- Preparation of carbocyclic phosphonate nucleosides
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A versatile and high-yielding synthesis of racemic carbocyclic phosphonate nucleosides of adenine, hypoxanthine, guanine, cytosine, uracil, and thymine has been developed. These newly prepared compounds are isosteric (and isoelectronic) with (carbo)cyclic 2′,3′-dideoxy- and 2′,3′-dideoxy-2′,3′-didehydronucleoside monophosphates.
- Jaehne, Gerhard,Mueller, Armin,Kroha, Herbert,Roesner, Manfred,Holzhaeuser, Otto,Meichsner, Christoph,Helsberg, Matthias,Winkler, Irvin,Riess, Guenther
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Read Online
- Biomimetic Iron Complex Achieves TET Enzyme Reactivity**
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The epigenetic marker 5-methyl-2′-deoxycytidine (5mdC) is the most prevalent modification to DNA. It is removed inter alia via an active demethylation pathway: oxidation by Ten-Eleven Translocation 5-methyl cytosine dioxygenase (TET) and subsequent removal via base excision repair or direct demodification. Recently, we have shown that the synthetic iron(IV)-oxo complex [FeIV(O)(Py5Me2H)]2+ (1) can serve as a biomimetic model for TET by oxidizing the nucleobase 5-methyl cytosine (5mC) to its natural metabolites. In this work, we demonstrate that nucleosides and even short oligonucleotide strands can also serve as substrates, using a range of HPLC and MS techniques. We found that the 5-position of 5mC is oxidized preferably by 1, with side reactions occurring only at the strand ends of the used oligonucleotides. A detailed study of the reactivity of 1 towards nucleosides confirms our results; that oxidation of the anomeric center (1′) is the most common side reaction.
- Carell, Thomas,Daumann, Lena J.,Jonasson, Niko S. W.,Korytiaková, Eva,Schmidl, David
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supporting information
p. 21457 - 21463
(2021/08/23)
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- Excision of 5-Carboxylcytosine by Thymine DNA Glycosylase
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5-Methylcytosine (mC) is an epigenetic mark that is written by methyltransferases, erased through passive and active mechanisms, and impacts transcription, development, diseases including cancer, and aging. Active DNA demethylation involves TET-mediated stepwise oxidation of mC to 5-hydroxymethylcytosine, 5-formylcytosine (fC), or 5-carboxylcytosine (caC), excision of fC or caC by thymine DNA glycosylase (TDG), and subsequent base excision repair. Many elements of this essential process are poorly defined, including TDG excision of caC. To address this problem, we solved high-resolution structures of human TDG bound to DNA with cadC (5-carboxyl-2′-deoxycytidine) flipped into its active site. The structures unveil detailed enzyme-substrate interactions that mediate recognition and removal of caC, many involving water molecules. Importantly, two water molecules contact a carboxylate oxygen of caC and are poised to facilitate acid-catalyzed caC excision. Moreover, a substrate-dependent conformational change in TDG modulates the hydrogen bond interactions for one of these waters, enabling productive interaction with caC. An Asn residue (N191) that is critical for caC excision is found to contact N3 and N4 of caC, suggesting a mechanism for acid-catalyzed base excision that features an N3-protonated form of caC but would be ineffective for C, mC, or hmC. We also investigated another Asn residue (N140) that is catalytically essential and strictly conserved in the TDG-MUG enzyme family. A structure of N140A-TDG bound to cadC DNA provides the first high-resolution insight into how enzyme-substrate interactions, including water molecules, are impacted by depleting the conserved Asn, informing its role in binding and addition of the nucleophilic water molecule.
- Pidugu, Lakshmi S.,Dai, Qing,Malik, Shuja S.,Pozharski, Edwin,Drohat, Alexander C.
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p. 18851 - 18861
(2019/12/11)
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- Opened-ring adducts of 5-methylcytosine and 1,5-dimethylcytosine with amines and water and evidence for an opened-ring hydrate of 2′- deoxycytidine
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A variety of nucleic acid components and related compounds undergo photoreaction with water to form so-called photohydrates (e.g. uracil forms 6-hydroxy-5,6-dihydrouracil). However, the corresponding hydrates of 5-methylcytosine (a minor nucleobase in eukaryotic DNA) and related compounds have not been characterized. We report the preparation of opened-ring forms of such products for 5-methylcytosine (m5C) and 1,5-dimethylcytosine (DMC). This was accomplished via thermal reaction of ring-opened amine adducts (e.g. N-carbamoyl-3-amino-2-methylacrylamidine (IVa) or N-(N′-methylcarbamoyl)- 3-amino-2-methylacrylamidine (IVb)) produced by photo-induced reactions of m5C with ammonia or methylamine. When these adducts were treated with dilute trifluoroacetic acid, the amino group at the 3-position was replaced with a hydroxyl group; with IVa, N-carbamoyl-3-hydroxy-2-methylacrylamidine (Va) was formed, while reaction of IVb led to N-(N′-methylcarbamoyl)-3-hydroxy-2- methylacrylamidine (Vb). These compounds are ring-opened isomers of 5,6-dihydro-6-hydroxy-5-methylcytosine (Ia and IIa) and 5,6-dihydro-6-hydroxy-1, 5-dimethylcytosine (Ib and IIb). Compounds Va and Vb each undergo thermal ring closure reactions to form two unstable compounds with chemical and UV spectral properties expected for Ia and IIa (or Ib and IIb). The latter compounds have been identified as minor products in UV-irradiated aqueous solutions of m5C and DMC. Evidence is also presented that the 2′-deoxycytidine photohydrates coexist with an opened-ring form, possibly similar in nature to Vb. The nucleobase 5-methylcytosine (m5C) reacts photochemically with ammonia to form an opened-ring adduct (IVa); similar reactions occur with 1,5-dimethylcytosine and with methylamine. Subjection of such adducts to hydrolysis in dilute acid (e.g. 0.1% trifluoracetic acid) produces opened-ring hydrates of m5C (Va) and DMC. Photoreaction of the nucleoside 2′-deoxycytidine in water results in formation of closed-ring hydrates that equilibrate thermally with a compound that has the UV spectral properties expected for an adduct similar in nature to Va.
- Shetlar, Martin D.,Chung, Janet
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experimental part
p. 818 - 832
(2012/07/27)
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- 2′-O-aminoethyl oligoribonucleotides containing novel base analogues: Synthesis and triple-helix formation at pyrimidine/purine inversion sites
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The synthesis of a common sugar intermediate for the 2′-aminoethyl- ribonucleoside synthesis in 9 steps and an overall yield of 33 % starting from D-ribose is described. This intermediate was successfully used for the synthesis of the 2′-aminoethyl-ribonucleosides containing the bases thymine (t), 5-methylcytosine (c), 5-methyl-2-pyrimidinone (x), 2-aminopurine (ap) and guanine (g). These were subsequently transformed into the corresponding cyanoethyl phosphoramidite building blocks for oligonucleotide synthesis. 2′-Aminoethyl oligonucleotide 15-mers were prepared with a DNA synthesizer, and an optimized post-synthetic deprotection protocol has been developed which prevents cyanoethylation of the 2′-amino side chains during conventional ammonia deprotection. A series of fully modified, triplex forming 2′-aminoethyl oligoribonucleotides (2′AE-TFOs) were prepared in which x was designed to bind to CG inversion sites and ap as well as g to TA inversion sites on a double-helical DNA target. The affinity of x-CG base-triple formation in different sequence contexts was assessed by UV- and CD melting analysis. It was found that TFO 15-mers containing up to 5 x residues still form stable triplexes even in the case where all x residues are consecutively arranged in the TFO. The nearest neighbor properties of x have been probed and it was found that triplex stability decreases in the local sequence order -txt- > -txc- ? -cxc-. TFOs containing ap and g were found to bind to their DNA targets with TA inversion sites with less affinity and less selectivity compared to TFOs containing the corresponding deoxyribonucleosides, irrespective whether they were incorporated in TFOs with a DNA or a 2′-AE-RNA backbone. The obtained data suggest that guanine-TA or aminopurine-TA base-triple formation is strongly sensitive to TFO conformation and more efficient in TFOs with a DNA than an RNA backbone. Wiley-VCH Verlag GmbH & Co. KGaA, 2006.
- Buchini, Sabrina,Leumann, Christian J.
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p. 3152 - 3168
(2007/10/03)
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- Compounds and methods for the characterization of oligonucleotides
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The present invention relates to oligonucleotide synthesis. In particular, the present invention provides methods for characterizing samples useful for making oligonucleotides.
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- Cycloalkltriols containing cyclic substituents, processes and intermediate products for their preparation and their use as antiviral and antiparasitic agents
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Cycloalkyltriols containing heterocyclic substituents, in particular cyclopentyl- and cyclohexyltriols containing heterocyclic substituents Compounds of the formulae I and II STR1 in which the substituents A, R1, R2 and R3 and n have the meanings given, have an antiviral and antiparasitic action.
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- Preparation of oligonucleotides containing 5-bromouracil and 5-methylcytidine
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A previously described side reaction on 5-bromouracil during standard oligonucleotide deprotection conditions has been studied in detail. The side product, 5-amino-2′-deoxyuridine, is isolated and characterized. The use of several 5-methylcytidine protected derivatives for the preparation of oligonucleotides containing 5-bromouracil and 5-methylcytidine free of 5-amino-2′-deoxyuridine is discussed.
- Ferrer, Elisenda,Fabrega, Carme,Garcia, Ramon Gueimil,Azorin, Ferran,Eritja, Ramon
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p. 907 - 921
(2007/10/03)
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- Process for the preparation of 5-substituted cytosines and other 4,5-disubstituted pyrimidin-2(1H)-ones, and intermediates arising in the course of this
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Process for the preparation of 5-substituted cytosines and other 4,5-disubstituted pyrimidin-2(1H)-ones, and intermediates arising in the course of this The process for the preparation of compounds of the formula I, STR1 wherein compounds of the formula II STR2 are converted into an azole derivative of the formula III or an azine derivative of the formula IV STR3 which can subsequently be converted into a compound of the formula I using a nucleophile XH.
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- Oxidation of Cytosine and 5-Methylcytosine Nucleosides and 5-Methyl-2'-deoxycytidine 5'-Monophosphate with Peroxosulfate Ions
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Reaction of 5-methylcytosine nucleosides and nucleotide with Na2S2O8 resulted in an oxidation of the 5-methyl group, while treatment of them and cytosine nucleosides with KHSO5 gave the corresponding N3-oxides.
- Itahara, Toshio
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p. 1591 - 1594
(2007/10/02)
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- Substituent effects on degradation rates and pathways of cytosine nucleosides.
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A previous report on the influence of a 6-methyl substituent on cytosine nucleoside degradation proposed that N-glycosyl hydrolysis predominated over the deamination pathway which was characteristic of the unsubstituted parent compounds. The UV absorption data which led to this hypothesis were not conclusive. Evidence for N-glycosyl hydrolysis was indirect and the product concentration was not quantitated. In the present study, specific HPLC methods were employed to assay four cytosine nucleosides and their corresponding bases, thus allowing comparison of the N-glycosyl hydrolysis rate to the overall rate of loss for each nucleoside. These data indicated that the 6-methyl nucleosides underwent partial or complete hydrolysis to yield their corresponding sugars and 6-methylcytosine, which then deaminated to 6-methyluracil. An increase in the reactivity and a change in the reaction products of the 6-methyl nucleosides were attributed to an alteration in conformation. In addition, the 6-methyl arabinosyl nucleoside reacted much faster than the 6-methyl ribosyl nucleoside, presumably due to 2'-OH participation. Degradation of 5-methyl deoxycytidine was also re-examined since its degradation was previously attributed solely to N-glycosyl hydrolysis. In the present study, simultaneous deamination and hydrolysis were measured, although N-glycosyl hydrolysis was found to predominate.
- Nguyen,Notari
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p. 802 - 806
(2007/10/02)
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- SYNTHESIS OF N-(2-PHOSPHONYLMETHOXYETHYL) DERIVATIVES OF HETEROCYCLIC BASES
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The preparation of N-(2-phosphonylmethoxyethyl) derivatives of purine and pyrimidine bases, IV, as analogs of the antiviral 9-(2-phosphonylmethoxyethyl)adenine (PMEA,I), is described.The synthesis consists in alkylation of alkali metal salts of heterocyclic bases or their N- or O-substituted derivatives with diethyl 2-p-toluenesulfonyloxyethoxymethylphosphonate (IIa), 2-chloroethoxymethylphosphonate (IIb) or 2-bromoethoxymethylphosphonate (IIc).The obtained N-(2-diethoxyphosphonylmethoxyethyl) derivatives of heterocyclic bases (III) were treated with bromotrimethylsilane to give phosphonic acids IV.Compounds IV were prepared from pyrimidines (uracil, cytosine and their 5-methyl derivatives), purines (adenine and its N6- and C(2)-substituted derivatives, hypoxanthine, guanine, 6-hydrazinopurine and 6-methylthiopurine etc.) and their analogs (3-deazaadenine etc.).
- Holy, Antonin,Rosenberg, Ivan,Dvorakova, Hana
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p. 2190 - 2210
(2007/10/02)
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- Nucleoside process
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Process for producing 1-halo-2-deoxy-2-fluoroarabinofuranoside derivatives bearing protective ester groups from 1,3,5-tri-O-acylribofuranose; the 1-halo compounds are intermediates in the synthesis of therapeutically active nucleosidic compounds.
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