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Prebiotic Photochemical Coproduction of Purine Ribo- And Deoxyribonucleosides
The hypothesis that life on Earth may have started with a heterogeneous nucleic acid genetic system including both RNA and DNA has attracted broad interest. The recent finding that two RNA subunits (cytidine, C, and uridine, U) and two DNA subunits (deoxyadenosine, dA, and deoxyinosine, dI) can be coproduced in the same reaction network, compatible with a consistent geological scenario, supports this theory. However, a prebiotically plausible synthesis of the missing units (purine ribonucleosides and pyrimidine deoxyribonucleosides) in a unified reaction network remains elusive. Herein, we disclose a strictly stereoselective and furanosyl-selective synthesis of purine ribonucleosides (adenosine, A, and inosine, I) and purine deoxynucleosides (dA and dI), alongside one another, via a key photochemical reaction of thioanhydroadenosine with sulfite in alkaline solution (pH 8-10). Mechanistic studies suggest an unexpected recombination of sulfite and nucleoside alkyl radicals underpins the formation of the ribo C2′-O bond. The coproduction of A, I, dA, and dI from a common intermediate, and under conditions likely to have prevailed in at least some primordial locales, is suggestive of the potential coexistence of RNA and DNA building blocks at the dawn of life.
Xu, Jianfeng,Green, Nicholas J.,Russell, David A.,Liu, Ziwei,Sutherland, John D.
supporting information
p. 14482 - 14486
(2021/09/18)

An enzymatic flow-based preparative route to vidarabine
The bi-enzymatic synthesis of the antiviral drug vidarabine (arabinosyladenine, ara-A), catalyzed by uridine phosphorylase from Clostridium perfringens (CpUP) and a purine nucleoside phosphorylase fromAeromonas hydrophila (AhPNP), was re-designed under continuous-flow conditions. Glyoxyl-agarose and EziGTM1 (Opal) were used as immobilization carriers for carrying out this preparative biotransformation. Upon setting-up reaction parameters (substrate concentration and molar ratio, temperature, pressure, residence time), 1 g of vidarabine was obtained in 55% isolated yield and >99% purity by simply running the flow reactor for 1 week and then collecting (by filtration) the nucleoside precipitated out of the exiting flow. Taking into account the substrate specificity of CpUP and AhPNP, the results obtained pave the way to the use of the CpUP/AhPNP-based bioreactor for the preparation of other purine nucleosides.
Annunziata, Francesca,Bavaro, Teodora,Calleri, Enrica,Conti, Paola,Pinto, Andrea,Previtali, Clelia,Rinaldi, Francesca,Speranza, Giovanna,Tamborini, Lucia,Terreni, Marco,Ubiali, Daniela
(2020/03/23)

RETRACTED ARTICLE: Convenient synthesis of pyrimidine 2′-deoxyribonucleoside monophosphates with important epigenetic marks at the 5-position
Methyl groups of thymine and 5-methylcytosine (5mC) bases in DNA undergo endogenous oxidation damage. Additionally, 5mC residues can be enzymatically deaminated or oxidized through either genetic alterations or the newly identified epigenetic reprogramming pathway. Several methods have been developed to measure the formation of modified DNA nucleobases including 32P-postlabeling. However, the postlabeling method is often limited by the absence of authentic chemical standards. The synthesis of monophosphate standards of nucleotide oxidation products is complicated by the presence of additional functional groups on the modified bases that require complex protection and deprotection strategies. Due to the emerging interest in the pyrimidine oxidation products, the corresponding protected 3′-phosphoramidites needed for solid-phase oligonucleotide synthesis have been reported, and several are commercially available. We report here an efficient synthesis of 3′-monophosphates from 3′-phosphoramidites and the subsequent enzymatic conversion of 3′-monophosphates to the corresponding 5′-monophosphates using commercially available enzymes. This journal is
Zheng, Song,Tran, Ai,Curry, Alyson M.,White, Dawanna S.,Cen, Yana
p. 5164 - 5173
(2020/07/23)

Thermodynamic Reaction Control of Nucleoside Phosphorolysis
Nucleoside analogs represent a class of important drugs for cancer and antiviral treatments. Nucleoside phosphorylases (NPases) catalyze the phosphorolysis of nucleosides and are widely employed for the synthesis of pentose-1-phosphates and nucleoside analogs, which are difficult to access via conventional synthetic methods. However, for the vast majority of nucleosides, it has been observed that either no or incomplete conversion of the starting materials is achieved in NPase-catalyzed reactions. For some substrates, it has been shown that these reactions are reversible equilibrium reactions that adhere to the law of mass action. In this contribution, we broadly demonstrate that nucleoside phosphorolysis is a thermodynamically controlled endothermic reaction that proceeds to a reaction equilibrium dictated by the substrate-specific equilibrium constant of phosphorolysis, irrespective of the type or amount of NPase used, as shown by several examples. Furthermore, we explored the temperature-dependency of nucleoside phosphorolysis equilibrium states and provide the apparent transformed reaction enthalpy and apparent transformed reaction entropy for 24 nucleosides, confirming that these conversions are thermodynamically controlled endothermic reactions. This data allows calculation of the Gibbs free energy and, consequently, the equilibrium constant of phosphorolysis at any given reaction temperature. Overall, our investigations revealed that pyrimidine nucleosides are generally more susceptible to phosphorolysis than purine nucleosides. The data disclosed in this work allow the accurate prediction of phosphorolysis or transglycosylation yields for a range of pyrimidine and purine nucleosides and thus serve to empower further research in the field of nucleoside biocatalysis. (Figure presented.).
Kaspar, Felix,Giessmann, Robert T.,Neubauer, Peter,Wagner, Anke,Gimpel, Matthias
p. 867 - 876
(2020/01/24)

Bio-catalytic synthesis of unnatural nucleosides possessing a large functional group such as a fluorescent molecule by purine nucleoside phosphorylase
Unnatural nucleosides are attracting interest as potential diagnostic tools, medicines, and functional molecules. However, it is difficult to couple unnatural nucleobases to the 1′-position of ribose in high yield and with β-regioselectivity. Purine nucleoside phosphorylase (PNP, EC2.4.2.1) is a metabolic enzyme that catalyses the conversion of inosine to ribose-1α-phosphate and free hypoxanthine in phosphate buffer with 100% α-selectivity. We explored whether PNP can be used to synthesize unnatural nucleosides. PNP catalysed the reaction of thymidine as a ribose donor with purine to produce 2′-deoxynebularine (3, β form) in high conversion (80%). It also catalysed the phosphorolysis of thymidine and introduced a pyrimidine base with a halogen atom substituted at the 5-position into the 1′-position of ribose in moderate yield (52-73%), suggesting that it exhibits loose selectivity. For a bulky purine substrate [e.g., 6-(N,N-di-propylamino)], the yield was lower, but addition of a polar solvent such as dimethyl sulfoxide (DMSO) increased the yield to 74%. PNP also catalysed the reaction between thymidine and uracil possessing a large functional fluorescent group, 5-(coumarin-7-oxyhex-5-yn) uracil (C4U). Conversion to 2′-deoxy-[5-(coumarin-7-oxyhex-5-yn)] uridine (dRC4U) was drastically enhanced by DMSO addition. Docking simulations between dRC4U and E. coli PNP (PDB 3UT6) showed the uracil moiety in the active-site pocket of PNP with the fluorescent moiety at the entrance of the pocket. Thus, the bulky fluorescent moiety has little influence on the coupling reaction. In summary, we have developed an efficient method for producing unnatural nucleosides, including purine derivatives and modified uracil, using PNP.
Hatano, Akihiko,Wakana, Hiroyuki,Terado, Nanae,Kojima, Aoi,Nishioka, Chisato,Iizuka, Yu,Imaizumi, Takuya,Uehara, Sanae
p. 5122 - 5129
(2019/10/05)

Solid-phase synthesis and structural characterisation of phosphoroselenolate-modified DNA: A backbone analogue which does not impose conformational bias and facilitates SAD X-ray crystallography
Oligodeoxynucleotides incorporating internucleotide phosphoroselenolate linkages have been prepared under solid-phase synthesis conditions using dimer phosphoramidites. These dimers were constructed following the high yielding Michaelis-Arbuzov (M-A) reaction of nucleoside H-phosphonate derivatives with 5′-deoxythymidine-5′-selenocyanate and subsequent phosphitylation. Efficient coupling of the dimer phosphoramidites to solid-supported substrates was observed under both manual and automated conditions and required only minor modifications to the standard DNA synthesis cycle. In a further demonstration of the utility of M-A chemistry, the support-bound selenonucleoside was reacted with an H-phosphonate and then chain extended using phosphoramidite chemistry. Following initial unmasking of methyl-protected phosphoroselenolate diesters, pure oligodeoxynucleotides were isolated using standard deprotection and purification procedures and subsequently characterised by mass spectrometry and circular dichroism. The CD spectra of both modified and native duplexes derived from self-complementary sequences with A-form, B-form or mixed conformational preferences were essentially superimposable. These sequences were also used to study the effect of the modification upon duplex stability which showed context-dependent destabilisation (-0.4 to-3.1 °C per phosphoroselenolate) when introduced at the 5′-Termini of A-form or mixed duplexes or at juxtaposed central loci within a B-form duplex (-1.0 °C per modification). As found with other nucleic acids incorporating selenium, expeditious crystallisation of a modified decanucleotide A-form duplex was observed and the structure solved to a resolution of 1.45 ?. The DNA structure adjacent to the modification was not significantly perturbed. The phosphoroselenolate linkage was found to impart resistance to nuclease activity.
Conlon, Patrick F.,Eguaogie, Olga,Wilson, Jordan J.,Sweet, Jamie S. T.,Steinhoegl, Julian,Englert, Klaudia,Hancox, Oliver G. A.,Law, Christopher J.,Allman, Sarah A.,Tucker, James H. R.,Hall, James P.,Vyle, Joseph S.
p. 10948 - 10957
(2019/12/23)

Identification of Flavin Mononucleotide as a Cell-Active Artificial N6-Methyladenosine RNA Demethylase
N6-Methyladenosine (m6A) represents a common and highly dynamic modification in eukaryotic RNA that affects various cellular pathways. Natural dioxygenases such as FTO and ALKBH5 are enzymes that demethylate m6A residues in mRNA. Herein, the first identification of a small-molecule modulator that functions as an artificial m6A demethylase is reported. Flavin mononucleotide (FMN), the metabolite produced by riboflavin kinase, mediates substantial photochemical demethylation of m6A residues of RNA in live cells. This study provides a new perspective to the understanding of demethylation of m6A residues in mRNA and sheds light on the development of powerful small molecules as RNA demethylases and new probes for use in RNA biology.
Xie, Li-Jun,Yang, Xiao-Ti,Wang, Rui-Li,Cheng, Hou-Ping,Li, Zhi-Yan,Liu, Li,Mao, Lanqun,Wang, Ming,Cheng, Liang
supporting information
p. 5028 - 5032
(2019/03/17)

Use of Nucleoside Phosphorylases for the Preparation of Purine and Pyrimidine 2′-Deoxynucleosides
Enzymatic transglycosylation – the transfer of the carbohydrate moiety from one heterocyclic base to another – is being actively developed and applied for the synthesis of practically important nucleosides. This reaction is catalyzed by nucleoside phosphorylases (NPs), which are responsible for reversible phosphorolysis of nucleosides to yield the corresponding heterocyclic bases and monosaccharide 1-phosphates. We found that 7-methyl-2′-deoxyguanosine (7-Me-dGuo) is an efficient and novel donor of the 2-deoxyribose moiety in the enzymatic transglycosylation for the synthesis of purine and pyrimidine 2′-deoxyribonucleosides in excellent yields. Unlike 7-methylguanosine, its 2′-deoxy derivative is dramatically less stable. Fortunately, we have found that 7-methyl-2′-deoxyguanosine hydroiodide may be stored for 24 h in Tris-HCl buffer (pH 7.5) at room temperature without significant decomposition. In order to optimize the reagent ratio, a series of analytical transglycosylation reactions were conducted at ambient temperature. According to HPLC analysis of the transglycosylation reactions, the product 5-ethyl-2′-deoxyuridine (5-Et-dUrd) was obtained in high yield (84–93%) by using a small excess (1.5 and 2.0 equiv.) of 7-Me-dGuo over 5-ethyluracil (5-Et-Ura) and 0.5 equiv. of inorganic phosphate. Thymidine is a less effective precursor of α-d-2-deoxyribofuranose 1-phosphate (dRib-1p) compared to 7-Me-dGuo. We synthesized 2′-deoxyuridine, 5-Et-dUrd, 2′-deoxyadenosine and 2′-deoxyinosine on a semi-preparative scale using the optimized reagent ratio (1.5:1:0.5) in high yields. Unlike other transglycosylation reactions, the synthesis of 2-chloro-2′-deoxyadenosine was performed in a heterogeneous medium because of the poor solubility of the initial 2-chloro-6-aminopurine. Nevertheless, this nucleoside was prepared in good yield. The developed enzymatic procedure for the preparation of 2′-deoxynucleosides may compete with the known chemical approaches. (Figure presented.).
Drenichev, Mikhail S.,Alexeev, Cyril S.,Kurochkin, Nikolay N.,Mikhailov, Sergey N.
p. 305 - 312
(2018/01/15)

Enzymatic Synthesis of Therapeutic Nucleosides using a Highly Versatile Purine Nucleoside 2’-DeoxyribosylTransferase from Trypanosoma brucei
The use of enzymes for the synthesis of nucleoside analogues offers several advantages over multistep chemical methods, including chemo-, regio- and stereoselectivity as well as milder reaction conditions. Herein, the production, characterization and utilization of a purine nucleoside 2’-deoxyribosyltransferase (PDT) from Trypanosoma brucei are reported. TbPDT is a dimer which displays not only excellent activity and stability over a broad range of temperatures (50–70 °C), pH (4–7) and ionic strength (0–500 mM NaCl) but also an unusual high stability under alkaline conditions (pH 8–10). TbPDT is shown to be proficient in the biosynthesis of numerous therapeutic nucleosides, including didanosine, vidarabine, cladribine, fludarabine and nelarabine. The structure-guided replacement of Val11 with either Ala or Ser resulted in variants with 2.8-fold greater activity. TbPDT was also covalently immobilized on glutaraldehyde-activated magnetic microspheres. MTbPDT3 was selected as the best derivative (4200 IU/g, activity recovery of 22 %), and could be easily recaptured and recycled for >25 reactions with negligible loss of activity. Finally, MTbPDT3 was successfully employed in the expedient synthesis of several nucleoside analogues. Taken together, our results support the notion that TbPDT has good potential as an industrial biocatalyst for the synthesis of a wide range of therapeutic nucleosides through an efficient and environmentally friendly methodology.
Pérez, Elena,Sánchez-Murcia, Pedro A.,Jordaan, Justin,Blanco, María Dolores,Manche?o, José Miguel,Gago, Federico,Fernández-Lucas, Jesús
p. 4406 - 4416
(2018/09/14)

Hydrogen peroxide-Triggered gene silencing in mammalian cells through boronated antisense oligonucleotides
Hydrogen peroxide (H2O2) is a reactive oxygen species (ROS) involved in various diseases, including neurodegeneration, diabetes, and cancer. Here, we introduce a new approach to use H2O2 to modulate specific gene expression in mammalian cells. H2O2-responsive nucleoside analogues, in which the Watson-Crick faces of the nucleobases are caged by arylboronate moieties, were synthesized. One of these analogues, boronated thymidine (dTB), was incorporated into oligodeoxynucleotides (ODNs) using an automated DNA synthesizer. The hybridization ability of this boronated ODN to complementary RNA was clearly switched in the off-To-on direction upon H2O2 addition. Furthermore, we demonstrated H2O2-Triggered gene silencing in mammalian cells using antisense oligonucleotides (ASOs) modified with dTB. Our approach can be used for the regulation of any gene of interest by the sequence design of boronated ASOs and will contribute to the development of targeted disease therapeutics.
Mori, Shohei,Morihiro, Kunihiko,Okuda, Takumi,Kasahara, Yuuya,Obika, Satoshi
p. 1112 - 1118
(2018/02/09)

An effective and convenient synthesis of cordycepin from adenosine
Cordycepin is a purine nucleoside analog with potent and diverse biological activities. Herein, we designed two methods to synthesize cordycepin. One method mainly converted the 3′-OH group into an iodide group and further dehalogenation to yield the final product. Although this method presented a short synthetic procedure, the synthesis had a low overall yield, resulting in only 13.5% overall yield. To improve the overall yield of cordycepin, another synthetic route was studied, which consisted of four individual steps: (1) 5′-OH protection (2) esterification (3) -O-tosyl (-OTs) group removal (4) deprotection. The key step in the synthetic method involved the conversion of 5′-O-triphenylmethyladenosine to 3′-O-tosyl-5′-O-triphenylmethyladenosine, using LiAlH4 as reducing agent. The main advantages of this route were an acceptable total product yield and the commercial availability of all starting materials. The optimal reaction conditions for each step of the route were identified. The overall yield of cordycepin obtained from adenosine as the starting material was 36%.
Huang, Shen,Liu, Hui,Sun, Yanhua,Chen, Jian,Li, Xiufang,Xu, Jiangfeng,Hu, Yuwei,Li, Yuqing,Deng, Zhiwei,Zhong, Shian
p. 149 - 160
(2018/01/17)

Independent Photochemical Generation and Reactivity of Nitrogen-Centered Purine Nucleoside Radicals from Hydrazines
Photochemical precursors that produce dA￠ and dG(N2-H)￠ are needed to investigate their reactivity. The synthesis of two 1,1-diphenylhydrazines (1, 2) and their use as photochemical sources of dA￠ and dG(N2-H)￠ is presented. Trapping studies indicate production of these radicals with good fidelity, and 1 was incorporated into an oligonucleotide via solid-phase synthesis. Cyclic voltammetric studies show that reduction potentials of 1 and 2 are lower than those of widely used "hole sinks", e.g., 8-oxodGuo and 7-deazadGuo, to investigate DNA-hole transfer processes. These molecules could be useful (a) as sources of dA￠ and dG(N2-H)￠ at specific sites in oligonucleotides and (b) as "hole sinks" for the study of DNA-hole transfer processes.
Zheng, Liwei,Lin, Lu,Qu, Ke,Adhikary, Amitava,Sevilla, Michael D.,Greenberg, Marc M.
p. 6444 - 6447
(2017/12/08)

Photochemical demethylation method for N6-methyladenine
The invention relates to a specific photochemical demethylation method for N6-methyladenine and application of N6-methyladenine. According to the method, light is used as an initiator; and under the existence of a solvent, N6-methyladenine as shown in the formula I or a derivative of N6-methyladenine reacts with a photosensitizer and an oxidant under an illumination condition, thus obtaining a demethylated product as shown in the formula II and realizing demethylation. By the use of the photochemical demethylation method for the N6-methyladenine and relevant compounds, chemical adjustment of nucleic acid under a non-enzymic condition can be adjusted. The invention provides an effective research method on nucleic acid demethylation for the field of researches on epigenetics and chemicobiology.
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Paragraph 0039; 0040

(2017/08/29)

Visible-light-mediated oxidative demethylation of: N 6-methyl adenines
We report a simple protocol that affords oxidative demethylation of N6-methyl groups in N6-methyl adenines (m6A). The biologically compatible photocatalyst riboflavin prompts a highly selective C-H abstraction from N6-methyl in adenines under the irradiation of a visible blue LED light, affording a novel and highly selective biomimetic demethylation of m6A and related N-methyl adenine analogues. andcopy; 2017 The Royal Society of Chemistry.
Xie, Li-Jun,Wang, Rui-Li,Wang, Dong,Liu, Li,Cheng, Liang
supporting information
p. 10734 - 10737
(2017/10/06)

Enzymatic synthesis of ribo- and 2′-deoxyribonucleosides from glycofuranosyl phosphates: An approach to facilitate isotopic labeling
Milligram quantities of α-D-ribofuranosyl 1-phosphate (sodium salt) (αR1P) were prepared by the phosphorolysis of inosine, catalyzed by purine nucleoside phosphorylase (PNPase). The αR1P was isolated by chromatography in >95% purity and characterized by 1H and 13C NMR spectroscopy. Aqueous solutions of αR1P were stable at pH 6.4 and 4 °C for several months. The isolated αR1P was N-glycosylated with different nitrogen bases (adenine, guanine and uracil) using PNPase or uridine phosphorylase (UPase) to give the corresponding ribonucleosides in high yield based on the glycosyl phosphate. This methodology is attractive for the preparation of stable isotopically labeled ribo- and 2′-deoxyribonucleosides because of the ease of product purification and convenient use and recycling of nitrogen bases. The approach eliminates the need for separate reactions to prepare individual furanose-labeled ribonucleosides, since only one ribonucleoside (inosine) needs to be labeled, if desired, in the furanose ring, the latter achieved by a high-yield chemical N-glycosylation. 2′-Deoxyribonucleosides were prepared from 2′-deoxyinosine using the same methodology with minor modifications.
Zhang, Wenhui,Turney, Toby,Surjancev, Ivana,Serianni, Anthony S.
p. 125 - 133
(2017/08/08)

The synthesis of nebularine and its analogs via oxidative desulfuration in aqueous nitric acid
The synthesis of nebularine and its analogs has been achieved via oxidative desulfuration in H2O for the first time. With 50% HNO3as an oxidant and solvent, 18 products were obtained in good yields (70%–94%). The oxidative desulfuration system could tolerate different functional groups including fluoro, chloro, amino, alkyl, allyl, ribosyl, deoxyribosyl, and arabinofuranosyl groups.More importantly, the drug nebularine could be obtained successfully on a 20 g scale, which made this route more attractive for industrial applications.
Xia, Ran,Sun, Li-Ping,Qu, Gui-Rong
p. 88 - 91
(2016/12/24)

Synthesis of Adenine Nucleosides by Transglycosylation using Two Sequential Nucleoside Phosphorylase-Based Bioreactors with On-Line Reaction Monitoring by using HPLC
Uridine phosphorylase from Clostridium perfringens (CpUP, EC 2.4.2.3) was immobilized covalently in an aminopropylsilica monolithic column (25 mm×4.6 mm) upon functionalization with glutaraldehyde. Imino bonds that result from the reaction between the enzyme and the support were reduced chemically to afford a 66 % yield (13 mg) determined spectrophotometrically. The CpUP immobilized enzyme reactor (IMER) was connected to a silica particle-based IMER that contained a purine nucleoside phosphorylase from Aeromonas hydrophila (AhPNP, EC 2.4.2.1), which was developed previously and used successfully for the fast synthesis of some purine ribonucleosides by a “one-enzyme” transglycosylation. CpUP-IMER and AhPNP-IMER were connected to a HPLC system by a six-way switching valve. In this set-up, the synthesis of 2′-deoxyadenosine (dAdo, 8), adenosine (Ado, 9), and arabinosyladenine (araA, 10) by a “two-enzyme” transglycosylation is coupled directly to on-line reaction monitoring. Under the optimized transglycosylation conditions (2:1 ratio sugar donor/base acceptor; 10 mm phosphate buffer; pH 7.25; temperature 37 °C, flow rate 0.1 mL min?1), defined by a 2(5-2) III experimental design, the conversion of dAdo and Ado was approximately 90 %, and araA was synthesized in 20 % yield.
Cattaneo, Giulia,Rabuffetti, Marco,Speranza, Giovanna,Kupfer, Tom,Peters, Benjamin,Massolini, Gabriella,Ubiali, Daniela,Calleri, Enrica
p. 4614 - 4620
(2017/12/13)

Synthesis method for 2'-deoxyadenosine monohydrate
The invention discloses a synthesis method for 2'-deoxyadenosine monohydrate. Adenosine is esterified and acylated by an acylating agent and an acid-binding agent so that acylate can be obtained, the acylate is subjected to reduction and purification, and then 2'-deoxyadenosine monohydrate is obtained, wherein in the esterifying process, dialkyl group stannic oxide is adopted as an esterifying agent. According to the method, reaction selectivity is high, chromatographic separation is not needed, cost is low, and the yield is high; the synthesis method is applicable to industrial production.
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Paragraph 0080; 0081

(2016/12/22)

Identification of a Fused-Ring 2′-Deoxyadenosine Adduct Formed in Human Cells Incubated with 1-Chloro-3-buten-2-one, a Potential Reactive Metabolite of 1,3-Butadiene
1-Chloro-3-buten-2-one (CBO) is an in vitro metabolite of 1,3-butadiene (BD), a carcinogenic air pollutant. CBO exhibited potent cytotoxicity and genotoxicity that have been attributed in part to its reactivity toward DNA. Previously, we have characterized the CBO adducts with 2′-deoxycytidine and 2′-deoxyguanosine. In the present study, we report on the reaction of CBO with 2′-deoxyadenosine (dA) under in vitro physiological conditions (pH 7.4, 37 °C). We used the synthesized standards and their decomposition and acid-hydrolysis products to characterize the CBO-DNA adducts formed in human cells. The fused-ring dA adducts (dA-1 and dA-2) were readily synthesized and were structurally characterized as 1,N6-(1-hydroxy-1-hydroxymethylpropan-1,3-diyl)-2′-deoxyadenosine and 1,N6-(1-hydroxy-1-chloromethylpropan-1,3-diyl)-2′-deoxyadenosine, respectively. dA-1 exhibited a half-life of 16.0 ± 0.7 h and decomposed to dA at pH 7.4 and 37 °C. At similar conditions, dA-2 decomposed to dA-1 and dA, and had a half-life of 0.9 ± 0.1 h. These results provide strong evidence for dA-1 being a degradation product of dA-2. dA-1 is formed by replacement of the chlorine atom of dA-2 by a hydroxyl group. The slow decomposition of dA-1 to dA, along with the detection of hydroxymethyl vinyl ketone (HMVK) as another degradation product, suggested equilibrium between dA-1 and a ring-opened carbonyl-containing intermediate that undergoes a retro-Michael reaction to yield dA and HMVK. Acid hydrolysis of dA-1 and dA-2 yielded the corresponding deribosylated products A-1D and A-2D, respectively. In the acid-hydrolyzed reaction mixture of CBO with calf thymus DNA, both A-1D and A-2D could be detected; however, the amount of A-2D was significantly larger than that of A-1D. Interestingly, only A-2D could be detected by LC-MS analysis of acid-hydrolyzed DNA from cells incubated with CBO, suggesting that dA-2 was stable in DNA and thus may play an important role in the genotoxicity and carcinogenicity of BD. In addition, A-2D could be developed as a biomarker of CBO formation in human cells.
Zeng, Fang-Mao,Liu, Ling-Yan,Zheng, Jin,Kong, Cong,An, Jing,Yu, Ying-Xin,Zhang, Xin-Yu,Elfarra, Adnan A.
p. 1041 - 1050
(2016/07/06)

PRODUCTION METHOD OF NUCLEOSIDE COMPOUND
PROBLEM TO BE SOLVED: To provide a production method of a nucleoside compound by which an isotopic labeled nucleoside compound can be produced efficiently. SOLUTION: A production method of a nucleoside compound comprises obtaining a target nucleoside compound by the base exchange reaction of a raw material nucleoside compound and a base in the solution containing a phosphoric acid ion by a nucleoside phosphorylase, wherein the target nucleoside compound is labeled with a stable isotope or a radioisotope. COPYRIGHT: (C)2015,JPOandINPIT
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Paragraph 0032-0033

(2017/03/24)

Mechanisms of allosteric activation and inhibition of the deoxyribonucleoside triphosphate triphosphohydrolase from Enterococcus faecalis
EF1143 from Enterococcus faecalis, a life-threatening pathogen that is resistant to common antibiotics, is a homo-tetrameric deoxyribonucleoside triphosphate (dNTP) triphosphohydrolase (dNTPase), converting dNTPs into the deoxyribonucleosides and triphosphate. The dNTPase activity of EF1143 is regulated by canonical dNTPs, which simultaneously act as substrates and activity modulators. Previous crystal structures of apo-EF1143 and the protein bound to both dGTP and dATP suggested allosteric regulation of its enzymatic activity by dGTP binding at four identical allosteric sites. However, whether and how other canonical dNTPsregulatetheenzyme activity was not defined.Here, wepresent the crystal structure of EF1143 in complex with dGTP and dTTP. The new structure reveals that the tetrameric EF1143 contains four additional secondary allosteric sites adjacent to the previously identified dGTP-binding primary regulatory sites. Structural and enzyme kinetic studies indicate that dGTP binding to the first allosteric site, with nanomolar affinity, is a prerequisite for substrate docking and hydrolysis. Then, the presence of a particular dNTP in the second site either enhances or inhibits the dNTPase activity of EF1143. Our results provide the first mechanistic insight into dNTP-mediated regulation of dNTPase activity.
Vorontsov, Ivan I.,Wu, Ying,Delucia, Maria,Minasov, George,Mehrens, Jennifer,Shuvalova, Ludmilla,Anderson, Wayne F.,Ahn, Jinwoo
p. 2815 - 2824
(2014/02/14)

Nucleoside 2′-deoxyribosyltransferase from psychrophilic bacterium Bacillus psychrosaccharolyticus - Preparation of an immobilized biocatalyst for the enzymatic synthesis of therapeutic nucleosides
Nucleoside 2′-deoxyribosyltransferase (NDT) from the psychrophilic bacterium Bacillus psychrosaccharolyticus CECT 4074 has been cloned and produced for the first time. A preliminary characterization of the recombinant protein indicates that the enzyme is an NDT type II since it catalyzes the transfer of 2′-deoxyribose between purines and pyrimidines. The enzyme (BpNDT) displays a high activity and stability in a broad range of pH and temperature. In addition, different approaches for the immobilization of BpNDT onto several supports have been studied in order to prepare a suitable biocatalyst for the one-step industrial enzymatic synthesis of different therapeutic nucleosides. Best results were obtained by adsorbing the enzyme on PEI-functionalized agarose and subsequent cross-linking with aldehyde-dextran (20 kDa and 70% oxidation degree). The immobilized enzyme could be recycled for at least 30 consecutive cycles in the synthesis of 2′-deoxyadenosine from 2′-deoxyuridine and adenine at 37 °C and pH 8.0, with a 25% loss of activity. High conversion yield of trifluridine (64.4%) was achieved in 2 h when 20 mM of 2′-deoxyuridine and 10 mM 5-trifluorothymine were employed in the transglycosylation reaction catalyzed by immobilized BpNDT at 37 °C and pH 7.5.
Fresco-Taboada, Alba,Serra, Immacolata,Fernandez-Lucas, Jesus,Acebal, Carmen,Arroyo, Miguel,Terreni, Marco,De La Mata, Isabel
p. 11231 - 11249
(2014/11/08)

Synthesis and stability of a 2′-O-[N-(aminoethyl)carbamoyl] methyladenosine-containing dinucleotide
Working towards the synthesis of 2′-O-[N-(aminoethyl)carbamoyl] methyl-modified di- and oligonucleotides, we have synthesised a protected 2′-O-[N-(aminoethyl)carbamoyl]methyl-modified adenosine where the modification is introduced in a convenient one-pot three-step procedure. The corresponding H-phosphonate building block was also synthesised, and from this intermediate, a 2′-O-[N-(aminoethyl)carbamoyl]methyl-containing dinucleotide could be made. We also performed studies on the chemical and enzymatic stability of this dinucleotide. The dinucleotide was subjected to different ammonolysis and other basic conditions, and HPLC analysis showed that the modification was intact to most conditions, but that there was some minor hydrolysis when NH3 (concd. aq.) was used at 55 °C. Under several other sets of conditions, including saturated NH3 in methanol, and ethylenediamine, the amide remained intact. Treatment of the dinucleotide with Phosphodiesterase I from Crotalus adamanteus venom and Phosphodiesterase II from bovine spleen showed that the N-(aminoethyl)carbamoylmethyl moiety gives the phosphodiester linkage substantial protection against enzymatic degradation; the phosphodiester was not degraded by PDE II at all after seven days. A 2′-O-[N-(aminoethyl)carbamoyl]methyl-modified adenosine and a corresponding dinucleotide were synthesised. Hydrolysis (1-2 %) was observed in concentrated aqueous NH3 at 55°C, but under several other sets of reaction conditions, the amide remained intact. The N-(aminoethyl) carbamoylmethyl moiety gave substantial protection against enzymatic degradation by nucleases from snake venom and bovine spleen. Copyright
Milton, Stefan,Ander, Charlotte,Honcharenko, Dmytro,Honcharenko, Malgorzata,Yeheskiely, Esther,Stroemberg, Roger
p. 7184 - 7192
(2013/11/06)

Developing a collection of immobilized nucleoside phosphorylases for the preparation of nucleoside analogues: Enzymatic synthesis of arabinosyladenine and 2',3'-dideoxyinosine
The use of nucleoside phosphorylases (NPs; EC 2.4.2.n) represents a convenient alternative to the chemical route for the synthesis of natural and modified nucleosides. We purified four recombinantly expressed nucleoside phosphorylases from the bacterial pathogens Citrobacter koseri, Clostridium perfringens, and Streptococcus pyogenes (CkPNPI, CkPNPII, CpUP, SpUP) and their substrate specificity was investigated towards either natural pyrimidine or purine nucleosides and some analogues, namely, arabinosyladenine (araA) and 2',3'-dideoxyinosine (ddI). A 2-3 % activity towards these latter compounds (compared to the natural substrates) was observed. Enzyme activities were compared to the specificities obtained for the enzymes pyrimidine nucleoside phosphorylase from Bacillus subtilis (BsPyNP) and purine nucleoside phosphorylase from Aeromonas hydrophila (AhPNPII) previously reported by some of the authors. The enzymes displaying the suitable specificity for the synthesis of araA and ddI were immobilized on aldehyde-agarose. The immobilized preparations were highly stable at alkaline pH and in the presence of methanol or acetonitrile as cosolvent. They were used in the synthesis of araA and ddI by a one-pot, bienzymatic transglycosylation achieving 74 and 44 % conversion, respectively. Something different: Nucleoside phosphorylases are a convenient alternative to the chemical route for the synthesis of natural and modified nucleosides. Four new nucleoside phosphorylases have been prepared, characterized, and tested for their use in biocatalyzed syntheses of araA and ddI (see scheme). A generally applicable immobilization technique has been found to provide active and stable biocatalysts.
Serra, Immacolata,Ubiali, Daniela,Piskur, Jure,Christoffersen, Stig,Lewkowicz, Elizabeth S.,Iribarren, Adolfo M.,Albertini, Alessandra M.,Terreni, Marco
p. 157 - 165
(2013/04/24)

2,3-Dicyclohexylsuccinimide as a directing/protecting group for the regioselective glycosylation or alkylation of purines
Here we describe the synthesis and application of a novel 2,3-dicyclohexylsuccinimide (Cy2SI) protecting group towards regioselective purine glycosylation and alkylation reactions. This bulky protecting group promotes high regioselectivity during the glycosylation (as well as diastereoselectivity) or alkylation of purines using Hoffer's chlorosugar or tert-butyl bromoacetate, respectively. Cy2SI offers the additional synthetic advantage that other base-labile protecting groups, such as toluoyl esters, can be selectively removed in its presence without affecting the imide. The Royal Society of Chemistry 2013.
Pal, Ayan,Salandria, Kerry J.,Arico, Joseph W.,Schlegel, Mark K.,McLaughlin, Larry W.
supporting information
p. 2936 - 2938
(2013/04/24)

Phosphorylating reagent-free synthesis of 5′-phosphate oligonucleotides by controlled oxidative degradation of their 5′-end
The 5′-phosphorylated oligonucleotides (5′-pONs) are currently synthesized using expensive and sensitive modified phosphoramidite reagents. In this work, a simple, cost-effective, efficient, and automatable method is presented, based on the controlled oxidation of the 5′-terminal alcohol followed by a β-elimination reaction. The latter reaction leads to the removal of the terminal 5′-nucleoside and subsequent formation of the 5′-phosphate moiety. Thus, chemical phosphorylation of oligonucleotides (DNA or RNA) is achieved without using modified phosphoramidites.
Sallamand, Corinne,Miscioscia, Audrey,Lartia, Remy,Defrancq, Eric
supporting information; body text
p. 2030 - 2033
(2012/06/18)

Use of Citrobacter koseri whole cells for the production of arabinonucleosides: A larger scale approach
Purine arabinosides are well known antiviral and antineoplastic drugs. Since their chemical synthesis is complex, time-consuming, and polluting, enzymatic synthesis provides an advantageous alternative. In this work, we describe the microbial whole cell synthesis of purine arabinosides through nucleoside phosphorylase-catalyzed transglycosylation starting from their pyrimidine precursors. By screening of our microbial collection, Citrobacter koseri (CECT 856) was selected as the best biocatalyst for the proposed biotransformation. In order to enlarge the scale of the transformations to 150 mL for future industrial applications, the biocatalyst immobilization by entrapment techniques and its behavior in different reactor configurations, considering both batch and continuous processes, were analyzed. C. koseri immobilized in agarose could be used up to 68 times and the storage stability was at least 9 months. By this approach, fludarabine (58% yield in 14 h), vidarabine (71% yield in 26 h) and 2,6-diaminopurine arabinoside (77% yield in 24 h), were prepared.
Nóbile, Matías,Médici, Rosario,Terreni, Marco,Lewkowicz, Elizabeth S.,Iribarren, Adolfo M.
p. 2182 - 2188
(2013/02/25)

5-Fluoro-4-thiouridine phosphoramidite: New synthon for introducing photoaffinity label into oligodeoxynucleotides
The synthesis of phosphoramidite of 5-fluoro-4-thio-2′-O- methyluridine is described. An appropriate set of protecting groups was optimized including the 4-thio function introduced via 4-triazolyl as the 4-(2-cyanoethyl)thio derivative, and the t-butyldimethyl silyl for 2′ and 3′ hydroxyl protection, enabling efficient synthesis of the phosphoramidite. These protecting groups prevented unwanted side reactions during oligonucleotide synthesis. The utility of the proposed synthetic route was proven by the preparation of several oligonucleotides via automated synthesis. Photochemical experiments confirmed the utility of the synthon.
Milecki, Jan,Nowak, Joanna,Skalski, Bohdan,Franzen, Stefan
experimental part
p. 6098 - 6106
(2011/11/07)

NANO-REAGENTS WITH COOPERATIVE CATALYSIS AND THEIR USES IN MULTIPLE PHASE REACTIONS
Nano-reagents with catalytic activity are provided herein. The nanocatalyst comprises at least one amino acid attached to a nanoparticle, wherein the reactive side chain of the amino acid catalyzes a chemical or biological reaction. Methods of using these nano-reagents to catalyze reactions in solution or in multiple phases are also provided, as are methods of making these nanocatalysts.
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(2011/08/08)

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 1-2nm, were measured. The spin-spin distance was 1.8A±0. 2nm for DNA duplex 17(dA7,10)·11 containing two spin labels that are separated by two nucleotides within one individual strand. A distance of 1.4A±0.2nm was found for the spin-labeled 'dA-dT' base pair 15(dA7)·16(dT6). 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. 'Clicked' DNA spin labels: Spin labels (spheres in the figure) have been incorporated by click chemistry into DNA duplexes at spatially separated positions or into a 'dA-dT' base pair with high efficiency. Very accurate distances between spin labels, within a range of 1-2nm, were measured by continuous wave and pulse EPR spectroscopy. Copyright
Ding, Ping,Wunnicke, Dorith,Steinhoff, Heinz-Juergen,Seela, Frank
supporting information; scheme or table
p. 14385 - 14396
(2011/03/22)

Synthesis of oligodeoxynucleotides using fully protected Deoxynucleoside 3c-Phosphoramidite building blocks and base recognition of Oligodeoxynucleotides incorporating N3-Cyano-Ethylthymine
Oligodeoxynucleotide (ODN) synthesis, which avoids the formation of side products, is of great importance to biochemistry-based technology development. One side reaction of ODN synthesis is the cyanoethylation of the nucleobases. We suppressed this reaction by synthesizing ODNs using fully protected deoxynucleoside 3c-phosphoramidite building blocks, where the remaining reactive nucleobase residues were completely protected with acyl-, diacyl-, and acyl-oxyethylene-type groups. The detailed analysis of cyanoethylation at the nucleobase site showed that N3-protection of the thymine base efficiently suppressed the Michael addition of acrylonitrile. An ODN incorporating N3-cyanoethylthymine was synthesized using the phosphoramidite method, and primer extension reactions involving this ODN template were examined. As a result, the modified thymine produced has been proven to serve as a chain terminator.
Tsunoda, Hirosuke,Kudo, Tomomi,Ohkubo, Akihiro,Seio, Kohji,Sekine, Mitsuo
experimental part
p. 7509 - 7531
(2011/02/28)

Generation of 2′-deoxyadenosine N6-aminyl radicals from the photolysis of phenylhydrazone derivatives
Nitrogen-centered radicals are major species generated by the addition of hydroxyl radicals and the one-electron oxidation of adenine derivatives. Aminyl radicals are also generated in the decomposition of adenine chloramines upon reaction of hypochlorite. Here, we report the photochemistry of modified 2′-deoxyadenosine (dAdo) containing photoactive hydrazone substituents as a model to investigate the chemistry of dAdo N6-aminyl radicals. Derivatives of dAdo containing a phenylhydrazone moiety at N6 displayed UV absorption between 300 and 400 nm. Upon UV photolysis in the presence of a H-donor, that is, glutathione, two major products were formed, dAdo and benzaldehyde, indicating efficient homolytic cleavage to dAdo N 6-aminyl radicals and benzylidene iminyl radicals. dAdo N 6-phenylhydrazone was photolyzed in the presence of a molar excess of nonmodified dAdo to mimic the reactions taking place in DNA, and the major photoproducts were identified by high-performance liquid chromatography, mass spectrometry, and nuclear magnetic resonance. The formation of 2-(benzylideneamino)-2′-deoxyadenosine as well as a more extensive oxidation product may be explained by the recombination of initial dAdo N 6-aminyl and benzylidene iminyl radicals. The formation of 2′-deoxyinosine may be explained by hydrolytic deamination of dAdo N 6-aminyl radicals. Interestingly, a dimeric product containing two dAdo moieties was identified in the photolysis mixture. The present studies demonstrate the ability of dAdo N6-aminyl radicals to undergo H-abstraction to give dAdo, deamination to give 2′-deoxyinosine, and addition to the adenine moiety to give dimers.
Kuttappan-Nair, Vandana,Samson-Thibault, Francois,Wagner, J. Richard
scheme or table
p. 48 - 54
(2011/01/12)

Photolysis and thermolysis of platinum(IV) 2,2′-bipyridine complexes lead to identical platinum(II)-DNA adducts
Two PtIV and two Pt11 complexes containing a 2,2′-bipyridine ligand were treated with a short DNA oligonucleotide under light irradiation at 37 °C or in the dark at 37 and 50 °C. Photolysis and thermolysis of the PtIV complexes led to spontaneous reduction of the PtIV to the corresponding PtII complexes and to binding of PtII 2,2′-bipyridine complexes to N7 of guanine. When the reduction product was [Pt(bpy)Cl2], formation of bis-oligonucleotide adducts was observed, whereas [Pt(bpy)(MeNH 2)Cl]+ gave monoad- ducts, with chloride ligands substituted in both cases. Neither in the dark nor under light irradiation was the reductive elimination process of these PtIV complexes accompanied by oxidative DNA damage. This work raises the question of the stability of photoacti- vatable PtIV complexes toward moderate heating conditions.
Loup, Christophe,Tesouro Vallina, Ana,Coppel, Yannick,Letinois, Ulla,Nakabayashi, Yasuo,Meunier, Bernard,Lippert, Bernhard,Pratviel, Genevieve
experimental part
p. 11420 - 11431
(2010/11/24)

Radiation-induced formation of purine 5′,8-cyclonucleosides in isolated and cellular DNA: High stereospecificity and modulating effect of oxygen
The present work is aimed at gaining conclusive mechanistic insights into the radiation-induced formation of the 5′R and 5′S diastereomers of both adenine and guanine 5′,8-cyclo-2′-deoxyribonucleosides, with emphasis on the delineation of the inhibitory effect of O2 in isolated and cellular DNA. The levels of purine 5′,8-cyclo-2′- deoxyribonucleosides as assessed by HPLC-MS/MS were found to decrease steadily with the increase of O2 concentration, the 5′,8-cyclo-2′- deoxyguanosine being produced more efficiently than the 5′,8-cyclo- 2′-deoxyadenosine for low O2 concentrations. A high stereoselectivity was observed in the intramolecular addition of the C5′ radical to the C8 of the purine leading, after the creation of the C5′-C8 bond and a subsequent oxidation step, to the predominant formation of the 5′R diastereomer for both purine 5′,8-cyclonucleosides. The reduced formation yield of the 4 tandem lesions in the presence of O2 explains, at least partly, the low efficiency of radiation-induced yields of the purine 5′,8-cyclo-2′-deoxyribonucleosides in cellular DNA, which are about two orders of magnitude lower than the previously reported data obtained from HPLC-MS analysis. The Royal Society of Chemistry 2010.
Belmadoui, Nourreddine,Boussicault, Fabien,Guerra, Maurizio,Ravanat, Jean-Luc,Chatgilialoglu, Chryssostomos,Cadet, Jean
experimental part
p. 3211 - 3219
(2010/08/21)

Oligonucleotide synthesis involving deprotection of amidine-type protecting groups for nucleobases under acidic conditions
Amidine-type protecting groups, i.e., N,N-dimethylformamidine (dmf) and N,N-dibutylformamidine (dbf) groups, introduced into nucleobases were rapidly removed under mild acidic conditions using imidazolium triflate (IMT) or 1-hydroxybenztriazole (HOBt). This new deprotection strategy allowed a 2′-O-methyl-RNA derivative bearing a base-labile group to be efficiently synthesized using a silyl-type linker. It was also found that our new method could be applied to the synthesis of an unmodified RNA oligomer.
Ohkubo, Akihiro,Kuwayama, Yasukazu,Nishino, Yudai,Tsunoda, Hirosuke,Seio, Kohji,Sekine, Mitsuo
supporting information; scheme or table
p. 2496 - 2499
(2010/08/07)

Aeromonas hydrophila strains as biocatalysts for transglycosylation
Microbial transglycosylation is useful as a green alternative in the preparation of purine nucleosides and analogues, especially for those that display pharmacological activities. In a search for new transglycosylation biocatalysts, two Aeromonas hydrophila strains were selected. The substrate specificity of both micro-organisms was studied and, as a result, several nucleoside analogues have been prepared. Among them, ribavirin, a broad spectrum antiviral, and the well-known anti HIV didanosine, were prepared, in 77 and 62% yield using A. hydrophila CECT 4226 and A. hydrophila CECT 4221, respectively. In order to scale-up the processes, the reaction conditions, product purification and biocatalyst preparation were analyzed and optimized.
Nbile, Matas,Terreni, Marco,Lewkowicz, Elizabeth,Iribarren, Adolfo M.
experimental part
p. 395 - 402
(2011/10/08)

Enzymatic transglycosylation of natural and modified nucleosides by immobilized thermostable nucleoside phosphorylases from Geobacillus stearothermophilus
Natural and modified purine nucleosides have been synthesized using the recombinant thermostable enzymes purine nucleoside phosphorylase II (E. C. 2.4.2.1) and pyrimidine nucleoside phosphorylase (E. C. 2.4.2.2) from Geobacillus stearothermophilus B-2194. The enzymes were produced in recombinant E. coli strains and covalently immobilized on aminopropylsilochrom AP-CPG-170 after heating the cell lysates and the removal of coagulated thermolabile proteins. The resulting preparations of thermostable nucleoside phosphorylases retained a high activity after 20 reuses in nucleoside transglycosylation reactions at 70-75°C with a yield of the target products as high as 96%. Owing to the high catalytic activity, thermal stability, the ease of application, and the possibility of repeated use, the immobilized preparations of thermostable nucleoside phosphorylases are suitable for the production of pharmacologically important natural and modified nucleosides.
Taran,Verevkina,Feofanov,Miroshnikov
experimental part
p. 739 - 745
(2010/08/07)

DNA interstrand cross-link formation by the 1,4-dioxobutane abasic lesion
The oxidized abasic lesion 5′-(2-phosphoryl-1,4-dioxobutane) (DOB) is produced concomitantly with a single-strand break by a variety of DNA-damaging agents that abstract a hydrogen atom from the C5′-position. Independent generation of the DOB lesion in DN
Guan, Lirui,Greenberg, Marc M.
supporting information; experimental part
p. 15225 - 15231
(2010/01/29)

New thermolytic carbamoyl groups for the protection of nucleobases
It was found that N-arylcarbamoyl and N-(phenylsulfonyl)carbamoyl (psc) groups could be effectively introduced onto the amino groups of deoxycytidine and deoxyadenosine derivatives and could be removed thermolytically. We succeeded in synthesizing DNA pro
Ohkubo, Akihiro,Kasuya, Rintaro,Miyata, Kenichi,Tsunoda, Hirosuke,Seio, Kohji,Sekine, Mitsuo
experimental part
p. 687 - 694
(2009/06/20)

Effect of base stacking on the acid-base properties of the adenine cation radical [A?+] in solution: ESR and DFT studies
In this study, the acid-base properties of the adenine cation radical are investigated by means of experiment and theory. Adenine cation radical (A?+) is produced by one-electron oxidation of dAdo and of the stacked DNA-oligomer (dA)6 by Cl2?- in aqueous glass (7.5 M LiCl in H2O and in D2O) and investigated by ESR spectroscopy. Theoretical calculations and deuterium substitution at C8-H and N6-H in dAdo aid in our assignments of structure. We find the pKa value of A?+ in this system to be ca. 8 at 150 K in seeming contradiction to the accepted value of ≤1 at ambient temperature. However, upon thermal annealing to ≥160 K, complete deprotonation of A?+ occurs in dAdo in these glassy systems even at pH ca. 3. A?+ found in (dA)6 at 150 K also deprotonates on thermal annealing. The stability of A?+ at 150 K in these systems is attributed to charge derealization between stacked bases. Theoretical calculations at various levels (DFT B3LYP/6-31G*, MPWB95, and HF-MP2) predict binding energies for the adenine stacked dimer cation radical of 12 to 16 kcal/mol. Further DFT B3LYP/6-31G* calculations predict that, in aqueous solution, monomeric A?+ should deprotonate spontaneously (a predicted pKa of ca. -0.3 for A?+). However, the charge resonance stabilized dimer AA?+ is predicted to result in a significant barrier to deprotonation and a calculated pK a of ca. 7 for the AA?+ dimer which is 7 pH units higher than the monomer. These theoretical and experimental results suggest that A?+ isolated in solution and A?+ in adenine stacks have highly differing acid-base properties resulting from the stabilization induced by hole derealization within adenine stacks.
Adhikary, Amitava,Kumar, Anil,Khanduri, Deepti,Sevilla, Michael D.
experimental part
p. 10282 - 10292
(2009/02/03)

A kinetic study of the rat liver adenosine kinase reverse reaction
Adenosine kinase is an enzyme catalyzing the reaction: adenosine + ATP → AMP + ADP. We studied some biochemical properties not hitherto investigated and demonstrated that the reaction can be easily reversed when coupled with adenosine deaminase, which transforms adenosine into inosine and ammonia. The overall reaction is: AMP + ADP → ATP + inosine + NH3. The exoergonic ADA reaction shifts the equilibrium and fills the energy gap necessary for synthesis of ATP. This reaction could be used by cells under particular conditions of energy deficiency and, together with myokinase activity, may help to restore physiological ATP levels. Copyright Taylor & Francis Group, LLC.
Vannoni,Giglioni,Santoro,Aceto,Marinello,Leoncini
p. 872 - 875
(2008/12/21)

Reaction of glycidamide with 2′-deoxyadenosine and 2′-deoxyguanosine - Mechanism for the amide hydrolysis
2′-Deoxyadenosine (dA) and 2′-deoxyguanosine (dG) were reacted with the mutagenic epoxide glycidamide (GA, Scheme 1). The reactions yielded three GA-dA adducts (N1-GA-dA, N6-GA-dA and N1-GA-dI) and two GA-dG adducts (N1-GA-dG I and N1-GA-dG II) (). The structures of the adducts were characterized by spectroscopic and spectrometric methods (1H-, 13C, and 2D NMR, MS, UV). The mechanism of the amide hydrolysis taking place during formation of the adducts N1-GA-dA and N1-GA-dG I was studied. We propose a mechanism where a transamidation is the key step in the hydrolysis of the amide function of GA. Copyright Taylor & Francis Group, LLC.
Backman, Josefin,Kronberg, Leif
p. 129 - 148
(2008/09/18)

Radical reactions in aqueous medium using (Me3Si)3SiH
(Chemical Equation Presented) (Me3Si)3SiH was used as a successful reagent in a variety of radical-based transformations in water. The system comprising substrate, silane, and initiator (ACCN) mixed in aqueous medium at 100°C worked well for both hydrophilic and hydrophobic substrates, with the only variation that an amphiphilic thiol was also needed in case of the water-soluble compounds.
Postigo, Al,Kopsov, Sergey,Ferreri, Carla,Chatgilialoglu, Chryssostomos
p. 5159 - 5162
(2008/09/17)

Flow injection amperometric detection of 2′-deoxyguanosine at a ruthenium oxide hexacyanoferrate modified electrode
A ruthenium oxide hexacyanoferrate (RuOHCF) modified electrode was developed. Hydrodynamic voltammetry was employed to demonstrate the remarkable electrocatalytic activity toward the oxidation of 2′-deoxyguanosine. The RuOHCF modified electrode was used as amperometric detector for 2′-deoxyguanosine determination in a FIA apparatus. The influence of various experimental conditions was explored for optimum analytical performance, and at these experimental conditions, the method exhibited a linear response range to 2′-deoxyguanosine extending from 3.8 to 252 μmol L -1 with detection limit of 94 nmol L-1. Applications in DNA samples were examined, and the results for determination of 2′-deoxyguanosine were in good agreement with those obtained by HPLC analysis. Studies on the kinetics of the in vitro consumption of 2′-deoxyguanosine by acetaldehyde were also performed.
Paixao, Thiago R. L. C.,Garcia, Camila C. M.,Medeiros, Marisa H. G.,Bertotti, Mauro
p. 5392 - 5398
(2008/02/10)

C5′-adenosinyl radical cyclization. A stereochemical investigation
A variety of substituted 2′-deoxyadenosin-5′-yl radicals 3 were generated under different reaction conditions. Radicals 3 underwent intramolecular cyclization onto the C8-N7 double bond of the adenine moiety leading to aminyl radicals (5′S,8R)-4 and (5′R,8R)-4 and, eventually, to the corresponding cyclonucleosides 5 and 6. The effect of the solvent, the nature of the substituents, and the generation method of radicals 3 on the stereoselectivity of the C5′-radical cyclization have been considered. The observed increase of the (5′S)/(5′R) ratio by increasing the bulkiness of the R1 group is explained in terms of steric repulsion between R1 and the purine moiety which favors the C5′-endo conformation, whereas the effect of the water solvent in promoting the (5′R)-stereoselective cyclization is ascribed to intermolecular hydrogen bonding stabilizing the C5′-exo confomation.
Navacchia, Maria Luisa,Chatsilialoglu, Chryssostomos,Montevecchi, Pier Carlo
p. 4445 - 4452
(2007/10/03)

Selective removal of the 2′- and 3′-0-acyl groups from 2′,3′,5′-tri-O-acylribonucleoside derivatives with lithium trifluoroethoxide
Selective cleavage of O2′ and O3′ ester groups from ribonucleoside derivatives has been accomplished with Dowex 1 × 2 (CF 3CH2O-) in 2,2,2-trifluoroethanol (TFE) or lithium trifluoroethoxide/TFE. Deacylations with Li+ -OCH2CF3/TFE proceed at ambient temperature (or with mild heating) to give the 5′-O-acyl derivatives in superior yields and higher purity than prior approaches for selective O2′ and O3′ ester deprotection.
Nowak, Ireneusz,Jones, Carl T.,Robins, Morris J.
p. 3077 - 3081
(2007/10/03)

A new protecting group for the exocyclic amino groups of nucleosides
A new protecting group has been developed for the exocyclic amino groups of nucleosides that occur in DNA. 3-Phenyl-[{N-(2-trimethylsilyl-ethoxycarbonyl)- 2-amino}]-propanoic acid used as the protective agent.
Varaprasad, Chamakura V.N.S.,Johnson, Francis
p. 2163 - 2165
(2007/10/03)

Two-color two-laser DNA damaging.
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Kawai, Kiyohiko,Cai, Xichen,Sugimoto, Akira,Tojo, Sachiko,Fujitsuka, Mamoru,Majima, Tetsuro
p. 2406 - 2409
(2007/10/03)

Resistance towards exonucleases of dinucleotides with stereochemically altered internucleotide phosphate bonds
Kinetic constants for the hydrolytic susceptibility of the internucleotide phosphate bond in normal dinucleotides [e.g., 2′-deoxycytidylyl- (3′>5′)-2′-deoxyuridine (dCpdU) and 2′-deoxyadenylyl- (3′→5′)-2′-deoxycytidine (dApdC)] and isomeric dinucleotides
Nair, Vasu,Pal, Suresh
p. 289 - 291
(2007/10/03)

Preparation of deoxynucleosides
Methods for preparing deoxynucleosides from their corresponding ribonucleosides by forming 3-tert-butylphenoxythiocarbonylderivatives of the ribonucleosides and subsequently effecting radical deoxygenation reactions at the carbon atoms to be deoxygenated.
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(2008/06/13)