65-71-4Relevant articles and documents
Photoinduced deoxyribose C2′ oxidation in DNA. Alkali-dependent cleavage of erythrose-containing sites via a retroaldol reaction
Sugiyama, Hiroshi,Tsutsumi, Yasushi,Fujimoto, Kenzo,Saito, Isao
, p. 4443 - 4448 (1993)
Photoreactions of various 5-iodouracil- (1U)-containing oligonucleotides have been investigated. It was found that d(GCA1UGC)2 undergoes a competitive Cl′ and C2′ oxidation at the 5′ side of the 1U residue to give deoxyribonolactone-containing hexamer 1 and erythrose-containing hexamer 2, respectively. Upon heating under alkaline conditions, erythrose-containing hexamer 2 was shown to undergo a remarkably facile retroaldol reaction to give two fragments, both having phosphoroglycoaldehyde termini in high yields. On the basis of the chemical reactivity of 2, a new specific method for detection of the erythrose-containing sites resulting from deoxyribose C2′ oxidation in DNA was devised. Erythrose-containing sites were prepared by photoirradiation of duplex 1U-containing 13 mer 5′-d(CG1UGT'UTA1UAC 1UG)-3′/5′-d(CAGTATAAACACG)-3′. After photoirradiation, the reaction mixture was treated with hot alkali and NaBH4 and then subjected to enzymatic digestion. HPLC analysis of the digested mixture revealed the formation of modified mononucleotides 25-28, allowing the quantification of the erythrose-containing sites being produced at the 5′ side of all four 1U residues of the 13 mer. These results indicate that this method can be used for the detection and quantification of erythrose-containing sites resulting from deoxyribose C2′ oxidation in DNA.
The Peculiar Case of the Hyper-thermostable Pyrimidine Nucleoside Phosphorylase from Thermus thermophilus**
Kaspar, Felix,Neubauer, Peter,Kurreck, Anke
, p. 1385 - 1390 (2021/01/29)
The poor solubility of many nucleosides and nucleobases in aqueous solution demands harsh reaction conditions (base, heat, cosolvent) in nucleoside phosphorylase-catalyzed processes to facilitate substrate loading beyond the low millimolar range. This, in turn, requires enzymes that can withstand these conditions. Herein, we report that the pyrimidine nucleoside phosphorylase from Thermus thermophilus is active over an exceptionally broad pH (4–10), temperature (up to 100 °C) and cosolvent space (up to 80 % (v/v) nonaqueous medium), and displays tremendous stability under harsh reaction conditions with predicted total turnover numbers of more than 106 for various pyrimidine nucleosides. However, its use as a biocatalyst for preparative applications is critically limited due to its inhibition by nucleobases at low concentrations, which is unprecedented among nonspecific pyrimidine nucleoside phosphorylases.
Efficient biocatalytic synthesis of dihalogenated purine nucleoside analogues applying thermodynamic calculations
Giessmann, Robert T.,Kaspar, Felix,Klare, Hendrik F. T.,Kurreck, Jens,Neubauer, Peter,Paulick, Katharina,R?hrs, Viola,Wagner, Anke,Westarp, Sarah,Yehia, Heba
, (2020/02/28)
The enzymatic synthesis of nucleoside analogues has been shown to be a sustainable and efficient alternative to chemical synthesis routes. In this study, dihalogenated nucleoside analogues were produced by thermostable nucleoside phosphorylases in transglycosylation reactions using uridine or thymidine as sugar donors. Prior to the enzymatic process, ideal maximum product yields were calculated after the determination of equilibrium constants through monitoring the equilibrium conversion in analytical-scale reactions. Equilibrium constants for dihalogenated nucleosides were comparable to known purine nucleosides, ranging between 0.071 and 0.081. To achieve 90% product yield in the enzymatic process, an approximately five-fold excess of sugar donor was needed. Nucleoside analogues were purified by semi-preparative HPLC, and yields of purified product were approximately 50% for all target compounds. To evaluate the impact of halogen atoms in positions 2 and 6 on the antiproliferative activity in leukemic cell lines, the cytotoxic potential of dihalogenated nucleoside analogues was studied in the leukemic cell line HL-60. Interestingly, the inhibition of HL-60 cells with dihalogenated nucleoside analogues was substantially lower than with monohalogenated cladribine, which is known to show high antiproliferative activity. Taken together, we demonstrate that thermodynamic calculations and small-scale experiments can be used to produce nucleoside analogues with high yields and purity on larger scales. The procedure can be used for the generation of new libraries of nucleoside analogues for screening experiments or to replace the chemical synthesis routes of marketed nucleoside drugs by enzymatic processes.
Thermodynamic Reaction Control of Nucleoside Phosphorolysis
Kaspar, Felix,Giessmann, Robert T.,Neubauer, Peter,Wagner, Anke,Gimpel, Matthias
supporting information, 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.).
Anion exchange resins in phosphate form as versatile carriers for the reactions catalyzed by nucleoside phosphorylases
Artsemyeva, Julia N.,Buravskaya, Tatiana N.,Esipov, Roman S.,Konstantinova, Irina D.,Litvinko, Natalia M.,Mikhailopulo, Igor A.,Miroshnikov, Anatoly I.,Remeeva, Ekaterina A.
, p. 2607 - 2622 (2020/11/26)
In the present work, we suggested anion exchange resins in the phosphate form as a source of phosphate, one of the substrates of the phosphorolysis of uridine, thymidine, and 1-(β-D-arabinofuranosyl)uracil (Ara-U) catalyzed by recombinant E. coli uridine (UP) and thymidine (TP) phosphorylases. α-D-Pentofuranose-1-phosphates (PF-1Pis) obtained by phosphorolysis were used in the enzymatic synthesis of nucleosides. It was found that phosphorolysis of uridine, thymidine, and Ara-U in the presence of Dowex 1X8 (phosphate; Dowex-nPi) proceeded smoothly in the presence of magnesium cations in water at 20-50 °C for 54-96 h giving rise to quantitative formation of the corresponding pyrimidine bases and PF-1Pis. The resulting PF-1Pis can be used in three routes: (1) preparation of barium salts of PF-1Pis, (2) synthesis of nucleosides by reacting the crude PF-1Pi with an heterocyclic base, and (3) synthesis of nucleosides by reacting the ionically bound PF-1Pi to the resin with an heterocyclic base. These three approaches were tested in the synthesis of nelarabine, kinetin riboside, and cladribine with good to excellent yields (52-93%).
Sources of 2,5-diaminoimidazolone lesions in DNA damage initiated by hydroxyl radical attack
Thomas, Caroline Suzanne,Pollard, Hannah Catherine,Razskazovskiy, Yuriy,Roginskaya, Marina
, p. 517 - 524 (2020/09/07)
The present study reports radiation-chemical yields of 2.5-diaminoimidazolone (Iz) derivatives in X-irradiated phosphate-buffered solutions of guanosine and double-stranded DNA. Various gassing conditions (air, N20/O2 (4:1), N2O, vacuum) were employed to elucidate the contribution of several alternative pathways leading to Iz in reactions initiated by hydroxyl radical attack on guanine. In all systems, Iz was identified as the second by abundance guanine degradation product after 8-oxoguanine, formed in 1:5 (guanosine) and 1:3.3 (DNA) ratio to the latter in air-saturated solutions. Experimental data strongly suggest that the addition of molecular oxygen to the neutral guanine radical G(-H)? plays a major in Iz production in oxygenated solutions of double-stranded DNA while in other systems it may compete with recombination of G(-H)? with superoxide and/or alkyl peroxyl radicals. The production of Iz through hydroxyl radical attack on 8-oxoguanine was also shown to take place although the chemical yield of Iz (ca 6%) in this process is too low to compete with the other pathways. The linearity of Iz accumulation with dose also indicates a negligible contribution of this channel to its yield in all systems.
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.
Biomimetic Oxidative Deamination Catalysis via ortho-Naphthoquinone-Catalyzed Aerobic Oxidation Strategy
Golime, Gangadhararao,Bogonda, Ganganna,Kim, Hun Young,Oh, Kyungsoo
, p. 4986 - 4990 (2018/05/14)
An ortho-naphthoquinone-catalyzed oxidative deamination reaction has been developed where the molecular oxygen and water serve as the sole oxidant and nucleophile. The current aerobic deamination reaction proceeds via the ketimine formation between ortho-naphthoquinones and amines followed by the prototropic rearrangement and hydrolysis by water, representing a biomimetic oxidative deamination of amine species in the human body by the liver and kidneys. The compatibility of ortho-naphthoquinone organocatalysts with molecular oxygen and water opens up a new biomimetic catalyst system that can function as versatile deaminases for a variety of amine-containing molecules such as amino acids and DNA nuclear bases.
Methyl ketone derivative, and preparation method and applications thereof
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Paragraph 0039, (2017/08/28)
The invention discloses a methyl ketone derivative, and a preparation method and applications thereof. The preparation method comprises following steps: a ketone derivative and an organic peroxide are dissolved in a solvent, and reaction is carried out at 80 to 130 DEG C so as to obtain methyl pyrimidone and a methyl pyrimidone derivative. According to the preparation method, the ketone derivative is taken as a starting material; the raw materials are easily and widely available; products of different kinds can be obtained via the preparation method, and can be used directly or used in other further reaction. No metal is involved, so that the preparation method is suitable to be applied in pharmaceutical preparation technology. The preparation method is short in route, mild in reaction conditions, simple in reaction operation and postprocessing process, and high in yield, and is suitable for large-scale production.
Substrate specificity of E. coli uridine phosphorylase. Further evidences of high-syn conformation of the substrate in uridine phosphorolysis
Alexeev,Sivets,Safonova,Mikhailov
, p. 107 - 121 (2017/02/05)
Twenty five uridine analogues have been tested and compared with uridine with respect to their potency to bind to E. coli uridine phosphorylase. The kinetic constants of the phosphorolysis reaction of uridine derivatives modified at 2′-, 3′- and 5′-positions of the sugar moiety and 2-, 4-, 5- and 6-positions of the heterocyclic base were determined. The absence of the 2′- or 5′-hydroxyl group is not crucial for the successful binding and phosphorolysis. On the other hand, the absence of both the 2′- and 5′-hydroxyl groups leads to the loss of substrate binding to the enzyme. The same effect was observed when the 3′-hydroxyl group is absent, thus underlining the key role of this group. Our data shed some light on the mechanism of ribo- and 2′-deoxyribonucleoside discrimination by E. coli uridine phosphorylase and E. coli thymidine phosphorylase. A comparison of the kinetic results obtained in the present study with the available X-ray structures and analysis of hydrogen bonding in the enzyme-substrate complex demonstrates that uridine adopts an unusual high-syn conformation in the active site of uridine phosphorylase.
