‐deaza‐7‐iodo‐dA, and 7‐cyclopropyl‐7‐deaza‐dA). 7‐Deazapurine nucleosides enforce silver ion binding and direct metal‐mediated base pair formation to their Watson–Crick face. New phosphoramidites were prepared from 7‐deaza‐dA, 7‐deaza‐7‐iodo‐dA, and 7‐cyclopropyl‐7‐deaza‐dA, which contain labile isobutyryl protecting groups. Solid‐phase synthesis furnished oligonucleotides that contain mismatches in near central positions. Increased thermal stabilities (higher T_m values) were observed for oligonucleotide duplexes with non‐canonical dA*?dC and dA?dC pairs in the presence of silver ions. The stability of the silver‐mediated base pairs was pH dependent. Silver ion binding was not observed for the dA?dG mismatch but took place when mismatches were formed between 7‐deazaadenine and guanine. The specific binding of silver ions was confirmed by stoichiometric UV titration experiments, which proved that one silver ion is captured by one mismatch. The stability increase of canonical DNA mismatches might have an impact on cellular DNA repair.     

New nucleotide pairs for stable DNA triplexes stabilized by stacking interaction

New nucleotide pairs applicable to formation of DNA triplexes were developed. We designed oligonucleotides incorporating 5-aryl deoxycytidine derivatives (dC5Ars) and cyclic deoxycytidine derivatives, dCPPP and dCPPI, having an expanded aromatic area, as the second strand. As pairing partners, two types of abasic residues (C3: propylene linker, phi: abasic base) were chosen. It was concluded that, when the 5-aryl-modified cytosine bases paired with the abasic sites in TFOs in a space-fitting manner, the stability of the resulting triplexes significantly increased. The recognition of C3 toward dC5Ars was selective because of the stacking interactions between their aromatic part and the nucleobases flanking the abasic site. These results indicate the potential utility of new nucleotide triplets for DNA triplex formation, which might expand the variety of structures and sequences and might be useful for biorelated fields such as DNA nanotechnologies.     

四甲基吡啶卟啉与核酸相互作用的稳态和时间分辨光谱

采用稳态吸收和荧光光谱、圆二色谱和皮秒时间分辨荧光光谱手段, 研究了5,10,15,20-四[4-(N-甲基吡啶)]卟啉(TMPyP4)与腺嘌呤(A)、鸟嘌呤(G)、胸腺嘧啶(T)和胞嘧啶(C)等4种碱基, 以及相应的核苷、核苷酸和单链DNA的结合能力和光谱学性质. 研究结果发现, 嘌呤与TMPyP4的结合能力比嘧啶的强. 对于某一碱基系列, 结合能力强弱顺序依次为: 碱基~核苷<核苷酸<单链DNA. 时间分辨荧光谱研究发现, 除鸟嘌呤外, 核酸和TMPyP4复合物的荧光动力学均含有快(1~2 ns)和慢(约10 ns)两个衰减过程, 它们分别是由激基复合体和环境极性对激发态TMPyP4分子的影响所致. 单链DNA能诱导TMPyP4产生诱导圆二色信号, 而单分子(碱基、核苷、核苷酸)则无此功能.     

Photoswitchable Formation of a DNA Interstrand Cross‐Link by a Coumarin‐Modified Nucleotide

A coumarin‐modified pyrimidine nucleoside ( 1 ) has been synthesized using a Cu^I‐catalyzed click reaction and incorporated into oligodeoxynucleotides (ODNs). Interstrand cross‐links are produced upon irradiation of ODNs containing 1 at 350 nm. Cross‐linking occurs through a [2+2] cycloaddition reaction with the opposing thymidine, 2′‐deoxycytidine, or 2′‐deoxyadenosine. A much higher reactivity was observed with dT than dC or dA. Irradiation of the dT‐ 1 and dC‐ 1 cross‐linked products at 254 nm leads to a reversible ring‐opening reaction, while such phenomena were not observed with dA‐ 1 adducts. The reversible reaction is ultrafast and complete within 50–90 s. Consistent photoswitching behavior was observed over 6 cycles of irradiation at 350 nm and 254 nm. To the best of our knowledge, this is the first example of photoswitchable interstrand cross‐linking formation induced by a modified pyrimidine nucleoside.     

Conformationally restricted triplex-forming oligonucleotides (TFOs). Binding properties of α-l-LNA and introduction of the N-glycosylated LNA-guanosine

The method for scaled-up production of α-l-LNA phosphoramidite building blocks containing thymine and 5-methylcytosine nucleobases is described. Binding properties of pyrimidine TFOs modified with α-l-LNA are reported. In contrast to LNA TFOs, the fully modified α-l-LNA forms a stable triplex with a model DNA duplex. Pyrimidine DNA/LNA/α-l-LNA chimeras also efficiently hybridize with a model DNA duplex in the parallel mode. LNA nucleoside containing unnatural N⁷-glycosylated guanine (LNA-⁷G) was synthesized by a convergent method and incorporated into LNA oligonucleotides. The triplex-forming alternating DNA/LNA oligonucleotides containing a single LNA-⁷G modification instead of internal LNA-mC demonstrate improved pH-dependent properties. The single LNA-⁷G modification can also discriminatively reduce competitive binding of TFOs to natural nucleic acids in the antiparallel duplex mode.     

Stability of Hoogsteen‐Type Triplexes – Electrostatic Attraction between Duplex Backbone and Triplex‐Forming Oligonucleotide (TFO) Using an Intercalating Conjugate

Syntheses are described for two novel twisted intercalating nucleic acid (TINA) monomers where the intercalator comprises a benzene ring linked to a naphthalimide moiety via an ethynediyl bridge. The intercalators Y and Z have a 2‐(dimethylamino)ethyl and a methyl residue on the naphthalimide moiety, respectively. When used as triplex‐forming oligonucleotides (TFOs), the novel naphthalimide TINAs show extraordinary high thermal stability in Hoogsteen‐type triplexes and duplexes with high discrimination of mismatch strands. DNA Strands containing the intercalator Y show higher thermal triplex stability than DNA strands containing the intercalator Z . This observation can be explained by the ionic interaction of the protonated dimethylamino group under physiological conditions, targeting the negatively charged phosphate backbone of the duplex. This interaction leads to an extra binding mode between the TFO and the duplex, in agreement with molecular‐modeling studies. We believe that this is the first example of an intercalator linking the TFO to the phosphate backbone of the duplex by an ionic interaction, which is a promising tool to achieve a higher triplex stability.     

Interstrand cross-link formation in duplex and triplex DNA by modified pyrimidines

DNA interstrand cross-links have important biological consequences and are useful biotechnology tools. Phenylselenyl substituted derivatives of thymidine (1) and 5-methyl-2'-deoxycytidine (5) produce interstrand cross-links in duplex DNA when oxidized by NaIO4. The mechanism involves a [2,3]-sigmatropic rearrangement of the respective selenoxides to the corresponding methide type intermediates, which ultimately produce the interstrand cross-links. Determination of the rate constants for the selenoxide rearrangements indicates that the rate-determining step for cross-linking is after methide formation. Cross-linking by the thymidine derivative in duplex DNA shows a modest kinetic preference when flanked by pyrimidines as opposed to purines. In contrast, the rate constant for cross-link formation from 5 opposite dG in duplex DNA is strongly dependent upon the flanking sequence and, in general, is at least an order of magnitude slower than that for 1 in an otherwise identical sequence. Introduction of mispairs at the base pairs flanking 5 or substitution of the opposing dG by dI significantly increases the rate constant and yield for cross-linking, indicating that stronger hydrogen bonding between the methide derived from it and dG compared to dA and the respective electrophile derived from 1 limits reaction by increasing the barrier to rotation into the required syn-conformation. Incorporation of 1 or 5 in triplex forming oligonucleotides (TFOs) that utilize Hoogsteen base pairing also yields interstrand cross-links. The dC derivative produces ICLs approximately 10x faster than the thymidine derivative when incorporated at the 5'-termini of the TFOs and higher yields when incorporated at internal sites. The slower, less efficient ICL formation emanating from 1 is attributed to reaction at N1-dA, which requires local melting of the duplex. In contrast, 5 produces cross-links by reacting with N7-dG. The cross-linking reactions of 1 and 5 illustrate the versatility and utility of these molecules as mechanistic probes and tools for biotechnology.     

Nucleosides and Oligonucleotides with Diynyl Side Chains: Base Pairing and Functionalization of 2′‐Deoxyuridine Derivatives by the Copper(I)‐Catalyzed Alkyne?Azide ‘Click’ Cycloaddition

Oligonucleotides containing the 5‐substituted 2′‐deoxyuridines 1b or 1d bearing side chains with terminal C?C bonds are described, and their duplex stability is compared with oligonucleotides containing the 5‐alkynyl compounds 1a or 1c with only one nonterminal C?C bond in the side chain. For this, 5‐iodo‐2′‐deoxyuridine ( 3 ) and diynes or alkynes were employed as starting materials in the Sonogashira cross‐coupling reaction (Scheme 1). Phosphoramidites 2b – d were prepared (Scheme 3) and used as building blocks in solid‐phase synthesis. T_m Measurements demonstrated that DNA duplexes containing the octa‐1,7‐diynyl side chain or a diprop‐2‐ynyl ether residue, i.e., containing 1b or 1d , are more stable than those containing only one triple bond, i.e., 1a or 1c (Table 3). The diyne‐modified nucleosides were employed in further functionalization reactions by using the protocol of the Cu^I‐catalyzed Huisgen–Meldal–Sharpless [2+3] cycloaddition (‘click chemistry’) (Scheme 2). An aliphatic azide, i. e., 3′‐azido‐3′‐deoxythymidine (AZT; 4 ), as well as the aromatic azido compound 5 were linked to the terminal alkyne group resulting in 1H‐1,2,3‐triazole‐modified derivatives 6 and 7 , respectively (Scheme 2), of which 6 forms a stable duplex DNA (Table 3). The Husigen–Meldal–Sharpless cycloaddition was also performed with oligonucleotides (Schemes 4 and 5).     

Synthesis of Nucleobase‐Functionalized Carbon Nanotubes and Their Hybridization with Single‐Stranded DNA

For the first time ssDNA (25‐aptamer of mixed dA, dT, dG, and dC) was wrapped around functionalized single‐walled carbon nanotubes (SWCNTs), whose external surfaces were attached to multiple triazole‐(ethylene glycol)‐dA ligands. This method of hybridization involved the formation of hydrogen bonds between dT of ssDNA and dA of functionalized SWCNTs. It deviates from the reported π–π stacking between the nucleobases of DNA and the external sidewalls of nanotubes. The structural properties of the functionalized SWCNTs and its ssDNA complex were characterized by spectroscopic (including CD and Raman), thermogravimetric, and microscopic (TEM) methods. The results thus obtained establish a new platform of DNA delivery by use of nanotubes as a new vehicle with great potential in biomedical applications and drug development.     

Stabilization of a Bimolecular Triplex by 3′‐S‐Phosphorothiolate Modifications: An NMR and UV Thermal Melting Investigation

Triplexes formed from oligonucleic acids are key to a number of biological processes. They have attracted attention as molecular biology tools and as a result of their relevance in novel therapeutic strategies. The recognition properties of single‐stranded nucleic acids are also relevant in third‐strand binding. Thus, there has been considerable activity in generating such moieties, referred to as triplex forming oligonucleotides (TFOs). Triplexes, composed of Watson–Crick (W–C) base‐paired DNA duplexes and a Hoogsteen base‐paired RNA strand, are reported to be more thermodynamically stable than those in which the third strand is DNA. Consequently, synthetic efforts have been focused on developing TFOs with RNA‐like structural properties. Here, the structural and stability studies of such a TFO, composed of deoxynucleic acids, but with 3′‐S‐phosphorothiolate (3′‐SP) linkages at two sites is described. The modification results in an increase in triplex melting temperature as determined by UV absorption measurements. ¹H NMR analysis and structure generation for the (hairpin) duplex component and the native and modified triplexes revealed that the double helix is not significantly altered by the major groove binding of either TFO. However, the triplex involving the 3′‐SP modifications is more compact. The 3′‐SP modification was previously shown to stabilise G‐quadruplex and i‐motif structures and therefore is now proposed as a generic solution to stabilising multi‐stranded DNA structures.     

8-Aza-7-deazaadenine N8- and N8-(β-D-2′-Deoxyribofuranosides): Building blocks for automated DNA synthesis and properties of oligodeoxyribonucleotides

Oligonucleotides with alternating 8-aza-7-deaza-2′-deoxyadenosine (= c⁷z⁸A_d2) and dT residues (see 11, 14 and 16 ) or 4-aminopyrazolo [3,4-d] pyrimidine N²-(β-D -2′-deoxyribofuranoside) (= c⁷z⁸A′_d¹); ( 3 ) and dT residues (see 12 ) have been prepared by solid-phase synthesis using P(III) chemistry, Additionally, palindromic oligomers derived from d(C-T-G-G-A-T-C-C-A-G) but containing 2 or 3 instead of dA (see 18 – 22 ) have been synthesized. Benzoylation of 2 or 3 , followed by 4,4′-dimethoxytritylation and subsequent phosphitylation yielded the methyl or the cyanoethyl phosphoramidites 8a,b and 9 . They were employed in automated. DNA synthesis. Alternating oligomers containing 2 or 3 showed increase dT_m values compared to those with dA, in particular 12 with an unusual N²-glycosylic bond. The palindromic oligomers 18 - 22 containing 2 or 3 instead of dA outside of the enzymic recognition side reduced the hydrolysis rate, replacement within d(G-A-T-C) abolished phosphodiester hydrolysis.     

Oligonucleotides Containing 7‐Deaza‐2′‐deoxyinosine as Universal Nucleoside: Synthesis of 7‐Halogenated and 7‐Alkynylated Derivatives,Ambiguous Base Pairing,and Dye Functionalization by the Alkyne–Azide ‘Click’ Reaction

Oligonucleotides containing 7‐deaza‐2′‐deoxyinosine derivatives bearing 7‐halogen substituents or 7‐alkynyl groups were prepared. For this, the phosphoramidites 2b – 2g containing 7‐substituted 7‐deaza‐2′‐deoxyinosine analogues 1b – 1g were synthesized (Scheme 2). Hybridization experiments with modified oligonucleotides demonstrate that all 2′‐deoxyinosine derivatives show ambiguous base pairing, as 2′‐deoxyinosine does. The duplex stability decreases in the order C_d>A_d>T_d>G_d when 2b – 2g pair with these canonical nucleosides (Table 6). The self‐complementary duplexes 5′‐d(F⁷c⁷I‐C)₆, d(Br⁷c⁷I‐C)₆, and d(I⁷c⁷I‐C)₆ are more stable than the parent duplex d(c⁷I‐C)₆ (Table 7). An oligonucleotide containing the octa‐1,7‐diyn‐1‐yl derivative 1g , i.e., 27 , was functionalized with the nonfluorescent 3‐azido‐7‐hydroxycoumarin ( 28 ) by the Huisgen–Sharpless–Meldal cycloaddition ‘click’ reaction to afford the highly fluorescent oligonucleotide conjugate 29 (Scheme 3). Consequently, oligonucleotides incorporating the derivative 1g bearing a terminal C?C bond show a number of favorable properties: i) it is possible to activate them by labeling with reporter molecules employing the ‘click’ chemistry. ii) Space demanding residues introduced in the 7‐position of the 7‐deazapurine base does not interfere with duplex structure and stability (Table 8). iii) The ambiguous pairing character of the nucleobase makes them universal probes for numerous applications in oligonucleotide chemistry, molecular biology, and nanobiotechnology.     

Synthesis of Oligonucleotides Containing the N6-(2-Deoxy-α,β-d-erythropentofuranosyl)-2,6-diamino-4-hydroxy-5-formamidopyrimidine (Fapy⋅dG) Oxidative Damage Product Derived from 2′-Deoxyguanosine

N⁶-(2-Deoxy-α,β-d -erythropentofuranosyl)-2,6-diamino-4-hydroxy-5-formamidopyrimidine (Fapy⋅dG) is a major DNA lesion produced from 2′-deoxyguanosine under oxidizing conditions. Fapy ⋅ dG is produced from a common intermediate that leads to 7,8-dihydro-8-oxo-2′-deoxyguanosine (8-OxodGuo), and in greater quantities in cells. The impact of Fapy ⋅ dG on DNA structure and function is much less well understood than that of 8-OxodGuo. This is largely due to the significantly greater difficulty in synthesizing oligonucleotides containing Fapy ⋅ dG than 8-OxodGuo. We describe a synthetic approach for preparing oligonucleotides containing Fapy ⋅ dG that will facilitate intensive studies of this lesion in DNA. A variety of oligonucleotides as long as 30 nucleotides are synthesized. We anticipate that the chemistry described herein will provide an impetus for a wide range of studies involving Fapy ⋅ dG.     

Regioselective arylation of 2′-deoxyribonucleosides on amido or imino sites by copper(II)-mediated direct coupling with arylboronic acids

N¹-Aryl derivatives of 2′-deoxyguanosine (dG) were synthesized by copper(II)-mediated coupling of dG with arylboronic acids. Analogous aryl derivatives of 2′-deoxyinosine (dIn), 2′-deoxyuridine (dU), thymidine (T), 2′-deoxyadenosine (dA), and 2′-deoxycytidine (dC) were also conveniently synthesized by this method. Arylation took place preferentially on the amido functions in dG and dIn and the imino functions in dU or T. Remarkably, the nucleosides themselves served as internal ligands as well as reactants.