7005-15-4Relevant academic research and scientific papers
Cleavage of oligodeoxyribonucleotides from polymer supports and their rapid deprotection under microwaves
Gupta,Kumar
, p. 1761 - 1766 (1998)
Novel conditions for the cleavage of oligodeoxynucleotides from polymer supports and their complete deprotection under microwaves have been developed. The oligonucleotides synthesized using phosphoramidite synthons carrying base labile (Pac, Dmf and t-Bpac) and conventional (Bz for A and C and Pac for G) protecting groups for nucleic bases were deprotected using 0.2M sodium hydroxide (MeOH : H2O :: 1:1, v/v) = Reagent A and 1M sodium hydroxide (MeOH : H2O :: 1:1, v/v) = Reagent B, respectively under microwaves. The deprotected oligonucleotides were found to be comparable with the corresponding oligonucleotides deprotected under standard conditions (aq. ammonia at 55°C).
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
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)
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.
Mechanisms of allosteric activation and inhibition of the deoxyribonucleoside triphosphate triphosphohydrolase from Enterococcus faecalis
Vorontsov, Ivan I.,Wu, Ying,Delucia, Maria,Minasov, George,Mehrens, Jennifer,Shuvalova, Ludmilla,Anderson, Wayne F.,Ahn, Jinwoo
, p. 2815 - 2824 (2014)
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.
Meteorite-catalyzed intermoleculartrans-glycosylation produces nucleosides under proton beam irradiation
Bizzarri, Bruno Mattia,Fanelli, Angelica,Kapralov, Michail,Krasavin, Eugene,Saladino, Raffaele
, p. 19258 - 19264 (2021/06/03)
Di-glycosylated adenines act as glycosyl donors in the intermoleculartrans-glycosylation of pyrimidine nucleobases under proton beam irradiation conditions. Formamide and chondrite meteorite NWA 1465 increased the yield and the selectivity of the reaction
Prebiotic Photochemical Coproduction of Purine Ribo- And Deoxyribonucleosides
Xu, Jianfeng,Green, Nicholas J.,Russell, David A.,Liu, Ziwei,Sutherland, John D.
supporting information, p. 14482 - 14486 (2021/09/18)
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.
An enzymatic flow-based preparative route to vidarabine
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)
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.
RETRACTED ARTICLE: Convenient synthesis of pyrimidine 2′-deoxyribonucleoside monophosphates with important epigenetic marks at the 5-position
Zheng, Song,Tran, Ai,Curry, Alyson M.,White, Dawanna S.,Cen, Yana
, p. 5164 - 5173 (2020/07/23)
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
Thermodynamic Reaction Control of Nucleoside Phosphorolysis
Kaspar, Felix,Giessmann, Robert T.,Neubauer, Peter,Wagner, Anke,Gimpel, Matthias
, p. 867 - 876 (2020/01/24)
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.).
Bio-catalytic synthesis of unnatural nucleosides possessing a large functional group such as a fluorescent molecule by purine nucleoside phosphorylase
Hatano, Akihiko,Wakana, Hiroyuki,Terado, Nanae,Kojima, Aoi,Nishioka, Chisato,Iizuka, Yu,Imaizumi, Takuya,Uehara, Sanae
, p. 5122 - 5129 (2019/10/05)
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.
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
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)
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.
