WorldWideScience

Sample records for base-functionalized carbocyclic nucleosides

  1. Synthesis and Antiviral Activity of Novel 4'-Branched Carbocyclic C-Nucleoside

    Energy Technology Data Exchange (ETDEWEB)

    Jin, Ying Lan; Hong, Joon Hee [Chosun University, Gwangju (Korea, Republic of)

    2005-09-15

    C-Nucleosides have received considerable attention due not only to the chemical stability but also to the interesting biological activities of naturally occurring compounds such as showdomycin, formycins, oxazinomycin, etc. Also, several biologically active synthetic C-nucleosides such as pseudoisocytidine, thiazofurin, and 9-deazaadenosine have been reported. Carbocyclic nucleosides are another class of metabolically stable nucleosides in which a methylene group replaces the oxygen in the furan ring of the natural nucleosides. Another interesting feature of carbocyclic nucleosides is that a number of carbocyclic adenosine analogues are assumed to exert their antiviral action via the inhibition of S-adenosyl-homocysteine hydrolase. Moreover, this mechanism might be exploited in a combination therapy in association with the nucleosides with a different mechanism of action. On the basis of these interesting chemical and biological properties of C-nucleosides and carbocyclic nucleosides, it was of interest to synthesize hybrid nucleosides, carbocyclic C-nucleosides. Although some carbocyclic and C-nucleosides are naturally occurring, so far no natural carbocyclic C-nucleosides have been reported. The history of synthesis of carbocyclic C-nucleosides dates back to the 1960s. Despite the long history of carbocyclic C-nucleosides, only a few carbocyclic C-nucleosides have been synthesized, probably due to the synthetic difficulties of these nucleosides. Herein, we would like to report the synthesis procedure of a novel 4'-branched carbocyclic C-nucleoside.

  2. Two convergent approaches toward novel carbocyclic C-nucleosides

    Czech Academy of Sciences Publication Activity Database

    Nencka, Radim; Šála, Michal; Dejmek, Milan; Dračínský, Martin; Holý, Antonín; Hřebabecký, Hubert

    -, č. 23 (2010), s. 4119-4130 ISSN 0039-7881 R&D Projects: GA MŠk 1M0508 Institutional research plan: CEZ:AV0Z40550506 Keywords : carbocyclic C-nucleosides * convergent approach * uracil * pyrimidines Subject RIV: CC - Organic Chemistry Impact factor: 2.260, year: 2010

  3. Synthesis of carbocyclic homo-N-nucleosides from iridoids

    DEFF Research Database (Denmark)

    Franzyk, Henrik; Rasmussen, Jon Holbech; Mazzei, Rafaele Antonio

    1998-01-01

    Two iridoid glucosides, antirrhinoside (1) and catalpol (2), were converted into selectively protected polysubstituted cyclopentylmethanols, which were subsequently used to prepare carbocyclic homo-N-nucleosides (5, 6 and 14). A purine moiety was introduced either by the Mitsunobu reaction...... or by substitution of a primary triflate with the tetrabutylammonium salt of 6-iodopurine. The latter method was superior with regard to both ease of purification and yield. The N-9 vs. N-7 regioselectivity of the salts of different 6-substituted purine derivatives was briefly investigated....

  4. Revaluation of biomass-derived furfuryl alcohol derivatives for the synthesis of carbocyclic nucleoside phosphonate analogues

    OpenAIRE

    Sidi Mohamed, Bemba; P?rigaud, Christian; Math?, Christophe

    2017-01-01

    The racemic synthesis of new carbocyclic nucleoside methylphosphonate analogues bearing purine bases (adenine and guanine) was accomplished using bio-sourced furfuryl alcohol derivatives. All compounds were prepared using a Mitsunobu coupling between the heterocyclic base and an appropriate carbocyclic precursor. After deprotection, the compounds were evaluated for their activity against a large number of viruses. However, none of them showed significant antiviral activity or cytotoxicity.

  5. Synthesis of novel racemic carbocyclic nucleosides derived from 5,6-disubstituted norbornene

    Czech Academy of Sciences Publication Activity Database

    Šála, Michal; Hřebabecký, Hubert; Dračínský, Martin; Masojídková, Milena; De Palma, A. M.; Neyts, J.; Holý, Antonín

    2010-01-01

    Roč. 75, č. 1 (2010), s. 1-20 ISSN 0010-0765 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : carbocyclic nucleosides * norbornenes * Coxsackie virus Subject RIV: CC - Organic Chemistry Impact factor: 0.853, year: 2010

  6. Synthesis of novel carbocyclic nucleoside analogues derived from 7-oxabicyclo[2.2.1]heptane-2-methanol

    Czech Academy of Sciences Publication Activity Database

    Hřebabecký, Hubert; Dračínský, Martin; De Palma, A.; Neyts, J.; Holý, Antonín

    2009-01-01

    Roč. 74, č. 3 (2009), s. 487-502 ISSN 0010-0765 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : carbocyclic nucleosides * purines * thymine Subject RIV: CC - Organic Chemistry Impact factor: 0.856, year: 2009

  7. New carbocyclic nucleoside analogues built on a bicyclo[2.2.2]octane-2,2-dimethanol template

    Czech Academy of Sciences Publication Activity Database

    Hřebabecký, Hubert; Dračínský, Martin; Holý, Antonín

    2009-01-01

    Roč. 74, č. 9 (2009), s. 1425-1441 ISSN 0010-0765 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : carbocyclic nucleosides * purines * Mitsunobu reaction Subject RIV: CC - Organic Chemistry Impact factor: 0.856, year: 2009

  8. Synthesis of novel carbocyclic nucleoside analogues containing bicyclo[2.2.1]hept-2-ene-2-methanol

    Czech Academy of Sciences Publication Activity Database

    Hřebabecký, Hubert; Dračínský, Martin; Holý, Antonín

    2008-01-01

    Roč. 73, č. 1 (2008), s. 44-58 ISSN 0010-0765 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501; GA ČR GA203/05/0132 Institutional research plan: CEZ:AV0Z40550506 Keywords : carbocyclic nucleosides * purines * norbornenes Subject RIV: CC - Organic Chemistry Impact factor: 0.784, year: 2008

  9. Synthesis of novel carbocyclic nucleosides and Pro-Tides derived from 4-oxatricyclo[4.2.1.03,7]nonane-9-methanol

    Czech Academy of Sciences Publication Activity Database

    Hřebabecký, Hubert; Dračínský, Martin; Holý, Antonín

    2007-01-01

    Roč. 72, č. 10 (2007), s. 1331-1349 ISSN 0010-0765 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : carbocyclic nucleosides * purines * thymine Subject RIV: CC - Organic Chemistry Impact factor: 0.879, year: 2007

  10. Synthesis of novel conformationally locked carbocyclic nucleosides derived from 3-(hydroxymethyl)bicyclo[2.2.1]heptane-2,5-diol

    Czech Academy of Sciences Publication Activity Database

    Hřebabecký, Hubert; Masojídková, Milena; Dračínský, Martin; Holý, Antonín

    2006-01-01

    Roč. 71, č. 6 (2006), s. 871-888 ISSN 0010-0765 R&D Projects: GA MŠk(CZ) 1M0508; GA ČR(CZ) GA203/05/0132 Institutional research plan: CEZ:AV0Z40550506 Keywords : azides * amines * cyclic sulfates * carbocyclic nucleosides Subject RIV: CC - Organic Chemistry Impact factor: 0.881, year: 2006

  11. Synthesis of novel racemic carbocyclic nucleoside analogues derived from 4,8-dioxatricyclo[4.2.1.0 3,7]nonane-9-methanol and 4-oxatricyclo[4.3.1.0 3,7]decane-10-methanol, compounds with activity against Coxsackie viruses

    Czech Academy of Sciences Publication Activity Database

    Hřebabecký, Hubert; Dračínský, Martin; De Palma, A.; Neyts, J.; Holý, Antonín

    2009-01-01

    Roč. 74, č. 3 (2009), s. 469-485 ISSN 0010-0765 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : amines * carbocyclic nucleosides * purines Subject RIV: CC - Organic Chemistry Impact factor: 0.856, year: 2009

  12. Cross-coupling reactions of nucleoside triphosphates followed by polymerase incorporation. Construction and applications of base-functionalized nucleic acids.

    Science.gov (United States)

    Hocek, Michal; Fojta, Miroslav

    2008-07-07

    Construction of functionalized nucleic acids (DNA or RNA) via polymerase incorporation of modified nucleoside triphosphates is reviewed and selected applications of the modified nucleic acids are highlighted. The classical multistep approach for the synthesis of modified NTPs by triphosphorylation of modified nucleosides is compared to the novel approach consisting of direct aqueous cross-coupling reactions of unprotected halogenated nucleoside triphosphates. The combination of cross-coupling of NTPs with polymerase incorporation gives an efficient and straightforward two-step synthesis of modified nucleic acids. Primer extension using biotinylated templates followed by separation using streptavidine-coated magnetic beads and DNA duplex denaturation is used for preparation of modified single stranded oligonucleotides. Examples of using this approach for electrochemical DNA labelling and bioanalytical applications are given.

  13. Synthesis of racemic 9-(6- and 2,6-substituted 9H-purin-9-yl)-5-oxatricyclo[4.2.1.03,7]nonane-3-methanols, novel conformationally locked carbocyclic nucleosides

    Czech Academy of Sciences Publication Activity Database

    Hřebabecký, Hubert; Masojídková, Milena; Holý, Antonín

    2005-01-01

    Roč. 70, č. 1 (2005), s. 103-123 ISSN 0010-0765 Institutional research plan: CEZ:AV0Z40550506 Keywords : amines * oximes * nucleosides Subject RIV: CC - Organic Chemistry Impact factor: 0.949, year: 2005

  14. Synthesis of a Novel Carbocyclic Analog of Bredinin

    Directory of Open Access Journals (Sweden)

    Vasu Nair

    2013-09-01

    Full Text Available The natural nucleoside antibiotic, bredinin, exhibits antiviral and other biological activities. While various nucleosides related to bredinin have been synthesized, its carbocyclic analog has remained unknown. Synthesis of this heretofore unknown analog of bredinin is described. The key precursor, (3aS,4R,6R,6aR-6-((methoxy-methoxymethyl-2,2-dimethyltetrahydro-3aH-cyclopenta[d][1,3]dioxol-4-amine (5, was prepared from the commercially available compound, (1R,4S-2-azabicyclo[2.2.1] hept-5-en-3-one (4. Our initial approach used intermediate 6, derived in three transformations from 5, for the key photolytic step to produce the desired ring-opened precursor to the target compound. This photochemical transformation was unsuccessful. However, an appropriately protected and related precursor was synthesized from 5 through the following side-chain functional group transformations: elaboration of the amino group through malonyl ester formation, oximation at the central carbon, conversion of ester to amide and catalytic reduction of the oxime group. This precursor, on treatment with triethylorthoformate and catalytic acetic acid in ethanol, underwent cyclization to produce the desired 4-carbamoyl-imidazolium-5-olate ring. Deprotection of the latter product proceeded smoothly to give the carbocyclic analog of bredinin. This target molecule exhibits antiviral activity, albeit low, against a number of RNA viruses. Further biological evaluations are in progress.

  15. Norbornane as the novel pseudoglycone moiety in nucleosides

    Czech Academy of Sciences Publication Activity Database

    Šála, Michal; Hřebabecký, Hubert; Dračínský, Martin; Masojídková, Milena; De Palma, A.; Neyts, J.; Holý, Antonín

    2009-01-01

    Roč. 65, č. 45 (2009), s. 9291-9299 ISSN 0040-4020 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : carbocyclic nucleosides * norbornane * coxsackie virus Subject RIV: CC - Organic Chemistry Impact factor: 3.219, year: 2009

  16. Diastereoselective Flexible Synthesis of Carbocyclic C-Nucleosides

    Czech Academy of Sciences Publication Activity Database

    Maier, L.; Khirsariya, P.; Hylse, O.; Adla, S.K.; Cernova, L.; Poljak, M.; Krajčovičová, S.; Weis, E.; Drápela, Stanislav; Souček, Karel; Paruch, K.

    2017-01-01

    Roč. 82, č. 7 (2017), s. 3382-3402 ISSN 0022-3263 Institutional support: RVO:68081707 Keywords : mitsunobu * catalysis * alcohols Subject RIV: CC - Organic Chemistry OBOR OECD: Organic chemistry Impact factor: 4.849, year: 2016

  17. Norbornane-based nucleoside and nucleotide analogues locked in North conformation

    Czech Academy of Sciences Publication Activity Database

    Dejmek, Milan; Šála, Michal; Hřebabecký, Hubert; Dračínský, Martin; Procházková, Eliška; Chalupská, Dominika; Klíma, Martin; Plačková, Pavla; Hájek, Miroslav; Andrei, G.; Naesens, L.; Leyssen, P.; Neyts, J.; Balzarini, J.; Bouřa, Evžen; Nencka, Radim

    2015-01-01

    Roč. 23, č. 1 (2015), s. 184-191 ISSN 0968-0896 R&D Projects: GA ČR GPP207/12/P625; GA MŠk LO1302 Institutional support: RVO:61388963 Keywords : carbocyclic nucleosides * purines * norbornane * antiviral * PI4KIIalpha Subject RIV: CC - Organic Chemistry Impact factor: 2.923, year: 2015

  18. Stereoselective synthesis of 2'-fluoro-6'-methylene carbocyclic adenosine via Vince lactam.

    Science.gov (United States)

    Singh, Uma S; Mishra, Ram C; Shankar, Ravi; Chu, Chung K

    2014-05-02

    2'-Fluoro-6'-methylene carbocyclic adenosine (FMCA) is a potent and selective inhibitor of wild type as well as drug-resistant hepatitis B virus (HBV) mutants. FMCA demonstrated excellent anti-HBV activity against both adefovir-resistant and lamivudine-resistant double (rtL180M/rtM204V) mutants as well as in lamivudine/entecavir triple mutants (L180M+S202G+M204V) in vitro. Its monophosphate prodrug (FMCAP) demonstrated a greater than 12-fold increase of anti-HBV activity in comparison to that of the nucleoside without elevation of cellular toxicity. In the preliminary in vivo study in chimeric mice harboring the lamivudine/entecavir triple mutant, FMCAP effectively reduced HBV viral load, while entecavir was not effective. Therefore, it was of great interest to develop an efficient synthetic procedure to support the preclinical investigation. In this article, a new approach for the synthesis of FMCA from a readily available starting material (Vince lactam) in 16 steps is described. An efficient and practical methodology for stereospecific preparation of a versatile carbocyclic key intermediate, D-2'-fluoro-6'-methylene cyclopentanol 14, has been developed from diazotization, elimination, stereoselective epoxidation, fluorination, and oxidation-reduction sequence of the Vince lactam in 14 steps. The utility of D-2'-fluoro-6'-methylene cyclopentanol 14 is demonstrated in the preparation of FMCA using the Mitsunobu coupling to introduce the adenine base to synthesize the final nucleoside.

  19. Sulfur ylides in the synthesis of heterocyclic and carbocyclic compounds

    International Nuclear Information System (INIS)

    Lakeev, Sergei N; Maydanova, Irina O; Galin, Fanur Z; Tolstikov, Genrikh A

    2001-01-01

    Data on the use of sulfonium ylides in the synthesis of carbocyclic and heterocyclic compounds published over the last 15 years are analysed, systematised and generalised. The bibliography includes 139 references.

  20. Model synthesis of six-membered carbocyclic spironucleosides

    Czech Academy of Sciences Publication Activity Database

    Nencka, Radim; Hřebabecký, Hubert; Dračínský, Martin

    2010-01-01

    Roč. 75, č. 12 (2010), s. 1259-1272 ISSN 0010-0765 R&D Projects: GA MŠk 1M0508 Institutional research plan: CEZ:AV0Z40550506 Keywords : spironucleosides * carbocyclic * Bucherer-Bergs reaction * Michaels addition * tandem reaction Subject RIV: CC - Organic Chemistry Impact factor: 0.853, year: 2010

  1. A versatile synthon for the preparation for the preparation of carbasugars and carbocyclic nucleosides:

    DEFF Research Database (Denmark)

    Johansen, Steen Karsk

    Der er i det sidste decennium været stor forskningsaktivitet i relation til at fremstille og undersøge egenskaber af nye kulhydrat mimics da enzyminhibitorer er interessante i mange biologisk sammenhænge.Den kemiske udfordringen er at fremstille sådanne molekyler i en optisk aktiv form. Dette kan...

  2. Stereoselective Hydrogenation and Ozonolysis of Iridoids. Conversion into Carbocyclic Nucleoside Analogues

    DEFF Research Database (Denmark)

    Franzyk, Henrik; Stermitz, Frank R.

    1999-01-01

    Stereoselective hydrogenation of the iridoids geniposide (9) and aucubin (19) was achieved by using the 1-methyl-1-methoxyethyl ether (MIP) as protecting group for the allylic alcohol, as it enhanced the stereoselectivity and prevented undesired hydrogenolysis. Ozonolysis of the hydrogenation...

  3. Stereoselective Hydrogenation and Ozonolysis of Iridoids. Conversion into Carbocyclic Nucleoside Analogues

    DEFF Research Database (Denmark)

    Franzyk, Henrik; Stermitz, Frank R.

    1999-01-01

    product from 9, adoxoside (11), with reductive work-up, afforded either a chiral lactone (25) or a chiral polyol (26), depending on the reduction conditions. Polyol 26 was subjected to protecting group manipulation and subsequent oxidation and reductions to yield cyclopentane building blocks (29......Stereoselective hydrogenation of the iridoids geniposide (9) and aucubin (19) was achieved by using the 1-methyl-1-methoxyethyl ether (MIP) as protecting group for the allylic alcohol, as it enhanced the stereoselectivity and prevented undesired hydrogenolysis. Ozonolysis of the hydrogenation...

  4. Construction of Eight-Membered Carbocycles with Trisubstituted Double Bonds Using the Ring Closing Metathesis Reaction

    Directory of Open Access Journals (Sweden)

    Motoo Tori

    2010-06-01

    Full Text Available Medium sized carbocycles are particularly difficult to synthesize. Ring closing metathesis reactions (RCM have recently been applied to construct eight-membered carbocycles, but trisubstituted double bonds in the eight-membered rings are more difficult to produce using RCM reactions. In this review, model examples and our own results are cited and the importance of the preparation of suitably designed precursors is discussed. Examples of RCM reactions used in the total synthesis of natural products are also outlined.

  5. Nucleotides, Nucleosides, and Nucleobases

    DEFF Research Database (Denmark)

    Jensen, Kaj Frank; Dandanell, Gert; Hove-Jensen, Bjarne

    2008-01-01

    We review literature on the metabolism of ribo- and deoxyribonucleotides, nucleosides, and nucleobases in Escherichia coli and Salmonella,including biosynthesis, degradation, interconversion, and transport. Emphasis is placed on enzymology and regulation of the pathways, at both the level of gene...

  6. Synthetic Strategies for Converting Carbohydrates into Carbocycles by the Use of Olefin Metathesis

    DEFF Research Database (Denmark)

    Madsen, Robert

    2007-01-01

    This microreview covers recent advances in the use of ring-closing metathesis for the synthesis of carbocycles from carbohydrates. Various strategies for the synthesis of a,w-dienes from carbohydrates are presented, which give rise to a large variety of dienes with different stereochemistry, prot...

  7. Microbial transformation of nucleosides

    Science.gov (United States)

    Lamba, S. S.

    1979-01-01

    A study involving the use of coulter counter in studying the effects of neomycin on E. coli, S. aureus and A. aerogenes was completed. The purpose of this was to establish proper technique for enumeration of cells per ml. It was found that inhibitory effects on growth of E. coli and A. aerogenes, both gram negative organisms, were directly related to the concentration of neomycin used. However, in case S. aureus, a gram positive organism, a decreased inhibition was noted at higher concentrations. A paper entitled, Use of Coulter Counter in Studying Effect of Drugs on Cells in Culture 1 - Effects of Neomycin on E. coli, S. aureus and A. aerogenes, is attached in the appendix. Laboratory procedures were also established to study the effects of nucleoside antibiotic cordycepin on He La cell grown in suspension cultures.

  8. Regio- and stereodivergent antibiotic oxidative carbocyclizations catalysed by Rieske oxygenase-like enzymes

    Science.gov (United States)

    Sydor, Paulina K.; Barry, Sarah M.; Odulate, Olanipekun M.; Barona-Gomez, Francisco; Haynes, Stuart W.; Corre, Christophe; Song, Lijiang; Challis, Gregory L.

    2011-05-01

    Oxidative cyclizations, exemplified by the biosynthetic assembly of the penicillin nucleus from a tripeptide precursor, are arguably the most synthetically powerful implementation of C-H activation reactions in nature. Here, we show that Rieske oxygenase-like enzymes mediate regio- and stereodivergent oxidative cyclizations to form 10- and 12-membered carbocyclic rings in the key steps of the biosynthesis of the antibiotics streptorubin B and metacycloprodigiosin, respectively. These reactions represent the first examples of oxidative carbocyclizations catalysed by non-haem iron-dependent oxidases and define a novel type of catalytic activity for Rieske enzymes. A better understanding of how these enzymes achieve such remarkable regio- and stereocontrol in the functionalization of unactivated hydrocarbon chains will greatly facilitate the development of selective man-made C-H activation catalysts.

  9. Nucleoside composition of Heloderma venoms.

    Science.gov (United States)

    Aird, Steven D

    2008-06-01

    Venoms of Heloderma horridum and Heloderma suspectum were analyzed for the possible presence of purine and pyrimidine nucleosides. Adenosine, cytidine, guanosine, hypoxanthine, inosine, and uridine were found in mug quantities. These amounts are much smaller than those seen in many elapid or viperine venoms, but greater and more varied than those found in crotaline venoms. While their contribution to the hypotension induced by Heloderma venoms may be minor, venom nucleosides nonetheless act in concert with kallikreins/hemorrhagins, alkaline phosphomonoesterase, 5'-nucleotidase, helodermin, helospectins, helothermine, and serotonin. The use of nucleosides as toxins is therefore a generalized squamate strategy, rather than the exclusive province of snakes. Both Heloderma venoms were found to be devoid of NADase and phosphodiesterase activities. Enzymes to release endogenous purines in the prey, are not significant components of Heloderma venoms.

  10. Nucleoside Inhibitors of Zika Virus

    Czech Academy of Sciences Publication Activity Database

    Eyer, L.; Nencka, Radim; Huvarová, I.; Palus, Martin; Alves, M. J.; Gould, E. A.; De Clercq, E.; Růžek, Daniel

    2016-01-01

    Roč. 214, č. 5 (2016), s. 707-711 ISSN 0022-1899 Institutional support: RVO:61388963 ; RVO:60077344 Keywords : Zika virus * flavivirus * nucleoside analogue * antiviral * therapy Subject RIV: CC - Organic Chemistry; EE - Microbiology, Virology (BC-A) Impact factor: 6.273, year: 2016

  11. Ring-size-selective construction of fluorine-containing carbocycles via intramolecular iodoarylation of 1,1-difluoro-1-alkenes

    Directory of Open Access Journals (Sweden)

    Takeshi Fujita

    2017-12-01

    Full Text Available 1,1-Difluoro-1-alkenes bearing a biaryl-2-yl group effectively underwent site-selective intramolecular iodoarylation by the appropriate cationic iodine species. Iodoarylation of 2-(2-aryl-3,3-difluoroallylbiaryls proceeded via regioselective carbon–carbon bond formation at the carbon atoms in β-position to the fluorine substituents, thereby constructing dibenzo-fused six-membered carbocycles bearing a difluoroiodomethyl group. In contrast, 2-(3,3-difluoroallylbiaryls underwent a similar cyclization at the α-carbon atoms to afford ring-difluorinated seven-membered carbocycles.

  12. Synthesis of Carbocyclic Hydantocidins via Regioselective and Diastereoselective Phosphine-Catalyzed [3 + 2]-Cycloadditions to 5-Methylenehydantoins

    Energy Technology Data Exchange (ETDEWEB)

    Pham, Tien Q.; Pyne, Stephen G.; Skelton, Brian W.; White, Allan H. (UWA); (Wollongong)

    2010-07-20

    The phosphine-catalyzed [3 + 2]-cycloaddition of 5-methylenehydantoins 4 with the ylides 5, derived from addition of tributylphosphine to the 2-butynoic acid derivatives, 6a-d, gives spiro-heterocyclic products. The camphor sultam derivative 6b gives optically active products. Noteable was that the ylides derived from ethyl 2-butynoate and the 3-(2-butynoyl)-1,3-oxazolidin-2-one derivatives 6c and 6d gave spiro-heterocyclic products with reverse regioselectivities. The N,N-dibenzylprotected cycloadduct has been converted to carbocyclic hydantocidin and 6,7-diepi-carbocyclic hydantocidin.

  13. Marine Nucleosides: Structure, Bioactivity, Synthesis and Biosynthesis

    Directory of Open Access Journals (Sweden)

    Ri-Ming Huang

    2014-12-01

    Full Text Available Nucleosides are glycosylamines that structurally form part of nucleotide molecules, the building block of DNA and RNA. Both nucleosides and nucleotides are vital components of all living cells and involved in several key biological processes. Some of these nucleosides have been obtained from a variety of marine resources. Because of the biological importance of these compounds, this review covers 68 marine originated nucleosides and their synthetic analogs published up to June 2014. The review will focus on the structures, bioactivities, synthesis and biosynthetic processes of these compounds.

  14. Carbocyclization versus oxycyclization on the metal-catalyzed reactions of oxyallenyl C3-linked indoles.

    Science.gov (United States)

    Alcaide, Benito; Almendros, Pedro; Alonso, José M; Fernández, Israel

    2013-07-05

    The preparation of previously unknown (indol-3-yl)-α-allenols and -allenones was accomplished from indole-3-carbaldehydes, through indium-mediated Barbier allenylation reaction taking advantage of the N-(2-pyridyl)sulfonyl group. Metal-catalyzed cyclizations of oxyallenyl C3-linked indoles proceeded in two ways depending on the presence or absence of the N-(2-pyridyl)sulfonyl group. For allenols, gold-catalyzed oxycyclization occurred in the presence of the protecting group; in the absence of the protecting group, palladium- and gold-catalyzed benzannulations operated. On the contrary, under gold catalysis furyl-indoles were obtained as exclusive products from NH-allenones, while 5-endo carbocyclization adducts were the major components starting from N-SO2py-protected allenones. These cyclization reactions have been developed experimentally, and their mechanisms have additionally been investigated by a computational study.

  15. Nucleoside antibiotics: biosynthesis, regulation, and biotechnology.

    Science.gov (United States)

    Niu, Guoqing; Tan, Huarong

    2015-02-01

    The alarming rise in antibiotic-resistant pathogens has coincided with a decline in the supply of new antibiotics. It is therefore of great importance to find and create new antibiotics. Nucleoside antibiotics are a large family of natural products with diverse biological functions. Their biosynthesis is a complex process through multistep enzymatic reactions and is subject to hierarchical regulation. Genetic and biochemical studies of the biosynthetic machinery have provided the basis for pathway engineering and combinatorial biosynthesis to create new or hybrid nucleoside antibiotics. Dissection of regulatory mechanisms is leading to strategies to increase the titer of bioactive nucleoside antibiotics. Copyright © 2014. Published by Elsevier Ltd.

  16. SOME RECENT FINDINGS IN THE BIOTECHNOLOGY OF BIOLOGICALLY IMPORTANT NUCLEOSIDES

    Directory of Open Access Journals (Sweden)

    A. Mikhailopulo

    2013-08-01

    Full Text Available Some recent findings in the biotechnology of biologically important nucleosides will be discussed, viz., (i a new strategy of the cascade one-pot transformation of D-pentoses into nucleosides based on the extension and deepening of the knowledge of the mechanism of functioning of the ribokinase, phosphopentomutase, and uridine, thymidine and purine nucleoside (PNP phosphorylases, and the role of different factors (structural, electronic, stereochemical in the glycoside bond formation, (ii the modern chemistries of the chemo-enzymatic syntheses of nucleosides, (iii the transglycosylation reaction using natural and sugar modified nucleosides as donors of carbohydrate residues and heterocyclic bases as acceptors catalyzed by nucleoside phosphorylases (NP.

  17. Synthesis of novel conformationally locked carbocyclic nucleosides derived from 5,5- and 6,6-bis(hydroxymethyl)bicyclo[2.2.1]heptan-2-ol

    Czech Academy of Sciences Publication Activity Database

    Hřebabecký, Hubert; Masojídková, Milena; Holý, Antonín

    2005-01-01

    Roč. 70, č. 4 (2005), s. 519-538 ISSN 0010-0765 R&D Projects: GA ČR(CZ) GA203/05/0132 Institutional research plan: CEZ:AV0Z40550506 Keywords : azides * amines * cyclic sulfates Subject RIV: CC - Organic Chemistry Impact factor: 0.949, year: 2005

  18. A compendium of cyclic sugar amino acids and their carbocyclic and heterocyclic nitrogen analogues.

    Science.gov (United States)

    Risseeuw, Martijn; Overhand, Mark; Fleet, George W J; Simone, Michela I

    2013-10-01

    This compendium focuses on functionalised sugar amino acids (SAAs) and their 3- to 6-membered nitrogen heterocyclic and carbocyclic analogues. The main benefit of using SAAs and their related nitrogen and carbon congeners in the production of peptidomimetics and glycomimetics is that their properties can be readily altered via modification of their ring size, chemical manipulation of their numerous functional groups and fine-tuning of the stereochemical arrangement of their ring substituents. These building blocks provide access to hydrophilic and hydrophobic peptide isosteres whose physical properties allow entry to a region of chemotherapeutic space which is still under-explored by medicinal chemists. These building blocks are also important in providing amino acids whose inherent conformational bias leads to predisposition to secondary structure upon oligomerisation in relatively short sequences. These foldamers, particularly those containing ω-amino acids, provide an additional opportunity to expand access to the control of structures by artificial peptides. The synthesis and biological evaluation of these building blocks in glycomimetics and peptidomimetics systems keep expanding the reach of the glycosciences to the medical sciences, provide a greater outlook onto the wide range of cellular functions of saccharides and their derivatives involved and greater insight into the nature of oligosaccharide and protein folding.

  19. Pyrrolidine analogues of nucleosides and nucleotides

    Czech Academy of Sciences Publication Activity Database

    Rejman, Dominik; Pohl, Radek; Kovačková, Soňa; Kočalka, Petr; Švenková, Alžběta; Šanderová, Hana; Krásný, Libor; Rosenberg, Ivan

    -, č. 52 (2008), s. 577-578 ISSN 0261-3166. [Joint Symposium of the International Roundtable on Nucleosides, Nucleotides and Nucleic Acids /18./ and the International Symposium on Nucleic Acid Chemistry /35./. Kyoto, 08.09.2008-12.09.2008] R&D Projects: GA MŠk(CZ) LC06077; GA MŠk 2B06065; GA MZd NR9138 Institutional research plan: CEZ:AV0Z40550506; CEZ:AV0Z50200510; CEZ:AV0Z50520514 Keywords : pyrrolidine * nucleoside * nucleotide Subject RIV: CC - Organic Chemistry

  20. Synthesis and turnover of purine nucleoside phosphorylase in human lymphocytes

    International Nuclear Information System (INIS)

    Snyder, F.F.; Kwan, E.; Mably, E.R.; Neote, K.

    1986-01-01

    The authors study the synthesis of purine nucleoside phosphorylase during phytohumagglutinin induced T cell transformation and have examined the turnover of purine nucleoside phosphorylase in thte human lymphoblast WI-L2. A polyclonal antibody to purine nucleoside phosphorylase in the synthetic and turnover studies is used. Fluorography of purine nucleoside phosphorylase immunoprecipitates from WI-L2 cells are shown. Lymphoblast proteins were labelled during 16hour cultrue with 4,5-tritium-leucine. Samples were 20 ug of total protein, immunoprecipitate of 200 ug protein with control serum and purine nucleoside phosphorylase antiserum

  1. Tetrofuranose nucleoside phosphonic acids: Synthesis and properties

    Czech Academy of Sciences Publication Activity Database

    Poláková, Ivana; Buděšínský, Miloš; Točík, Zdeněk; Rosenberg, Ivan

    2011-01-01

    Roč. 76, č. 5 (2011), s. 503-536 ISSN 0010-0765 R&D Projects: GA AV ČR KAN200520801; GA ČR GA203/09/0820; GA MŠk(CZ) LC06061; GA MŠk(CZ) LC06077 Institutional research plan: CEZ:AV0Z40550506 Keywords : tetrofuranosyl phosphonate * nucleotide analogues * phosphonomethoxy nucleosides * sugar hydroxyphosphonates Subject RIV: CC - Organic Chemistry Impact factor: 1.283, year: 2011

  2. Synthesis and Biological Activity of Reversed Pyrimidine Nucleosides

    Directory of Open Access Journals (Sweden)

    Nataša Župančić

    2015-03-01

    Full Text Available An efficient approach to reversed nucleosides which enables their synthesis in gram quantities is described. N-1′-Pyrimidine reversed nucleosides were prepared by treating of the sodium salt of pyrimidine bases with protected 5-tosyl ribose. Additionally, N-1′,N-3′-disubstituted reversed nucleosides were isolated in the condensation reactions with the 5-halogen pyrimidines. Using the Sonogashira coupling of 5′-iodouracil reversed nucleoside with ethynyltrimethyl silane gave 5′-ethynyl derivative which was further transformed into 5′-acetyl reversed nucleoside. Biological activity of deprotected reversed nucleosides was validated on the panel of six human carcinoma cell lines (HeLa, MIAPaCa2, Hep2, NCI-H358, CaCo-2, and HT-29. 5′-Iodouracil derivative displayed moderate growth inhibition activity against human colon carcinoma (CaCo-2 cells.

  3. Nucleoside transport in primary cultured rabbit tracheal epithelial cells.

    Science.gov (United States)

    Mathias, Neil R; Wu, Sharon K; Kim, Kwang-Jin; Lee, Vincent H L

    2005-01-01

    The present study aimed at elucidating the mechanisms of nucleoside transport in primary cultured rabbit tracheal epithelial cells (RTEC) grown on a permeable filter support. Uptake of (3)H-uridine, the model nucleoside substrate, from the apical fluid of primary cultured RTEC was examined with respect to its dependence on Na(+), substrate concentration, temperature and its sensitivity to inhibitors, other nucleosides and antiviral nucleoside analogs. Apical (3)H-uridine uptake in primary cultured RTEC was strongly dependent on an inward Na(+) gradient and temperature. Ten micromolar nitro-benzyl-mercapto-purine-ribose (NBMPR) (an inhibitor of es-type nucleoside transport in the nanomolar range) did not further inhibit this process. (3)H-uridine uptake from apical fluid was inhibited by basolateral ouabain (10 microM) and apical phloridzin (100 microM), indicating that uptake may involve a secondary active transport process. Uridine uptake was saturable with a K(m) of 3.4 +/- 1.8 microM and the V(max) of 24.3 +/- 5.2 pmoles/mg protein/30 s. Inhibition studies indicated that nucleoside analogs that have a substitution on the nucleobase competed with uridine uptake from apical fluid, but those with modifications on the ribose sugar including acyclic analogs were ineffective. The pattern of inhibition of apical (3)H-uridine, (3)H-inosine and (3)H-thymidine uptake into RTEC cells by physiological nucleosides was consistent with multiple systems: A pyrimidine-selective transport system (CNT1); a broad nucleoside substrate transport system that excludes inosine (CNT4) and an equilibrative NBMPR-insensitive nucleoside transport system (ei type). These results indicate that the presence of apically located nucleoside transporters in the epithelial cells lining the upper respiratory tract can lead to a high accumulation of nucleosides in the trachea. At least one Na(+)-dependent, secondary, active transport process may mediate the apical absorption of nucleosides or

  4. Practical Considerations For Developing Nucleoside Reverse Transcriptase Inhibitors

    OpenAIRE

    Hurwitz, Selwyn J.; Schinazi, Raymond F.

    2012-01-01

    Nucleoside reverse transcriptase inhibitors (NRTI) remain a cornerstone of current antiretroviral regimens in combinations usually with a non-nucleoside reverse transcriptase inhibitor (NNRTI), a protease inhibitor (PI), or an integrase inhibitor (INI). The antiretroviral efficacy and relative safety of current NRTI results from a tight and relatively specific binding of their phosphorylated nucleoside triphosphates (NRTI-TP) with the HIV-1 reverse transcriptase which is ess...

  5. Structure of grouper iridovirus purine nucleoside phosphorylase

    International Nuclear Information System (INIS)

    Kang, You-Na; Zhang, Yang; Allan, Paula W.; Parker, William B.; Ting, Jing-Wen; Chang, Chi-Yao; Ealick, Steven E.

    2010-01-01

    The crystal structure of purine nucleoside phosphorylase from grouper iridovirus was solved at 2.38 Å resolution. Purine nucleoside phosphorylase (PNP) catalyzes the reversible phosphorolysis of purine ribonucleosides to the corresponding free bases and ribose 1-phosphate. The crystal structure of grouper iridovirus PNP (givPNP), corresponding to the first PNP gene to be found in a virus, was determined at 2.4 Å resolution. The crystals belonged to space group R3, with unit-cell parameters a = 193.0, c = 105.6 Å, and contained four protomers per asymmetric unit. The overall structure of givPNP shows high similarity to mammalian PNPs, having an α/β structure with a nine-stranded mixed β-barrel flanked by a total of nine α-helices. The predicted phosphate-binding and ribose-binding sites are occupied by a phosphate ion and a Tris molecule, respectively. The geometrical arrangement and hydrogen-bonding patterns of the phosphate-binding site are similar to those found in the human and bovine PNP structures. The enzymatic activity assay of givPNP on various substrates revealed that givPNP can only accept 6-oxopurine nucleosides as substrates, which is also suggested by its amino-acid composition and active-site architecture. All these results suggest that givPNP is a homologue of mammalian PNPs in terms of amino-acid sequence, molecular mass, substrate specificity and overall structure, as well as in the composition of the active site

  6. Two nucleoside transporters in Lactococcus lactis with different substrate specificities

    DEFF Research Database (Denmark)

    Martinussen, Jan; Sørensen, Claus; Jendresen, Christian Bille

    2010-01-01

    , and the utilization of nucleotides is dependent on exogenous phosphatases. The composition of transporters with specificity for purine and pyrimidine nucleosides and nucleobases is subject to variation. The ability of Lactococcus lactis to transport different nucleosides across the cell membrane was characterized...

  7. Asymmetric synthesis of densely functionalized medium-ring carbocycles and lactones through modular assembly and ring-closing metathesis of sulfoximine-substituted trienes and dienynes.

    Science.gov (United States)

    Lejkowski, Michal; Banerjee, Prabal; Schüller, Sabine; Münch, Alexander; Runsink, Jan; Vermeeren, Cornelia; Gais, Hans-Joachim

    2012-03-19

    An asymmetric synthesis of densely functionalized 7-11-membered carbocycles and 9-11-membered lactones has been developed. Its key steps are a modular assembly of sulfoximine-substituted C- and O-tethered trienes and C-tethered dienynes and their Ru-catalyzed ring-closing diene and enyne metathesis (RCDEM and RCEYM). The synthesis of the C-tethered trienes and dienynes includes the following steps: 1) hydroxyalkylation of enantiomerically pure titanated allylic sulfoximines with unsaturated aldehydes, 2) α-lithiation of alkenylsulfoximines, 3) alkylation, hydroxy-alkylation, formylation, and acylation of α-lithioalkenylsulfoximines, and 4) addition of Grignard reagents to α-formyl(acyl)alkenylsulfoximines. The sulfoximine group provided for high asymmetric induction in steps 1) and 4). RCDEM of the sulfoximine-substituted trienes with the second-generation Ru catalyst stereoselectively afforded the corresponding functionalized 7-11-membered carbocyles. RCDEM of diastereomeric silyloxy-substituted 1,6,12-trienes revealed an interesting difference in reactivity. While the (R)-diastereomer gave the 11-membered carbocyle, the (S)-diastereomer delivered in a cascade of cross metathesis and RCDEM 22-membered macrocycles. RCDEM of cyclic trienes furnished bicyclic carbocycles with a bicyclo[7.4.0]tridecane and bicyclo[9.4.0]pentadecane skeleton. Selective transformations of the sulfoximine- and bissilyloxy-substituted carbocycles were performed including deprotection, cross-coupling reaction and reduction of the sulfoximine moiety. Esterification of a sulfoximine-substituted homoallylic alcohol with unsaturated carboxylic acids gave the O-tethered trienes, RCDEM of which yielded the sulfoximine-substituted 9-11-membered lactones. RCEYM of a sulfoximine-substituted 1,7-dien-10-yne showed an unprecedented dichotomy in ring formation depending on the Ru catalyst. While the second-generation Ru catalyst gave the 9-membered exo 1,3-dienyl carbocycle, the first-generation Ru

  8. An Efficient and Facile Methodology for Bromination of Pyrimidine and Purine Nucleosides with Sodium Monobromoisocyanurate (SMBI

    Directory of Open Access Journals (Sweden)

    Roger Stromberg

    2013-10-01

    Full Text Available An efficient and facile strategy has been developed for bromination of nucleosides using sodium monobromoisocyanurate (SMBI. Our methodology demonstrates bromination at the C-5 position of pyrimidine nucleosides and the C-8 position of purine nucleosides. Unprotected and also several protected nucleosides were brominated in moderate to high yields following this procedure.

  9. Vitamin E Phosphate Nucleoside Prodrugs: A Platform for Intracellular Delivery of Monophosphorylated Nucleosides

    Directory of Open Access Journals (Sweden)

    Richard Daifuku

    2018-02-01

    Full Text Available Vitamin E phosphate (VEP nucleoside prodrugs are designed to bypass two mechanisms of tumor resistance to therapeutic nucleosides: nucleoside transport and kinase downregulation. Certain isoforms of vitamin E (VE have shown activity against solid and hematologic tumors and result in chemosensitization. Because gemcitabine is one of the most common chemotherapeutics for the treatment of cancer, it was used to demonstrate the constructs utility. Four different VE isoforms were conjugated with gemcitabine at the 5′ position. Two of these were δ-tocopherol-monophosphate (MP gemcitabine (NUC050 and δ-tocotrienol-MP gemcitabine (NUC052. NUC050 was shown to be able to deliver gemcitabine-MP intracellularly by a nucleoside transport independent mechanism. Its half-life administered IV in mice was 3.9 h. In a mouse xenograft model of non-small cell lung cancer (NSCLC NCI-H460, NUC050 at a dose of 40 mg/kg IV qwk × 4 resulted in significant inhibition to tumor growth on days 11–31 (p < 0.05 compared to saline control (SC. Median survival was 33 days (NUC050 vs. 25.5 days (SC ((hazard ratio HR = 0.24, p = 0.017. Further, NUC050 significantly inhibited tumor growth compared to historic data with gemcitabine at 135 mg/kg IV q5d × 3 on days 14–41 (p < 0.05. NUC052 was administered at a dose of 40 mg/kg IV qwk × 2 followed by 50 mg/kg qwk × 2. NUC052 resulted in inhibition to tumor growth on days 14–27 (p < 0.05 and median survival was 34 days (HR = 0.27, p = 0.033. NUC050 and NUC052 have been shown to be safe and effective in a mouse xenograft of NSCLC.

  10. Natural and engineered biosynthesis of nucleoside antibiotics in Actinomycetes.

    Science.gov (United States)

    Chen, Wenqing; Qi, Jianzhao; Wu, Pan; Wan, Dan; Liu, Jin; Feng, Xuan; Deng, Zixin

    2016-03-01

    Nucleoside antibiotics constitute an important family of microbial natural products bearing diverse bioactivities and unusual structural features. Their biosynthetic logics are unique with involvement of complex multi-enzymatic reactions leading to the intricate molecules from simple building blocks. Understanding how nature builds this family of antibiotics in post-genomic era sets the stage for rational enhancement of their production, and also paves the way for targeted persuasion of the cell factories to make artificial designer nucleoside drugs and leads via synthetic biology approaches. In this review, we discuss the recent progress and perspectives on the natural and engineered biosynthesis of nucleoside antibiotics.

  11. Medicinal Chemistry of Fluorinated Cyclic and Acyclic Nucleoside Phosphonates

    Czech Academy of Sciences Publication Activity Database

    Baszczyňski, Ondřej; Janeba, Zlatko

    2013-01-01

    Roč. 33, č. 6 (2013), s. 1304-1344 ISSN 0198-6325 Institutional support: RVO:61388963 Keywords : nucleoside phosphosphonates * fluorine * antiviral * anticancer * antiparasitic Subject RIV: CC - Organic Chemistry Impact factor: 8.131, year: 2013

  12. Impact of unnatural nucleosides on the control of microbial growth.

    Science.gov (United States)

    Hatano, Akihiko; Nishimura, Makoto; Souta, Ikuo

    2009-06-01

    This research investigated the antimicrobial activities of unnatural nucleosides. We tested the MIC and MBC of 17 synthetic nucleoside analogues against 10 microbial strains. These nucleoside analogues were classified into four groups according to their structural characteristics. Inhibition was observed with compounds 1-1, 3-1, and 4-3. In particular, 5'-deoxythymidine (3-1) was most effective at 50 micro g/mL against Bacillus subtilis and Staphylococcus aureus. This analogue has had the hydroxyl group at the 5' position replaced with a hydrogen atom. All compounds had weak effects against various species of mold. The MBC of 5'-deoxythymidine was 50 g/mL in 0.5 h against S. aureus. These results showed that 5'-deoxythymidine had the most effective antimicrobial activity of the 17 different unnatural nucleosides. The inhibitory effect of 3-1 suggests that it may be useful as an antibacterial agent in medical situations.

  13. Current prodrug strategies for improving oral absorption of nucleoside analogues

    Directory of Open Access Journals (Sweden)

    Youxi Zhang

    2014-04-01

    Full Text Available Nucleoside analogues are first line chemotherapy in various severe diseases: AIDS (acquired immunodeficiency disease syndrome, cytomegalovirus infections, cancer, etc. However, many nucleoside analogues exhibit poor oral bioavailability because of their high polarity and low intestinal permeability. In order to get around this drawback, prodrugs have been utilized to improve lipophilicity by chemical modification of the parent drug. Alternatively, prodrugs targeting transporters present in the intestine have been applied to promote the transport of the nucleoside analogues. Valacyclovir and valganciclovir are two classic valine ester prodrugs transported by oligopeptide transporter 1. The ideal prodrug achieves delivery of a parent drug by attaching a non-toxic moiety that is stable during transport, but is readily degraded to the parent drug once at the target. This article presents advances of prodrug approaches for enhancing oral absorption of nucleoside analogues.

  14. Meteorite-catalyzed synthesis of nucleosides and other prebiotic compounds

    Czech Academy of Sciences Publication Activity Database

    Ferus, Martin; Knížek, Antonín; Civiš, Svatopluk

    2015-01-01

    Roč. 112, č. 23 (2015), s. 7109-7110 ISSN 0027-8424 Institutional support: RVO:61388955 Keywords : meteorite-catalzzed synthesis * nucleosides * prebiotic compounds Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 9.423, year: 2015

  15. Crystal structure of a concentrative nucleoside transporter from Vibrio cholerae at 2.4;#8201;Å

    Energy Technology Data Exchange (ETDEWEB)

    Johnson, Zachary Lee; Cheong, Cheom-Gil; Lee, Seok-Yong (Duke)

    2012-07-11

    Nucleosides are required for DNA and RNA synthesis, and the nucleoside adenosine has a function in a variety of signalling processes. Transport of nucleosides across cell membranes provides the major source of nucleosides in many cell types and is also responsible for the termination of adenosine signalling. As a result of their hydrophilic nature, nucleosides require a specialized class of integral membrane proteins, known as nucleoside transporters (NTs), for specific transport across cell membranes. In addition to nucleosides, NTs are important determinants for the transport of nucleoside-derived drugs across cell membranes. A wide range of nucleoside-derived drugs, including anticancer drugs (such as Ara-C and gemcitabine) and antiviral drugs (such as zidovudine and ribavirin), have been shown to depend, at least in part, on NTs for transport across cell membranes. Concentrative nucleoside transporters, members of the solute carrier transporter superfamily SLC28, use an ion gradient in the active transport of both nucleosides and nucleoside-derived drugs against their chemical gradients. The structural basis for selective ion-coupled nucleoside transport by concentrative nucleoside transporters is unknown. Here we present the crystal structure of a concentrative nucleoside transporter from Vibrio cholerae in complex with uridine at 2.4 {angstrom}. Our functional data show that, like its human orthologues, the transporter uses a sodium-ion gradient for nucleoside transport. The structure reveals the overall architecture of this class of transporter, unravels the molecular determinants for nucleoside and sodium binding, and provides a framework for understanding the mechanism of nucleoside and nucleoside drug transport across cell membranes.

  16. Thermus thermophilus Strains Active in Purine Nucleoside Synthesis

    Directory of Open Access Journals (Sweden)

    Marcos Almendros

    2009-03-01

    Full Text Available Several strains of Thermus thermophilus were tested in order to detect purine nucleoside synthase activity using pyrimidine nucleosides as the sugar-donor and adenine or hypoxanthine as bases. High productivity values (t =1 hr were obtained while completely avoiding adenosine-deaminase degradation of the products. N-2-deoxy-ribosyltransferase activity is described for the first time in hyperthermophilic bacteria.

  17. Synthesis of coumarin or ferrocene labeled nucleosides via Staudinger ligation

    Directory of Open Access Journals (Sweden)

    Kois Pavol

    2006-11-01

    Full Text Available Abstract Background Reaction of azides with triaryl phosphines under mild conditions gives iminophosphoranes which can react with almost any kind of electrophilic reagent, e.g. aldehydes/ketones to form imines or esters to form amides. This so-called Staudinger ligation has been employed in a wide range of applications as a general tool for bioconjugation including specific labeling of nucleic acids. Results A new approach for the preparation of labeled nucleosides via intermolecular Staudinger ligation is described. Reaction of azidonucleosides with triphenylphosphine lead to iminophosphorane intermediates, which react subsequently with derivatives of coumarin or ferrocene to form coumarin or ferrocene labeled nucleosides. Fluorescent properties of coumarin labeled nucleosides are determined. Conclusion New coumarin and ferrocene labeled nucleosides were prepared via intermolecular Staudinger ligation. This reaction joins the fluorescent coumarin and biospecific nucleoside to the new molecule with promising fluorescent and electrochemical properties. The isolated yields of products depend on the structure of azidonucleoside and carboxylic acids. A detailed study of the kinetics of the Staudinger ligation with nucleoside substrates is in progress.

  18. Total syntheses of hyperforin and papuaforins A-C, and formal synthesis of nemorosone through a gold(I)-catalyzed carbocyclization.

    Science.gov (United States)

    Bellavance, Gabriel; Barriault, Louis

    2014-06-23

    The remarkable biological activities of polyprenylated polycyclic acylphloroglucinols (PPAPs) combined with their highly decorated bicyclo[3.3.1]nonane-2,4,9-trione frameworks have inspired synthetic organic chemists over the last decade. The concise total syntheses of four natural products PPAPs; hyperforin and papuaforins A-C, and the formal synthesis of nemorosone are reported. Key to the realization of this strategy is the short and scalable synthesis of densely substituted PPAP scaffolds through a gold(I)-catalyzed 6-endo-dig carbocyclization of cyclic enol ethers for late-stage functionalization. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Novel Applications of Donor-Acceptor Cyclopropanes and Dearomatization towards the Expedient Synthesis of Highly Substituted Carbocycles

    Science.gov (United States)

    Mackay, William Daniel

    I. Lewis Acid Catalyzed (3+2)-Annulations of Donor-Acceptor Cyclopropanes and Ynamides. The Sc(OTf)3-catalyzed (3+2)-annulation of donor-acceptor cyclopropanes and ynamides is described, providing the corresponding cyclopentene sulfonamides in good to excellent yield. Deprotection and hydrolysis of the resulting cyclopentenesulfonamides delivers 2,3-substituted cyclopentanones with high diastereoselectivity. II. Kinetic Separation and Asymmetric Reactions of Norcaradiene Cycloadducts: Facilitated Access via H2O-Accelerated Cycloaddition. We exploit the Buchner reaction to access 1,2-disubstituted cyclohexadiene synthons (norcaradienes), which participate in H2O-accelerated cycloaddition with dienophiles to provide cyclopropyl-fused [2.2.2]-bicyclooctene derivatives in good yields. Regioisomeric mixtures can be kinetically separated exploiting different reaction rates in Diels-Alder reactions. meso -Diels-Alder products may be enantioselectively desymmetrized, providing highly substituted cyclohexanes with up to seven contiguous stereocenters. III. The Development of Regioisomerically Enriched Buchner Products for Use as Cyclohexadienyl Synthetic Intermediates. We have investigated two conceptual methods to generate highly regioisomerically enriched norcaradienyl intermediates through arene cyclopropanation. Intermolecular Buchner reaction of aryl diazoacetates under either thermolysis or silver(I) catalysis provide expedient routes to single regioisomeric norcaradienes, in some cases favoring the least sterically encumbered site of cyclopropanation. Intramolecular Buchner reaction of benzyl cyanodiazoacetates allow for the site-selective cyclopropanation of the tethered arene, and the installation of an activated cyclopropane for downstream functionalization. Both methods generate norcaradienes that are amenable to further transformations to generate highly stereochemically complex carbocyclic products.

  20. Cross-coupling reactions of nucleoside triphosphates followed by polymerase incorporation. Construction and applications of base-functionalized nucleic acids

    Czech Academy of Sciences Publication Activity Database

    Hocek, Michal; Fojta, Miroslav

    2008-01-01

    Roč. 6, č. 13 (2008), s. 2233-2241 ISSN 1477-0520 R&D Projects: GA ČR GA203/05/0043; GA ČR GA203/07/1195; GA MŠk LC512; GA MŠk(CZ) LC06035 Institutional research plan: CEZ:AV0Z40550506; CEZ:AV0Z50040507 Keywords : DNA * nucleotides * cross-coupling reactions Subject RIV: CC - Organic Chemistry Impact factor: 3.550, year: 2008

  1. 2'-Fluoro-6'-methylene carbocyclic adenosine and its phosphoramidate prodrug: A novel anti-HBV agent, active against drug-resistant HBV mutants.

    Science.gov (United States)

    Singh, Uma S; Mulamoottil, Varughese A; Chu, Chung K

    2018-05-01

    Chronic hepatitis B (CHB) is one of the major causes of morbidity and mortality worldwide. Currently, clinically approved nucleos(t)ide analogs (NAs) are very efficient in reducing the load of hepatitis B virus (HBV) with minimum side effects. However, the long-term administration of antiviral drugs promotes HBV for potential drug resistance. To overcome this problem, combination therapies are administered, but HBV progressively altered mutations remain a threat. Therefore, optimally designed NAs are urgently needed to treat drug-resistant HBV. Herein, 2'-fluoro-6'-methylene carbocyclic adenosine (FMCA) and its phosphoramidate (FMCAP) have been discovered, which may be utilized in combination therapies for curing drug-resistant chronic hepatitis B. In preclinical studies, these carbocyclic NAs demonstrated potential anti-HBV activity against adefovir, as well as lamivudine (LMV/LAM) drug-resistant mutants. In vitro, these molecules have demonstrated significant activity against LMV/entecavir (ETV) triple mutants (L180M + S202G + M204V). Also, preliminary studies of FMCA/FMCAP in chimeric mice and female Non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mouse models having the LMV/ETV triple mutant have shown a high rate of reduction of HBV DNA levels compared to ETV. In this review, we have summarized preclinical studies of FMCA and its phosphoramidate prodrug (FMCAP). © 2018 Wiley Periodicals, Inc.

  2. Synthesis and Antiviral Activity of 3-Aminoindole Nucleosides of 2-Acetamido-2-deoxy-D-glucose

    Energy Technology Data Exchange (ETDEWEB)

    Abdelrahman, Adel A. H.; Elessawy, Farag A.; Barakat, Yousif A. [Menoufia Univ., Shebin El-Koam (Egypt); Ellatif, Mona M. Abd [The British Univ. in Egypt, Cairo (Egypt)

    2012-10-15

    A new method for the construction of 3-aminoindole nucleosides of 2-acetamido-2-deoxy-D-glucose based is presented. Nitration and acetylation of the indole nucleosides by acetic anhydride-nitric acid mixture followed by reduction using silver catalyst (SNSM) impregnated on silica gel, afforded the corresponding amino indole nucleosides. The nucleosides were tested for antiviral activity against hepatitis B virus (HBV) to show different degrees of antiviral activities or inhibitory actions.

  3. Advanced Prodrug Strategies in Nucleoside and Non-Nucleoside Antiviral Agents: A Review of the Recent Five Years

    Directory of Open Access Journals (Sweden)

    Hanadi Sinokrot

    2017-10-01

    Full Text Available Background: Poor pharmacokinetic profiles and resistance are the main two drawbacks from which currently used antiviral agents suffer, thus make them excellent targets for research, especially in the presence of viral pandemics such as HIV and hepatitis C. Methods: The strategies employed in the studies covered in this review were sorted by the type of drug synthesized into ester prodrugs, targeted delivery prodrugs, macromolecular prodrugs, other nucleoside conjugates, and non-nucleoside drugs. Results: Utilizing the ester prodrug approach a novel isopropyl ester prodrug was found to be potent HIV integrase inhibitor. Further, employing the targeted delivery prodrug zanamivir and valine ester prodrug was made and shown a sole delivery of zanamivir. Additionally, VivaGel, a dendrimer macromolecular prodrug, was found to be very efficient and is now undergoing clinical trials. Conclusions: Of all the strategies employed (ester, targeted delivery, macromolecular, protides and nucleoside analogues, and non-nucleoside analogues prodrugs, the most promising are nucleoside analogues and macromolecular prodrugs. The macromolecular prodrug VivaGel works by two mechanisms: envelope mediated and receptor mediated disruption. Nucleotide analogues have witnessed productive era in the recent past few years. The era of non-interferon based treatment of hepatitis (through direct inhibitors of NS5A has dawned.

  4. Synthesis of O-Amino Sugars and Nucleosides.

    Science.gov (United States)

    Chen, Na; Xie, Juan

    2018-03-12

    Nucleic acids and carbohydrates are essential biomolecules involved in numerous biological and pathological processes. Development of multifunctional building blocks based on nucleosides and sugars is in high demand for the generation of novel oligonucleotide mimics and glycoconjugates for biomedical applications. Recently, aminooxyl-functionalized compounds have attracted increasing research interest because of their easy derivatization through oxime ligation or N -oxyamide formation reactions. Various biological applications have been reported for O -amino carbohydrate- and nucleoside-derived compounds. Here, we report our efforts in the design and synthesis of glyco-, glycosyl, nucleoside- and nucleo-aminooxy acid derivatives from readily available sugars and amino acids, and their use for the generation of N -oxyamide-linked oligosaccharides, glycopeptides, glycolipids, oligonucleosides and nucleopeptides as novel glycoconjugates or oligonucleotide mimics. Delicate and key points in the synthesis will be emphasized.

  5. Synthesis of some novel hydrazono acyclic nucleoside analogues

    Directory of Open Access Journals (Sweden)

    Mohammad N. Soltani Rad

    2010-05-01

    Full Text Available The syntheses of novel hydrazono acyclic nucleosides similar to miconazole scaffolds are described. In this series of acyclic nucleosides, pyrimidine as well as purine and other azole derivatives replaced the imidazole function in miconazole and the ether group was replaced with a hydrazone moiety using phenylhydrazine. To interpret the dominant formation of (E-hydrazone derivatives rather than (Z-isomers, PM3 semiempirical quantum mechanic calculations were carried out which indicated that the (E-isomers had the lower heats of formation.

  6. New prodrugs based on phospholipid-nucleoside conjugates

    Energy Technology Data Exchange (ETDEWEB)

    MacCoss, M.

    1982-02-03

    A method is described for the preparation of defined, isomerically pure phospholipid-nucleoside conjugates as a prodrug in which the drug (araC) is attached to the phospholipid by a monophosphate linkage. Key intermediates in the process involve selective blocking and deblocking of the nucleoside derivative. These particular monophosphate-linked derivatives represent a new class of prodrug, which are useful by themselves or in combination with diphosphate linked derivatives. Several new compositions involving diphosphate linked derivatives are described in which the products are isomerically pure and having defined fatty acid chain lengths.

  7. Prolinol-based nucleoside phosphonic acids: Synthesis and properties

    Czech Academy of Sciences Publication Activity Database

    Vaněk, Václav; Buděšínský, Miloš; Liboska, Radek; Hurychová, Vladimíra; Rosenberg, Ivan

    -, č. 52 (2008), s. 537-538 ISSN 0261-3166. [Joint Symposium of the International Roundtable on Nucleosides, Nucleotides and Nucleic Acids /18./ and the International Symposium on Nucleic Acid Chemistry /35./. Kyoto, 08.09.2008-12.09.2008] R&D Projects: GA MŠk(CZ) LC06061; GA MŠk(CZ) LC06077; GA MŠk LC512 Institutional research plan: CEZ:AV0Z40550506 Keywords : oligonucleotide phosphonate * nucleoside analogue * pyrrolidine * prolinol Subject RIV: CC - Organic Chemistry

  8. Two nucleoside uptake systems in Lactococcus lactis: Competition between purine nucleosides and cytidine allows for modulation of intracellular nucleotide pools

    DEFF Research Database (Denmark)

    Martinussen, Jan; Wadskov-Hansen, Steen Lyders Lerche; Hammer, Karin

    2003-01-01

    and with an H3BO3-LiOH buffer for separation in the second dimension. We report here the sizes of the ribo- and deoxyribonucleotide pools in laboratory strain MG1363 during growth in a defined medium. We found that purine and pyrimidine-requiring strains may be used to establish physiological conditions...... in batch fermentations with altered nucleotide pools and growth rates by addition of nucleosides in different combinations. Addition of cytidine together with inosine to a purine-requiring strain leads to a reduction in the internal purine nucleotide pools and a decreased growth rate. This effect...... was found between uridine and either cytidine or inosine. These findings suggest that there are two different high-affinity nucleoside transporters, one system responsible for uridine uptake and another system responsible for the uptake of all purine nucleosides and cytidine....

  9. Aryl nucleoside H-phosphonates. Part 15: Synthesis, properties and, anti-HIV activity of aryl nucleoside 5'-alpha-hydroxyphosphonates.

    Science.gov (United States)

    Szymańska, Agnieszka; Szymczak, Marzena; Boryski, Jerzy; Stawiński, Jacek; Kraszewski, Adam; Collu, Gabriella; Sanna, Giseppina; Giliberti, Gabriele; Loddo, Roberta; La Colla, Paolo

    2006-03-15

    Aryl nucleoside 5'-H-phosphonates 4 bearing AZT or 2',3'-dideoxyuridine moieties were subjected to reaction with various aromatic aldehydes to produce nucleoside 5'-alpha-hydroxyphosphonate derivatives 2 as potential anti-HIV agents. Stability of the title compounds in cell culture media was investigated and three distinct decomposition pathways were identified. The anti-HIV activity of hydroxyphosphonates 2 correlates well with the type and extent of their chemical or enzymatic degradation in culture medium (RPMI 1640 containing 10% FBS), suggesting that aryl nucleoside 5'-hydroxyphosphonates 2 act as depot forms of the parent antiviral nucleosides.

  10. Amino Acid Ester Prodrugs of Nucleoside and Nucleotide Antivirals

    Czech Academy of Sciences Publication Activity Database

    Krečmerová, Marcela

    2017-01-01

    Roč. 17, č. 10 (2017), s. 818-833 ISSN 1389-5575 Grant - others:AV ČR(CZ) M200551201 Institutional support: RVO:61388963 Keywords : acyclic nucleoside analogues * antiherpetics * antiretrovirals * cidofovir * peptidomimetics * prodrugs Subject RIV: CC - Organic Chemistry OBOR OECD: Organic chemistry Impact factor: 2.661, year: 2016

  11. Antiviral acyclic nucleoside phosphonates: New structures and prodrugs

    Czech Academy of Sciences Publication Activity Database

    Krečmerová, Marcela; Tichý, Tomáš; Pomeisl, Karel; Andrei, G.; Balzarini, J.; Snoeck, R.

    2016-01-01

    Roč. 1, č. 2 (2016), s. 37 [PharmaMed-2016. International Conference on Medicinal and Pharmaceutical Chemistry . 05.12.2016-07.12.2016, Dubai] R&D Projects: GA ČR(CZ) GA14-00522S Institutional support: RVO:61388963 Keywords : acyclic nucleoside phosphonates * prodrugs * antivirals * 5-azacytosine Subject RIV: CC - Organic Chemistry

  12. New clinical perspectives on non-nucleoside reverse transcriptase ...

    African Journals Online (AJOL)

    New clinical perspectives on non-nucleoside reverse transcriptase inhibitors. S Miller. Abstract. No Abstract. Full Text: EMAIL FREE FULL TEXT EMAIL FREE FULL TEXT · DOWNLOAD FULL TEXT DOWNLOAD FULL TEXT · AJOL African Journals Online. HOW TO USE AJOL... for Researchers · for Librarians · for Authors ...

  13. Synthesis of (Purin-6-yl)methylphosphonate Bases and Nucleosides

    Czech Academy of Sciences Publication Activity Database

    Hasník, Zbyněk; Pohl, Radek; Hocek, Michal

    2010-01-01

    Roč. 51, č. 18 (2010), s. 2464-2466 ISSN 0040-4039 R&D Projects: GA MŠk 1M0508 Institutional research plan: CEZ:AV0Z40550506 Keywords : purines * nucleosides * phosphonates * cross-coupling Subject RIV: CC - Organic Chemistry Impact factor: 2.618, year: 2010

  14. Deoxyribonucleoside kinases activate nucleoside antibiotics in severely pathogenic bacteria

    DEFF Research Database (Denmark)

    Sandrini, Michael; Shannon, O.; Clausen, A.R.

    2007-01-01

    Common bacterial pathogens are becoming progressively more resistant to traditional antibiotics, representing a major public-health crisis. Therefore, there is a need for a variety of antibiotics with alternative modes of action. In our study, several nucleoside analogs were tested against...

  15. new clinical perspectives on non- nucleoside reverse transcriptase ...

    African Journals Online (AJOL)

    NUCLEOSIDE REVERSE TRANSCRIPTASE. INHIBITORS. TilE SOUTIIERN AFRICAN JOU NAL OF IIIV MEDICINE metabolised in the liver via the P450 cytochrome system, a pathway shared with many other compounds. This creates a risk for drug/drug interactions and the potential to modify the efficacy and/or toxicity of ...

  16. Reactivity Studies of 2,6-Ditriazolylpurine Nucleosides with Nucleophiles

    OpenAIRE

    Kovaļovs, A; Novosjolova, I; Bizdēna, Ē; Turks, M

    2012-01-01

    Reaction of 2,6-ditriazolylpurine nucleosides with nucleophiles is mild and efficient route to C6 derivatization of purine base. To explore scope and limitations of the method, we studied reactivity of various N- and S-nucleophiles as well as kinetics for selected reactions.

  17. Synthesis, bioanalysis and pharmacology of nucleoside and nucleotide analogs

    NARCIS (Netherlands)

    Jansen, R.S.

    2009-01-01

    Nucleoside analogs are an important class of drugs in anticancer and antiviral therapy. The compounds are, however, only active after intracellular conversion to their mono-, di- and triphosphate nucleotide form. In this thesis the development of sensitive liquid chromatography coupled to tandem

  18. Efficient synthesis and biological properties of the 2‘-trifluoromethyl analogues of acyclic nucleosides and acyclic nucleoside phosphonates

    Czech Academy of Sciences Publication Activity Database

    Jansa, Petr; Kolman, Viktor; Kostinová, Alexandra; Dračínský, Martin; Mertlíková-Kaiserová, Helena; Janeba, Zlatko

    2011-01-01

    Roč. 76, č. 10 (2011), s. 1187-1198 ISSN 0010-0765 R&D Projects: GA MV VG20102015046 Institutional research plan: CEZ:AV0Z40550506 Keywords : nucleosides * nucleotides * phosphorus * fluorine * biological activity * antibiotics Subject RIV: CC - Organic Chemistry Impact factor: 1.283, year: 2011

  19. Investigation of six-membered carbocyclic compounds as a molecular switch block of room temperature phosphorescence in nondeoxygenated β-cyclodextrin solution

    International Nuclear Information System (INIS)

    Zhang Hairong; Wei Yansheng; Jin Weijun; Liu Changsong

    2003-01-01

    An aerated aqueous solution, intense room temperature phosphorescence (RTP) of nitrogen heterocyclic compounds (NHCs) and polyaromatic hydrocarbons (PAHs) can be observed when micro amounts of six-membered carbocyclic compounds (6-MCCs) are introduced in β-cyclodextrin (β-CD) solution. In order to find the predominating factors of the enhanced phosphorescence observed with this novel approach, 22 typical phosphors of NHCs and PAHs were carefully screened and served as model compounds. The role of the inner heavy atom, the substituent group and the host-guest molecules space-matching on the RTP of different phosphors were investigated. The results demonstrated that the enhancement effects of cyclohexane, bromocyclohexane and cyclohexanol for the RTP of NHCs and PAHs have precedence over traditional halide alkanes such as 1,2-dibromoethane (DBE), exhibiting an obvious sequence as following: cyclohexane > bromocyclohexane > cyclohexanol. This new approach compared with other RTP methods is simple, convenient and fast

  20. The First Synthesis of Dually Modified Southern-Mimicking Nucleoside: 4'-Methyl Branched and Bicyclo [3.1.0] Hexane Locked Nucleoside

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Kwan Woo; Hong, Joon Hee [Chosun University, Gwangju (Korea, Republic of)

    2004-05-15

    This paper reports the stereoselective synthesis of a novel nucleoside, 4'-methyl branched and bicyclo [3.1.0] hexane locked-nucleoside 12, using a sequential [3,3]-sigmatropic rearrangement/carbene cycloaddition reaction/Curtius reaction strategy with a high stereoselectivity.

  1. Locked and unlocked nucleosides in functional nucleic acids

    DEFF Research Database (Denmark)

    Doessing, Holger; Vester, Birte

    2011-01-01

    Nucleic acids are able to adopt a plethora of structures, many of which are of interest in therapeutics, bio- or nanotechnology. However, structural and biochemical stability is a major concern which has been addressed by incorporating a range of modifications and nucleoside derivatives. This rev......Nucleic acids are able to adopt a plethora of structures, many of which are of interest in therapeutics, bio- or nanotechnology. However, structural and biochemical stability is a major concern which has been addressed by incorporating a range of modifications and nucleoside derivatives....... This review summarizes the use of locked nucleic acid (LNA) and un-locked nucleic acid (UNA) monomers in functional nucleic acids such as aptamers, ribozymes, and DNAzymes....

  2. Evaluation of Anti-HIV-1 Mutagenic Nucleoside Analogues*

    Science.gov (United States)

    Vivet-Boudou, Valérie; Isel, Catherine; El Safadi, Yazan; Smyth, Redmond P.; Laumond, Géraldine; Moog, Christiane; Paillart, Jean-Christophe; Marquet, Roland

    2015-01-01

    Because of their high mutation rates, RNA viruses and retroviruses replicate close to the threshold of viability. Their existence as quasi-species has pioneered the concept of “lethal mutagenesis” that prompted us to synthesize pyrimidine nucleoside analogues with antiviral activity in cell culture consistent with an accumulation of deleterious mutations in the HIV-1 genome. However, testing all potentially mutagenic compounds in cell-based assays is tedious and costly. Here, we describe two simple in vitro biophysical/biochemical assays that allow prediction of the mutagenic potential of deoxyribonucleoside analogues. The first assay compares the thermal stabilities of matched and mismatched base pairs in DNA duplexes containing or not the nucleoside analogues as follows. A promising candidate should display a small destabilization of the matched base pair compared with the natural nucleoside and the smallest gap possible between the stabilities of the matched and mismatched base pairs. From this assay, we predicted that two of our compounds, 5-hydroxymethyl-2′-deoxyuridine and 5-hydroxymethyl-2′-deoxycytidine, should be mutagenic. The second in vitro reverse transcription assay assesses DNA synthesis opposite nucleoside analogues inserted into a template strand and subsequent extension of the newly synthesized base pairs. Once again, only 5-hydroxymethyl-2′-deoxyuridine and 5-hydroxymethyl-2′-deoxycytidine are predicted to be efficient mutagens. The predictive potential of our fast and easy first line screens was confirmed by detailed analysis of the mutation spectrum induced by the compounds in cell culture because only compounds 5-hydroxymethyl-2′-deoxyuridine and 5-hydroxymethyl-2′-deoxycytidine were found to increase the mutation frequency by 3.1- and 3.4-fold, respectively. PMID:25398876

  3. Evaluation of anti-HIV-1 mutagenic nucleoside analogues.

    Science.gov (United States)

    Vivet-Boudou, Valérie; Isel, Catherine; El Safadi, Yazan; Smyth, Redmond P; Laumond, Géraldine; Moog, Christiane; Paillart, Jean-Christophe; Marquet, Roland

    2015-01-02

    Because of their high mutation rates, RNA viruses and retroviruses replicate close to the threshold of viability. Their existence as quasi-species has pioneered the concept of "lethal mutagenesis" that prompted us to synthesize pyrimidine nucleoside analogues with antiviral activity in cell culture consistent with an accumulation of deleterious mutations in the HIV-1 genome. However, testing all potentially mutagenic compounds in cell-based assays is tedious and costly. Here, we describe two simple in vitro biophysical/biochemical assays that allow prediction of the mutagenic potential of deoxyribonucleoside analogues. The first assay compares the thermal stabilities of matched and mismatched base pairs in DNA duplexes containing or not the nucleoside analogues as follows. A promising candidate should display a small destabilization of the matched base pair compared with the natural nucleoside and the smallest gap possible between the stabilities of the matched and mismatched base pairs. From this assay, we predicted that two of our compounds, 5-hydroxymethyl-2'-deoxyuridine and 5-hydroxymethyl-2'-deoxycytidine, should be mutagenic. The second in vitro reverse transcription assay assesses DNA synthesis opposite nucleoside analogues inserted into a template strand and subsequent extension of the newly synthesized base pairs. Once again, only 5-hydroxymethyl-2'-deoxyuridine and 5-hydroxymethyl-2'-deoxycytidine are predicted to be efficient mutagens. The predictive potential of our fast and easy first line screens was confirmed by detailed analysis of the mutation spectrum induced by the compounds in cell culture because only compounds 5-hydroxymethyl-2'-deoxyuridine and 5-hydroxymethyl-2'-deoxycytidine were found to increase the mutation frequency by 3.1- and 3.4-fold, respectively. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. C-Nucleosides: Synthetic Strategies and Biological Applications

    Czech Academy of Sciences Publication Activity Database

    Štambaský, J.; Hocek, Michal; Kočovský, P.

    2009-01-01

    Roč. 109, č. 12 (2009), s. 6729-6764 ISSN 0009-2665 R&D Projects: GA MŠk LC512; GA AV ČR IAA400550902 Grant - others:NIH(US) 1R03TW007372-01 Institutional research plan: CEZ:AV0Z40550506 Keywords : nucleosides * nucleobases * biological activity * extension of genetic alphabet Subject RIV: CC - Organic Chemistry Impact factor: 35.957, year: 2009

  5. Antiviral acyclic nucleoside phosphonates: A novel group of immunomodulatroy agents

    Czech Academy of Sciences Publication Activity Database

    Zídek, Zdeněk; Franková, Daniela; Holý, A.

    2002-01-01

    Roč. 22, č. 1 (2002), s. 166-167 ISSN 1079-9907. [Cytokines and Interferons 2002. Turin - Italy, 06.10.2002-10.10.2002] R&D Projects: GA ČR GA305/00/0048 Institutional research plan: CEZ:AV0Z5008914 Keywords : nucleoside phosphonates Subject RIV: FR - Pharmacology ; Medidal Chemistry Impact factor: 1.885, year: 2002

  6. Nucleoside-Lipid-Based Nanocarriers for Sorafenib Delivery

    Science.gov (United States)

    Benizri, Sebastien; Ferey, Ludivine; Alies, Bruno; Mebarek, Naila; Vacher, Gaelle; Appavoo, Ananda; Staedel, Cathy; Gaudin, Karen; Barthélémy, Philippe

    2018-01-01

    Although the application of sorafenib, a small inhibitor of tyrosine protein kinases, to cancer treatments remains a worldwide option in chemotherapy, novel strategies are needed to address the low water solubility (use of nanocarriers is currently investigated in order to overcome these drawbacks. In this contribution, we report a new type of sorafenib-based nanoparticles stabilized by hybrid nucleoside-lipids. The solid lipid nanoparticles (SLNs) showed negative or positive zeta potential values depending on the nucleoside-lipid charge. Transmission electron microscopy of sorafenib-loaded SLNs revealed parallelepiped nanoparticles of about 200 nm. Biological studies achieved on four different cell lines, including liver and breast cancers, revealed enhanced anticancer activities of Sorafenib-based SLNs compared to the free drug. Importantly, contrast phase microscopy images recorded after incubation of cancer cells in the presence of SLNs at high concentration in sorafenib (> 80 μM) revealed a total cancer cell death in all cases. These results highlight the potential of nucleoside-lipid-based SLNs as drug delivery systems.

  7. [Determination of Nucleosides and HPLC Fingerprints of Cordyceps].

    Science.gov (United States)

    Wang, Bing; Li, Ning; Dong, Ting-xia; Zhan, Hua-qiang

    2015-05-01

    To establish the HPLC fingerprints method of Cordyceps and to determine the contents of uridine, inosine, guanosine and adenosine. The HPLC separation was performed on a Grace Prevail C18 column( 150 mm x 4.6 mm, 5 μm) in a gradient elution mode with a mixture consisting of water and methanol at a flow rate of 1.0 mL/min, the detection wavelength was set at 260 nm, the column temperature was 25 degrees C. The contents of four nucleosides were determined in Cordyceps from different habitats, and the HPLC fingerprint of Cordyceps was set up with 13 common peaks. Among of them, uridine, inosine, guanosine and adenosine were identified. The similarities of ten fingerprints were greater than 0.95 with good separation of each chromatographic peak, and met the requirement of the fingerprints. There were similar results in cluster analysis and principal component analysis of the major nucleosides and the fingerprints of 10 batches of Cordyceps. The results of sample classification in principal component analysis showed a good similarity with cluster analysis. This method showed the information of chemical composition in Cordyceps, with good repeatability and similarity between samples, indicating that the stable chemical distribution and proportion of the major nucleosides in the medical materials. Fingerprints, principal component analysis and cluster analysis, which are applied to identify the different sources of Cordyceps, provide an experimental basis for establishing the characteristics evaluation methodology of medicinal materials.

  8. Synthesis of pyrimidine containing nucleoside β-(R/S)-hydroxyphosphonate analogues

    OpenAIRE

    Meurillon, Maïa; Chaloin, Laurent; Peyrottes, Suzanne; Périgaud, Christian

    2011-01-01

    A concise route to nucleoside β-hydroxyphosphonate analogues is described. The use of a nucleoside β-ketophosphonate as the key intermediate allowed both the (R) and (S) isomers of β-hydroxyphosphonate analogues in the pyrimidine series to be accessed. Such derivatives may be considered as stable mimics of 5′-monophosphate nucleosides and, therefore, could be the starting point for the development of potential therapeutic agents.

  9. Synthesis and antiproliferative evaluation of novel azido nucleosides and their phosphoramidate derivatives

    Czech Academy of Sciences Publication Activity Database

    Xavier, N.M.; Goncalves-Pereira, R.; Jorda, Radek; Řezníčková, Eva; Kryštof, Vladimír; Oliveira, M.C.

    2017-01-01

    Roč. 89, č. 9 (2017), s. 1267-1281 ISSN 0033-4545 R&D Projects: GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : anticancer activity * azido nucleosides * bioactive molecules * ics-28 * N-glycosylation * nucleoside phosphoramidates * nucleoside/nucleotide analogs * Staudinger-phosphite reaction Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Organic chemistry Impact factor: 2.626, year: 2016

  10. Nucleosides of 4-methylthio-1,2,3-triazol-5-yl-carboxylic acid derivatives

    International Nuclear Information System (INIS)

    Shingarova, I.D.; Yartseva, I.V.; Preobrazhenskaya, M.N.

    1987-01-01

    2-β-D-Ribofuranosyl-4-methylthio-5-methoxycarbonyl-1,2,3-triazole was obtained by fusing 4-methylthio-5-methoxycarbonyl-1,2,3-triazole together with tetraacyl-D-ribofuranose, followed by deacylation, and its amide and hydrazide were prepared. The structures of the new nucleosides were established by converting them into known 2-nucleosides of 1,2,3-triazol-4-yl-carboxylic acid derivatives. By comparing PMR spectra with previously reported PMR spectra for the isomeric 1- and 2-nucleosides of 1,2,3-triazol-4-yl-carboxylic acid derivatives, the synthesized nucleosides could be assigned to 2-substituted triazoles

  11. Two purine nucleoside phosphorylases in Bacillus subtilis. Purification and some properties of the adenosine-specific phosphorylase

    DEFF Research Database (Denmark)

    Jensen, Kaj Frank

    1978-01-01

    Two purine nucleoside phosphorylases (purine-nucleoside:orthophosphate ribosyltransferase, EC 2.4.2.1) were purified from vegetative Bacillus subtilis cells. One enzyme, inosine-guanosine phosphorylase, showed great similarity to the homologous enzyme of Bacillus cereus. It appeared...

  12. An enzymatic glycosylation of nucleoside analogues using beta-galactosidase from Escherichia coli

    Czech Academy of Sciences Publication Activity Database

    Blažek, Jiří; Jansa, Petr; Baszczyňski, Ondřej; Kaiser, Martin Maxmilian; Otmar, Miroslav; Krečmerová, Marcela; Dračínský, Martin; Holý, Antonín; Králová, B.

    2012-01-01

    Roč. 20, č. 9 (2012), s. 3111-3118 ISSN 0968-0896 R&D Projects: GA MŠk 1M0508 Institutional research plan: CEZ:AV0Z40550506 Keywords : glycosylation * galactosylation * beta-galactosidase * enzymatic synthesis * nucleoside * acyclic nucleoside analogues Subject RIV: CC - Organic Chemistry Impact factor: 2.903, year: 2012

  13. Nucleoside phosphonic acids in thymidine phosphorylase inhibition: Structure - activity relationship

    Czech Academy of Sciences Publication Activity Database

    Panova, Natalya; Kóšiová, Ivana; Petrová, Magdalena; Vaněk, Václav; Liboska, Radek; Kovačková, Soňa; Kočalka, Petr; Králíková, Šárka; Točík, Zdeněk; Páv, Ondřej; Pačes, Ondřej; Rejman, Dominik; Rosenberg, Ivan

    -, č. 52 (2008), s. 665-666 ISSN 0261-3166. [Joint Symposium of the International Roundtable on Nucleosides, Nucleotides and Nucleic Acids /18./ and the International Symposium on Nucleic Acid Chemistry /35./. Kyoto, 08.09.2008-12.09.2008] R&D Projects: GA MŠk(CZ) LC06061; GA MŠk(CZ) LC06077; GA MŠk LC512 Institutional research plan: CEZ:AV0Z40550506 Keywords : thymidine phosphorylase * inhibitors * phosphonic acids Subject RIV: CC - Organic Chemistry

  14. Purine nucleoside phosphorylase deficiency in two unrelated Saudi patients

    International Nuclear Information System (INIS)

    Alangari, Abdullah; AlHarbi, Abdullah; AlGhonaium Abdulaziz; Santisteban, Ines; Hershfield, Michael

    2009-01-01

    Purine nucleoside phosphorylase (PNP) deficiency is a rare autosomal recessive metabolic disorder that results in combined immunodeficiency, neurologic dysfunction and autoimmunity. PNP deficiency has never been reported from Saudi Arabia or in patients with an Arabic ethnic background. We report on two Saudi girls with PNP deficiency. Both showed severe lymphopenia and neurological involvement. Sequencing of the PNP gene of one girl revealed a novel missense mutation Pro146>Leu in exon 4 due to a change in the codon from CCT>CTT. Expression of PNP (146L) cDNA in E coli indicated that the mutation greatly reduced, but did not completely eliminate PNP activity. (author)

  15. Purine nucleoside analogues inhibit the repair of radiation-induced potentially lethal damage in mammalian cells in culture

    International Nuclear Information System (INIS)

    Nakatsugawa, S.; Kumar, A.

    1982-01-01

    Three purine nucleoside analogues were found to be much more effective than pyrimidine nucleoside analogues in the screening of PLD repair inhibitors using X-irradiated Chinese hamster HA-1 cells. Among the three purine nucleoside analogues, 3'-dG was the most effective at a non-toxic concentration. (author)

  16. Metabolism of the carbocyclic analogue of (E)-5-(2-iodovinyl)-2'-deoxyuridine in herpes simplex virus-infected cells. Incorporation of C-IVDU into DNA

    International Nuclear Information System (INIS)

    De Clercq, E.; Bernaerts, R.; Balzarini, J.; Herdewijn, P.; Verbruggen, A.

    1985-01-01

    The carbocyclic analogues of (E)-5-(2-bromovinyl)-2'-deoxyuridine (BVDU) and (E)-5-(2-iodovinyl)-2'-deoxyuridine (IVDU), in which the sugar moiety is replaced by a cyclopentane ring and which have been designated as C-BVDU and C-IVDU, respectively, are, like their parent compounds BVDU and IVDU, potent and selective inhibitors of herpes simplex virus type 1 (HSV-1) and, to a lesser extent, herpes simplex virus type 2 (HSV-2) replication. The authors have now synthesized the radiolabeled C-IVDU analogue, C-[ 125 I]IVDU, and determined its metabolism by HSV-infected and mock-infected Vero cells. C-[ 125 I]IVDU was effectively phosphorylated by HSV-1-infected cells and, to a lesser extent, HSV-2-infected cells. C-[ 125 I]IVDU was not phosphorylated to an appreciable extent by either mock-infected cells or cells that had been infected with a thymidine kinase-deficient mutant of HSV-1. Furthermore, C-[ 125 I]IVDU was incorporated into both viral and cellular DNA of HSV-1-infected Vero cells. This finding represents the first demonstration of the incorporation of a cyclopentylpyrimidine into DNA

  17. Design and Synthesis of Polyimides Based on Carbocyclic Pseudo-Tröger’s Base-Derived Dianhydrides for Membrane Gas Separation Applications

    KAUST Repository

    Ma, Xiaohua

    2017-07-24

    Two novel carbocyclic pseudo-Tröger’s base-derived dianhydrides, 5,6,11,12-tetrahydro-5,11-methanodibenzo[a,e][8]annulene-2,3,8,9-tetracarboxylic anhydride (CTB1) and its dione-substituted analogue 6,12-dioxo-5,6,11,12-tetrahydro-5,11-methanodibenzo[a,e][8]annulene-2,3,8,9-tetracarboxylic dianhydride (CTB2), were made and used for the synthesis of soluble polyimides of intrinsic microporosity with 3,3′-dimethylnaphthidine (DMN). The polyimides CTB1-DMN and CTB2-DMN exhibited excellent thermal stability of ∼500 °C and high BET surface areas of 580 and 469 m2 g–1, respectively. A freshly made dione-substituted CTB2-DMN membrane demonstrated promising gas separation performance with O2 permeability of 206 barrer and O2/N2 selectivity of 5.2. A higher O2 permeability of 320 barrer and lower O2/N2 selectivity of 4.2 were observed for a fresh CTB1-DMN film due to its higher surface area and less tightly packed structure as indicated by weaker charge-transfer complex interactions. Physical aging over 60 days resulted in reduction in gas permeability and moderately enhanced selectivity. CTB2-DMN exhibited notable performance with gas permeation data located between the 2008 and 2015 permeability/selectivity upper bounds for O2/N2 and H2/CH4.

  18. Nucleoside Derived Antibiotics to Fight Microbial Drug Resistance: New Utilities for an Established Class of Drugs?

    Science.gov (United States)

    Serpi, Michaela; Ferrari, Valentina; Pertusati, Fabrizio

    2016-12-08

    Novel antibiotics are urgently needed to combat the rise of infections due to drug-resistant microorganisms. Numerous natural nucleosides and their synthetically modified analogues have been reported to have moderate to good antibiotic activity against different bacterial and fungal strains. Nucleoside-based compounds target several crucial processes of bacterial and fungal cells such as nucleoside metabolism and cell wall, nucleic acid, and protein biosynthesis. Nucleoside analogues have also been shown to target many other bacterial and fungal cellular processes although these are not well characterized and may therefore represent opportunities to discover new drugs with unique mechanisms of action. In this Perspective, we demonstrate that nucleoside analogues, cornerstones of anticancer and antiviral treatments, also have great potential to be repurposed as antibiotics so that an old drug can learn new tricks.

  19. Nucleosides as a carbon source in Bacillus subtilis: characterization of the drm-pupG operon.

    Science.gov (United States)

    Schuch, R; Garibian, A; Saxild, H H; Piggot, P J; Nygaard, P

    1999-10-01

    In Bacillus subtilis, nucleosides are readily taken up from the growth medium and metabolized. The key enzymes in nucleoside catabolism are nucleoside phosphorylases, phosphopentomutase, and deoxyriboaldolase. The characterization of two closely linked loci, drm and pupG, which encode phosphopentomutase (Drm) and guanosine (inosine) phosphorylase (PupG), respectively, is reported here. When expressed in Escherichia coli mutant backgrounds, drm and pupG confer phosphopentomutase and purine-nucleoside phosphorylase activity. Northern blot and enzyme analyses showed that drm and pupG form a dicistronic operon. Both enzymes are induced when nucleosides are present in the growth medium. Using mutants deficient in nucleoside catabolism, it was demonstrated that the low-molecular-mass effectors of this induction most likely were deoxyribose 5-phosphate and ribose 5-phosphate. Both Drm and PupG activity levels were higher when succinate rather than glucose served as the carbon source, indicating that the expression of the operon is subject to catabolite repression. Primer extension analysis identified two transcription initiation signals upstream of drm; both were utilized in induced and non-induced cells. The nucleoside-catabolizing system in B. subtilis serves to utilize the base for nucleotide synthesis while the pentose moiety serves as the carbon source. When added alone, inosine barely supports growth of B. subtilis. This slow nucleoside catabolism contrasts with that of E. coli, which grows rapidly on a nucleoside as a carbon source. When inosine was added with succinate or deoxyribose, however, a significant increase in growth was observed in B. subtilis. The findings of this study therefore indicate that the B. subtilis system for nucleoside catabolism differs greatly from the well-studied system in E. coli.

  20. Alkylsulfanylphenyl derivatives of cytosine and 7-deazaadenine nucleosides, nucleotides and nucleoside triphosphates. Synthesis, polymerase incorporation to DNA and electrochemical study

    Czech Academy of Sciences Publication Activity Database

    Macíčková-Cahová, Hana; Pohl, Radek; Horáková Brázdilová, Petra; Havran, Luděk; Špaček, Jan; Fojta, Miroslav; Hocek, Michal

    2011-01-01

    Roč. 17, č. 21 (2011), s. 5833-5841 ISSN 0947-6539 R&D Projects: GA MŠk(CZ) LC06035; GA MŠk LC512; GA ČR GA203/09/0317; GA AV ČR(CZ) IAA400040901 Institutional research plan: CEZ:AV0Z40550506; CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : DNA polymerases * electrochemistry * nucleosides * nucleotides * organosulfur compounds Subject RIV: CC - Organic Chemistry Impact factor: 5.925, year: 2011

  1. Rhodium-catalyzed [5 + 2 + 1] cycloaddition of ene-vinylcyclopropanes and CO: reaction design, development, application in natural product synthesis, and inspiration for developing new reactions for synthesis of eight-membered carbocycles.

    Science.gov (United States)

    Wang, Yi; Yu, Zhi-Xiang

    2015-08-18

    Practical syntheses of natural products and their analogues with eight-membered carbocyclic skeletons are important for medicinal and biological investigations. However, methods and strategies to construct the eight-membered carbocycles are limited. Therefore, developing new methods to synthesize the eight-membered carbocycles is highly desired. In this Account, we describe our development of three rhodium-catalyzed cycloadditions for the construction of the eight-membered carbocycles, which have great potential in addressing the challenges in the synthesis of medium-sized ring systems. The first reaction described in this Account is our computationally designed rhodium-catalyzed two-component [5 + 2 + 1] cycloaddition of ene-vinylcyclopropanes (ene-VCPs) and CO for the diastereoselective construction of bi- and tricyclic cyclooctenones. The design of this reaction is based on the hypothesis that the C(sp(3))-C(sp(3)) reductive elimination of the eight-membered rhodacycle intermediate generated from the rhodium-catalyzed cyclopropane cleavage and alkene insertion, giving Wender's [5 + 2] cycloadduct, is not easy. Under CO atmosphere, CO insertion may occur rapidly, converting the eight-membered rhodacycle into a nine-membered rhodacycle, which then undergoes an easy C(sp(2))-C(sp(3)) reductive elimination process and furnishes the [5 + 2 + 1] product. This hypothesis was supported by our preliminary DFT studies and also served as inspiration for the development of two [7 + 1] cycloadditions: the [7 + 1] cycloaddition of buta-1,3-dienylcyclopropanes (BDCPs) and CO for the construction of cyclooctadienones, and the benzo/[7 + 1] cycloaddition of cyclopropyl-benzocyclobutenes (CP-BCBs) and CO to synthesize the benzocyclooctenones. The efficiency of these rhodium-catalyzed cycloadditions can be revealed by the application in natural product synthesis. Two eight-membered ring-containing natural products, (±)-asterisca-3(15),6-diene and (+)-asteriscanolide, have been

  2. Enhancement of Nucleoside Production in Hirsutella sinensis Based on Biosynthetic Pathway Analysis

    Science.gov (United States)

    Liu, Zhi-Qiang; Zhang, Bo; Lin, Shan; Baker, Peter James; Chen, Mao-Sheng; Xue, Ya-Ping; Wu, Hui; Xu, Feng; Yuan, Shui-Jin; Teng, Yi; Wu, Ling-Fang

    2017-01-01

    To enhance nucleoside production in Hirsutella sinensis, the biosynthetic pathways of purine and pyrimidine nucleosides were constructed and verified. The differential expression analysis showed that purine nucleoside phosphorylase, inosine monophosphate dehydrogenase, and guanosine monophosphate synthase genes involved in purine nucleotide biosynthesis were significantly upregulated 16.56-fold, 8-fold, and 5.43-fold, respectively. Moreover, dihydroorotate dehydrogenase, uridine nucleosidase, uridine/cytidine monophosphate kinase, and inosine triphosphate pyrophosphatase genes participating in pyrimidine nucleoside biosynthesis were upregulated 4.53-fold, 10.63-fold, 4.26-fold, and 5.98-fold, respectively. To enhance the nucleoside production, precursors for synthesis of nucleosides were added based on the analysis of biosynthetic pathways. Uridine and cytidine contents, respectively, reached 5.04 mg/g and 3.54 mg/g when adding 2 mg/mL of ribose, resulting in an increase of 28.6% and 296% compared with the control, respectively. Meanwhile, uridine and cytidine contents, respectively, reached 10.83 mg/g 2.12 mg/g when adding 0.3 mg/mL of uracil, leading to an increase of 176.3% and 137.1%, respectively. This report indicated that fermentation regulation was an effective way to enhance the nucleoside production in H. sinensis based on biosynthetic pathway analysis. PMID:29333435

  3. Aqueous microwaves assisted cross-coupling reactions applied to unprotected nucleosides.

    Directory of Open Access Journals (Sweden)

    CHRISTOPHE eLEN

    2015-02-01

    Full Text Available Nucleoside analogues have attracted much attention due to their potential biological activities. Amongst all synthetic nucleosides, C5-modified pyrimidines and C7- or C8-modified purines have mostly been prepared using palladium cross-coupling reactions and then studied as antitumoral and antiviral agents. Our objective is to focus this review on the Suzuki-Miyaura and on the Heck cross-couplings of nucleosides using microwave irradiations which are an alternative technology compatible with green chemistry and sustainable development.

  4. Identification of a novel compound that inhibits osteoclastogenesis by suppressing nucleoside transporters.

    Science.gov (United States)

    Katsuyama, Shun; Sugino, Kumi; Sasazawa, Yukiko; Nakano, Yoshihiko; Aono, Harumi; Morishita, Keisuke; Kawatani, Makoto; Umezawa, Kazuo; Osada, Hiroyuki; Simizu, Siro

    2016-04-01

    We screened small-molecule compounds that inhibit osteoclast differentiation to find new anti-osteoporosis agents and found that a novel compound, SUKU-1, suppressed osteoclastogenesis. We also synthesized 38 derivatives of SUKU-1 and discovered that nine of them had inhibitory effects on osteoclastogenesis and that SUKU-33 was the most potent inhibitor. Next, we investigated the mechanisms by which SUKU-33 suppressed osteoclast differentiation. By measuring the uptake of [(3) H]-uridine in cells, we found that SUKU-33 suppressed both equilibrative nucleoside transporters and concentrative nucleoside transporters. These results suggest that SUKU-33 inhibits osteoclast differentiation by suppressing nucleoside transporters. © 2016 Federation of European Biochemical Societies.

  5. A simple chemical synthesis of sugar nucleoside diphosphates in water.

    Science.gov (United States)

    Tanaka, Hidenori; Yoshimura, Yayoi; Hindsgaul, Ole

    2013-10-08

    Chemoenzymatic oligosaccharide synthesis is attractive since it eliminates the tedious multistep protection-deprotection requirements of pure chemical synthesis. Chemoenzymatic synthesis using glycosyltransferases, however, requires not only the correct enzyme to control both regio- and stereospecificity, but also the glycosyl donor to provide the sugar that is added. This unit describes a simple synthesis of sugar-nucleoside diphosphates (sugar-NDPs), the type of glycosyl donor (e.g., UDP-Glc, UDP-Gal, ADP-Glc) required by most glycosyltransferases, by using a chemical coupling reaction in water. The preparation of sugar-NDPs by this method therefore does not require any skills in synthetic organic chemistry. Copyright © 2013 John Wiley & Sons, Inc.

  6. Electrospray ionization mass spectrometric study of thallium complexes with nucleosides.

    Science.gov (United States)

    Frańska, Magdalena

    2017-10-01

    The complexes between Tl + , K + , and nucleosides were studied by using electrospray ionization mass spectrometry. It was found that for complexes of 1:1 stoichiometry, thallium complexes with cytidine were the most abundant and thallium complexes with guanosine were the second most abundant ones. The relative abundances of cytidine-Tl + to cytidine-K + complexes depended on stoichiometry (at higher stoichiometry the potassium complexes were more abundant). In other words, the relative affinity of Tl + and K + to form cytidine complexes depends on the stoichiometry of the formed complexes. Guanosine-Tl + complexes were more abundant than guanosine-K + complexes, irrespective of stoichiometry. Both guanosine tetramer and mixed cytidine/guanosine tetramer were more abundant when they were stabilized by thallium than potassium. Therefore, Tl + may affect the K + stabilization of these tetramers.

  7. Attenuation of nucleoside and anti-cancer nucleoside analog drug uptake in prostate cancer cells by Cimicifuga racemosa extract BNO-1055.

    Science.gov (United States)

    Dueregger, Andrea; Guggenberger, Fabian; Barthelmes, Jan; Stecher, Günther; Schuh, Markus; Intelmann, Daniel; Abel, Gudrun; Haunschild, Jutta; Klocker, Helmut; Ramoner, Reinhold; Sampson, Natalie

    2013-11-15

    This study aimed to investigate the mechanisms underlying the anti-proliferative effects of the ethanolic Cimicifuga racemosa extract BNO-1055 on prostate cells and evaluate its therapeutic potential. BNO-1055 dose-dependently attenuated cellular uptake and incorporation of thymidine and BrdU and significantly inhibited cell growth after long-time exposure. Similar results were obtained using saponin-enriched sub-fractions of BNO-1055. These inhibitory effects of BNO-1055 could be mimicked using pharmacological inhibitors and isoform-specific siRNAs targeting the equilibrative nucleoside transporters ENT1 and ENT2. Moreover, BNO-1055 attenuated the uptake of clinically relevant nucleoside analogs, e.g. the anti-cancer drugs gemcitabine and fludarabine. Consistent with inhibition of the salvage nucleoside uptake pathway BNO-1055 potentiated the cytotoxicity of the de novo nucleotide synthesis inhibitor 5-FU without significantly altering its uptake. Collectively, these data show for the first time that the anti-proliferative effects of BNO-1055 result from hindered nucleoside uptake due to impaired ENT activity and demonstrate the potential therapeutic use of BNO-1055 for modulation of nucleoside transport. Copyright © 2013 Elsevier GmbH. All rights reserved.

  8. Comparison of nucleoside concentrations in blood of fish with and without tumors

    Energy Technology Data Exchange (ETDEWEB)

    Kuehl, D.W.; Johnson, R.D. (Environmental Protection Agency, Duluth, MN (United States)); Eisenschenk, L.; Naumann, S. (Univ. of Wisconsin, Superior (United States)); Regal, R.; Barnidge, P. (Univ. of Minnesota, Duluth (United States)); McKim, J. Jr. (Oregon State Univ., Corvallis (United States))

    1991-05-01

    The objective of this study was to develop and use HPLC based analytical methodology to characterize nucleosides in blood plasma and serum from fish with and without tumors, with a goal of determining if fish blood nucleoside concentrations could similarly be used as a bioindicator of tumor development in fish. The approach was to develop analytical methodology and quality assurance criteria for the analysis of nucleosides in fish blood, and to characterize nucleoside concentrations in blood of fish for which both healthy and tumor-bearing samples were available. Data would then be used to establish parameters with which tumor-bearing fish could be distinguished from healthy fish. Blood samples used to establish the diagnostic parameters were from control rainbow trout (Oncorhynchus mykiss) and those with tumors developed after exposure to aflatoxins. A second set of blood samples was from field collected black bullheads (Ictalurus melas).

  9. Modification of Purine and Pyrimidine Nucleosides by Direct C-H Bond Activation

    Directory of Open Access Journals (Sweden)

    Yong Liang

    2015-03-01

    Full Text Available Transition metal-catalyzed modifications of the activated heterocyclic bases of nucleosides as well as DNA or RNA fragments employing traditional cross-coupling methods have been well-established in nucleic acid chemistry. This review covers advances in the area of cross-coupling reactions in which nucleosides are functionalized via direct activation of the C8-H bond in purine and the C5-H or C6-H bond in uracil bases. The review focuses on Pd/Cu-catalyzed couplings between unactivated nucleoside bases with aryl halides. It also discusses cross-dehydrogenative arylations and alkenylations as well as other reactions used for modification of nucleoside bases that avoid the use of organometallic precursors and involve direct C-H bond activation in at least one substrate. The scope and efficiency of these coupling reactions along with some mechanistic considerations are discussed.

  10. Molecular moment similarity between several nucleoside analogs of thymidine and thymidine. sil@watson.ibm.com.

    Science.gov (United States)

    Silverman, B D; Pitman, M C; Platt, D E

    1999-06-01

    Molecular moment descriptors of the shape and charge distributions of twenty five nucleoside structures have been examined. The structures include thymidine as well as the difluorotoluene nucleoside analog which has been found to pair efficiently with adenine by polymerase catalysis. The remaining twenty three structures have been chosen to be as structurally similar to thymidine and to the difluorotoluene nucleoside analog as possible. The moment descriptors which include a description of the relationship of molecular charge to shape show the difluorotoluene nucleoside to be one of the most proximate molecules to thymidine in the space of the molecular moments. The calculations, therefore, suggest that polymerase specificity might be not only a consequence of molecular steric features alone but also of the molecular electrostatic environment and its registration with molecular shape.

  11. The preparation of trisubstituted alkenyl nucleoside phosphonates under ultrasound-assisted olefin cross-metathesis.

    Science.gov (United States)

    Sari, Ozkan; Hamada, Manabu; Roy, Vincent; Nolan, Steven P; Agrofoglio, Luigi A

    2013-09-06

    Intermolecular ultrasound-assisted olefin cross-metathesis is reported. This approach allows an easy access to challenging trisubstituted alkenyl nucleoside phosphonates. Regioselective chemoenzymatic deacetylation and Mitsunobu coupling are also described.

  12. Peptides derived from nucleoside beta-amino acids form an unusual 8-helix.

    Science.gov (United States)

    Threlfall, Richard; Davies, Andrew; Howarth, Nicola M; Fisher, Julie; Cosstick, Richard

    2008-02-07

    Peptides of varying length (dimers to octamers) were prepared from nucleoside beta-amino acids and conformational studies, based on NOE observations, show that the beta-peptides form an unusual 8-helix.

  13. [Bicyclic furano[2,3-D] derivatives of pyrimidine nucleosides--synthesis and antiviral properties].

    Science.gov (United States)

    Ivanov, M A; Aleksandrova, L A

    2013-01-01

    The methods of synthesis of furano- and pyrrolo[2,3-dlpyrimidine nucleosides as well as structure activity relationship of obtained compounds towards viruses of varicella zoster, hepatitis C, bovine viral diarrhea and some others are reviewed.

  14. Cell free phosphorylation method to assess the utility of new nucleotides as nucleoside reserve transcriptase inhibitors

    CSIR Research Space (South Africa)

    Lebea, Phiyani J

    2007-06-01

    Full Text Available The acyclic nucleoside phosphonates such as cidofovir, adefovir and tenofovir have proved to be effective in vitro (cell culture systems) and in vivo (animal models, clinical studies) against a wide variety of DNA virus and retrovirus infections...

  15. Enzymatic synthesis and phosphorolysis of 4(2-thioxo- and 6(5-azapyrimidine nucleosides by E. coli nucleoside phosphorylases

    Directory of Open Access Journals (Sweden)

    Vladimir A. Stepchenko

    2016-12-01

    Full Text Available The trans-2-deoxyribosylation of 4-thiouracil (4SUra and 2-thiouracil (2SUra, as well as 6-azauracil, 6-azathymine and 6-aza-2-thiothymine was studied using dG and E. coli purine nucleoside phosphorylase (PNP for the in situ generation of 2-deoxy-α-D-ribofuranose-1-phosphate (dRib-1P followed by its coupling with the bases catalyzed by either E. coli thymidine (TP or uridine (UP phosphorylases. 4SUra revealed satisfactory substrate activity for UP and, unexpectedly, complete inertness for TP; no formation of 2’-deoxy-2-thiouridine (2SUd was observed under analogous reaction conditions in the presence of UP and TP. On the contrary, 2SU, 2SUd, 4STd and 2STd are good substrates for both UP and TP; moreover, 2SU, 4STd and 2’-deoxy-5-azacytidine (Decitabine are substrates for PNP and the phosphorolysis of the latter is reversible. Condensation of 2SUra and 5-azacytosine with dRib-1P (Ba salt catalyzed by the accordant UP and PNP in Tris∙HCl buffer gave 2SUd and 2’-deoxy-5-azacytidine in 27% and 15% yields, respectively. 6-Azauracil and 6-azathymine showed good substrate properties for both TP and UP, whereas only TP recognizes 2-thio-6-azathymine as a substrate. 5-Phenyl and 5-tert-butyl derivatives of 6-azauracil and its 2-thioxo derivative were tested as substrates for UP and TP, and only 5-phenyl- and 5-tert-butyl-6-azauracils displayed very low substrate activity. The role of structural peculiarities and electronic properties in the substrate recognition by E. coli nucleoside phosphorylases is discussed.

  16. Ring Enlargement of Cyclic Acetals and Ketals: A Way to Seven-Membered Nucleoside Phostones

    Czech Academy of Sciences Publication Activity Database

    Páv, Ondřej; Barvík, I.; Buděšínský, Miloš; Masojídková, Milena; Rosenberg, Ivan

    2007-01-01

    Roč. 9, č. 26 (2007), s. 5469-5472 ISSN 1523-7060 R&D Projects: GA ČR GA203/05/0827; GA ČR GA202/05/0628; GA MŠk(CZ) LC06061; GA MŠk(CZ) LC06077 Institutional research plan: CEZ:AV0Z40550506 Keywords : nucleoside phostones * nucleoside derivatives * chlorodiethyl phosphite * ring enlargement Subject RIV: CC - Organic Chemistry Impact factor: 4.802, year: 2007

  17. Synthesis and anti-HIV activity of novel phenyl branched cyclopropyl nucleosides.

    Science.gov (United States)

    Wu, Ying; Hong, Joon Hee

    2005-09-01

    Novel phenyl branched cyclopropyl nucleoside analogues were designed and synthesized as potential antiviral agents. Cyclopropanation was performed via classical Simmons-Smith reaction using Zn(Et)2 and CH2I2. Coupling of the mesylates 11 and 12 with natural bases (A,C,T,U) and desilylation afforded a series of novel cyclopropyl nucleosides 21-28. The synthesized compounds were evaluated for their antiviral and antitumor activity against various viruses such as HIV, HSV-1, HSV-2 and HCMV.

  18. Lipases in green chemistry: acylation and alcoholysis on steroids and nucleosides.

    Science.gov (United States)

    Baldessari, Alicia; Iglesias, Luis E

    2012-01-01

    In this article, we describe the application of lipases in acylation and alcoholysis reactions on steroids and nucleosides. In the field of steroids, a variety of acetyl and fatty acid derivatives of androstanes, pregnanes, and cholestanes have been prepared through lipase-catalyzed acylation and alcoholysis reactions taking advantage of the high regio- and stereoselectivity of these enzymes. The substrates as well as the products show a high degree of biological activity as neurosteroids, hormones, and glucocorticoids. The regioselective preparation of diacylated nucleosides by means of an enzymatic alcoholysis allowed the synthesis of nucleosides prodrugs or modified nucleosides. The quantitative full deacylation and dealkoxycarbonylation of nucleosides and steroids is a mild synthetic method for the deprotection of these labile compounds. Some of the reported steroid and nucleoside products are novel, and it is not possible to obtain them satisfactorily by following traditional synthetic procedures. The advantages presented by this methodology, such as selectivity, mild reaction conditions, and low environmental impact, make the lipases an important tool in the application of the principles of Green Chemistry, offering a convenient way to prepare derivatives of natural compounds with a great potential in the pharmaceutical industry.

  19. Nature's combinatorial biosynthesis and recently engineered production of nucleoside antibiotics in Streptomyces.

    Science.gov (United States)

    Chen, Shawn; Kinney, William A; Van Lanen, Steven

    2017-04-01

    Modified nucleosides produced by Streptomyces and related actinomycetes are widely used in agriculture and medicine as antibacterial, antifungal, anticancer and antiviral agents. These specialized small-molecule metabolites are biosynthesized by complex enzymatic machineries encoded within gene clusters in the genome. The past decade has witnessed a burst of reports defining the key metabolic processes involved in the biosynthesis of several distinct families of nucleoside antibiotics. Furthermore, genome sequencing of various Streptomyces species has dramatically increased over recent years. Potential biosynthetic gene clusters for novel nucleoside antibiotics are now apparent by analysis of these genomes. Here we revisit strategies for production improvement of nucleoside antibiotics that have defined mechanisms of action, and are in clinical or agricultural use. We summarize the progress for genetically manipulating biosynthetic pathways for structural diversification of nucleoside antibiotics. Microorganism-based biosynthetic examples are provided and organized under genetic principles and metabolic engineering guidelines. We show perspectives on the future of combinatorial biosynthesis, and present a working model for discovery of novel nucleoside natural products in Streptomyces.

  20. Atlas-based functional radiosurgery: Early results

    Energy Technology Data Exchange (ETDEWEB)

    Stancanello, J.; Romanelli, P.; Pantelis, E.; Sebastiano, F.; Modugno, N. [Politecnico di Milano, Bioengineering Department and NEARlab, Milano, 20133 (Italy) and Siemens AG, Research and Clinical Collaborations, Erlangen, 91052 (Germany); Functional Neurosurgery Deptartment, Neuromed IRCCS, Pozzilli, 86077 (Italy); CyberKnife Center, Iatropolis, Athens, 15231 (Greece); Functional Neurosurgery Deptartment, Neuromed IRCCS, Pozzilli, 86077 (Italy)

    2009-02-15

    Functional disorders of the brain, such as dystonia and neuropathic pain, may respond poorly to medical therapy. Deep brain stimulation (DBS) of the globus pallidus pars interna (GPi) and the centromedian nucleus of the thalamus (CMN) may alleviate dystonia and neuropathic pain, respectively. A noninvasive alternative to DBS is radiosurgical ablation [internal pallidotomy (IP) and medial thalamotomy (MT)]. The main technical limitation of radiosurgery is that targets are selected only on the basis of MRI anatomy, without electrophysiological confirmation. This means that, to be feasible, image-based targeting must be highly accurate and reproducible. Here, we report on the feasibility of an atlas-based approach to targeting for functional radiosurgery. In this method, masks of the GPi, CMN, and medio-dorsal nucleus were nonrigidly registered to patients' T1-weighted MRI (T1w-MRI) and superimposed on patients' T2-weighted MRI (T2w-MRI). Radiosurgical targets were identified on the T2w-MRI registered to the planning CT by an expert functional neurosurgeon. To assess its feasibility, two patients were treated with the CyberKnife using this method of targeting; a patient with dystonia received an IP (120 Gy prescribed to the 65% isodose) and a patient with neuropathic pain received a MT (120 Gy to the 77% isodose). Six months after treatment, T2w-MRIs and contrast-enhanced T1w-MRIs showed edematous regions around the lesions; target placements were reevaluated by DW-MRIs. At 12 months post-treatment steroids for radiation-induced edema and medications for dystonia and neuropathic pain were suppressed. Both patients experienced significant relief from pain and dystonia-related problems. Fifteen months after treatment edema had disappeared. Thus, this work shows promising feasibility of atlas-based functional radiosurgery to improve patient condition. Further investigations are indicated for optimizing treatment dose.

  1. Atlas-based functional radiosurgery: Early results

    International Nuclear Information System (INIS)

    Stancanello, J.; Romanelli, P.; Pantelis, E.; Sebastiano, F.; Modugno, N.

    2009-01-01

    Functional disorders of the brain, such as dystonia and neuropathic pain, may respond poorly to medical therapy. Deep brain stimulation (DBS) of the globus pallidus pars interna (GPi) and the centromedian nucleus of the thalamus (CMN) may alleviate dystonia and neuropathic pain, respectively. A noninvasive alternative to DBS is radiosurgical ablation [internal pallidotomy (IP) and medial thalamotomy (MT)]. The main technical limitation of radiosurgery is that targets are selected only on the basis of MRI anatomy, without electrophysiological confirmation. This means that, to be feasible, image-based targeting must be highly accurate and reproducible. Here, we report on the feasibility of an atlas-based approach to targeting for functional radiosurgery. In this method, masks of the GPi, CMN, and medio-dorsal nucleus were nonrigidly registered to patients' T1-weighted MRI (T1w-MRI) and superimposed on patients' T2-weighted MRI (T2w-MRI). Radiosurgical targets were identified on the T2w-MRI registered to the planning CT by an expert functional neurosurgeon. To assess its feasibility, two patients were treated with the CyberKnife using this method of targeting; a patient with dystonia received an IP (120 Gy prescribed to the 65% isodose) and a patient with neuropathic pain received a MT (120 Gy to the 77% isodose). Six months after treatment, T2w-MRIs and contrast-enhanced T1w-MRIs showed edematous regions around the lesions; target placements were reevaluated by DW-MRIs. At 12 months post-treatment steroids for radiation-induced edema and medications for dystonia and neuropathic pain were suppressed. Both patients experienced significant relief from pain and dystonia-related problems. Fifteen months after treatment edema had disappeared. Thus, this work shows promising feasibility of atlas-based functional radiosurgery to improve patient condition. Further investigations are indicated for optimizing treatment dose.

  2. Characterization of reactive intermediates in laser photolysis of nucleoside using of sodium salt anthraquinone-2-sulfonic acid as photosensitizer

    International Nuclear Information System (INIS)

    Ma Jianhua; Lin Weizhen; Wang Wenfeng; Han Zhenhui; Yao Side; Lin Nianyun

    1999-01-01

    The interaction of triplet state of sodium salt of anthraquinone-2-sulfonic acid (AQS) with nucleosides has been investigated in CH 3 CN using KrF(248 nm) laser flash photolysis. The transient absorption spectra and kinetics obtained from the interaction of triplet AQS and nucleoside demonstrated that the primary ionic radical pair, radical cation of nucleosides and radical anion of AQS has been detected simultaneously for the first time

  3. Formation of Mixed-Ligand Complexes of Pd2+ with Nucleoside 5'-Monophosphates and Some Metal-Ion-Binding Nucleoside Surrogates

    Directory of Open Access Journals (Sweden)

    Oleg Golubev

    2014-10-01

    Full Text Available Formation of mixed-ligand Pd2+ complexes between canonical nucleoside 5'-monophosphates and five metal-ion-binding nucleoside analogs has been studied by 1H-NMR spectroscopy to test the ability of these nucleoside surrogates to discriminate between unmodified nucleobases by Pd2+-mediated base pairing. The nucleoside analogs studied included 2,6-bis(3,5-dimethylpyrazol-1-yl-, 2,6-bis(1-methylhydrazinyl- and 6-(3,5-dimethylpyrazol-1-yl-substituted 9-(β-d-ribofuranosylpurines 1–3, and 2,4-bis(3,5-dimethylpyrazol-1-yl- and 2,4-bis(1-methylhydrazinyl-substituted 5-(β-d-ribofuranosyl-pyrimidines 4–5. Among these, the purine derivatives 1-3 bound Pd2+ much more tightly than the pyrimidine derivatives 4, 5 despite apparently similar structures of the potential coordination sites. Compounds 1 and 2 formed markedly stable mixed-ligand Pd2+ complexes with UMP and GMP, UMP binding favored by 1 and GMP by 2. With 3, formation of mixed-ligand complexes was retarded by binding of two molecules of 3 to Pd2+.

  4. Rilpivirine: a new non-nucleoside reverse transcriptase inhibitor.

    Science.gov (United States)

    Sharma, Mamta; Saravolatz, Louis D

    2013-02-01

    Rilpivirine is a new non-nucleoside reverse transcriptase inhibitor (NNRTI) that is approved for HIV-1 treatment-naive adult patients in combination with other antiretroviral agents. The recommended dose is a 25 mg tablet once daily taken orally with a meal. Due to cytochrome P450 3A4 enzyme induction or gastric pH increase, rilpivirine cannot be coadministered with a number of other drugs (anticonvulsants, rifabutin, rifampicin, rifapentine, proton pump inhibitors, systemic dexamethasone and St John's wort). Rilpivirine should be used with caution when coadministered with a drug with a known risk for torsade de pointes. Rilpivirine has a better tolerability than a comparative NNRTI, efavirenz, in clinical trials, with fewer central nervous system adverse effects, rashes, lipid abnormalities and discontinuation rates. Virological failure occurs more commonly with higher baseline viral loads (>100,000 copies/mL) and lower baseline CD4 counts (<50 cells/mm(3)). Seventeen NNRTI mutations have been associated with decreased susceptibility to rilpivirine: K101E/P, E138A/G/K/Q/R, V179L, Y181C/I/V, H221Y, F227C, M230I/L, Y188L and the combination L100I + K103N. Resistance to rilpivirine largely excludes future use of the NNRTI class.

  5. Antibacterial Nucleoside-Analog Inhibitor of Bacterial RNA Polymerase.

    Science.gov (United States)

    Maffioli, Sonia I; Zhang, Yu; Degen, David; Carzaniga, Thomas; Del Gatto, Giancarlo; Serina, Stefania; Monciardini, Paolo; Mazzetti, Carlo; Guglierame, Paola; Candiani, Gianpaolo; Chiriac, Alina Iulia; Facchetti, Giuseppe; Kaltofen, Petra; Sahl, Hans-Georg; Dehò, Gianni; Donadio, Stefano; Ebright, Richard H

    2017-06-15

    Drug-resistant bacterial pathogens pose an urgent public-health crisis. Here, we report the discovery, from microbial-extract screening, of a nucleoside-analog inhibitor that inhibits bacterial RNA polymerase (RNAP) and exhibits antibacterial activity against drug-resistant bacterial pathogens: pseudouridimycin (PUM). PUM is a natural product comprising a formamidinylated, N-hydroxylated Gly-Gln dipeptide conjugated to 6'-amino-pseudouridine. PUM potently and selectively inhibits bacterial RNAP in vitro, inhibits bacterial growth in culture, and clears infection in a mouse model of Streptococcus pyogenes peritonitis. PUM inhibits RNAP through a binding site on RNAP (the NTP addition site) and mechanism (competition with UTP for occupancy of the NTP addition site) that differ from those of the RNAP inhibitor and current antibacterial drug rifampin (Rif). PUM exhibits additive antibacterial activity when co-administered with Rif, exhibits no cross-resistance with Rif, and exhibits a spontaneous resistance rate an order-of-magnitude lower than that of Rif. PUM is a highly promising lead for antibacterial therapy. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Visualizing multistep elevator-like transitions of a nucleoside transporter.

    Science.gov (United States)

    Hirschi, Marscha; Johnson, Zachary Lee; Lee, Seok-Yong

    2017-05-04

    Membrane transporters move substrates across the membrane by alternating access of their binding sites between the opposite sides of the membrane. An emerging model of this process is the elevator mechanism, in which a substrate-binding transport domain moves a large distance across the membrane. This mechanism has been characterized by a transition between two states, but the conformational path that leads to the transition is not yet known, largely because the available structural information has been limited to the two end states. Here we present crystal structures of the inward-facing, intermediate, and outward-facing states of a concentrative nucleoside transporter from Neisseria wadsworthii. Notably, we determined the structures of multiple intermediate conformations, in which the transport domain is captured halfway through its elevator motion. Our structures present a trajectory of the conformational transition in the elevator model, revealing multiple intermediate steps and state-dependent conformational changes within the transport domain that are associated with the elevator-like motion.

  7. Synthesis and characterization of a microporous 6FDA-polyimide made from a novel carbocyclic pseudo Tröger's base diamine: Effect of bicyclic bridge on gas transport properties

    KAUST Repository

    Abdulhamid, Mahmoud A.

    2017-10-12

    A newly designed carbocyclic pseudo Tröger\\'s base diamine (CTB) monomer, 2,8-dimethyl-3,9-diamino-5,6,11,12-tetrahydro-5,11-methanodibenzo[a,e][8]annulene (CTBDA) and its isomeric analogue 2,8-dimethyl-(1,7)(4,10)(3,9)-diamino-5,6,11,12-tetrahydro-5,11-methanodibenzo[a,e][8]annulene (iCTBDA), were designed for the synthesis of microporous 6FDA-based polyimides (6FDA-CTBDA and 6FDA-iCTBDA). Both polyimides were soluble, exhibited excellent thermal stability of ∼490 °C, and had high surface areas of 587 m2 g−1 (6FDA-CTBDA) and 562 m2 g−1 (6FDA-iCTBDA). A 6FDA-based polyimide derived from 4,10-dimethyl-3,9-diamino-6H,12H-5,11-methanodibenzo[b,f][1,5]-diazocine (6FDA-TBDA) was made for comparison to investigate the effects of the basic tertiary nitrogen functionality in the Tröger\\'s base diamine on the polymer properties relative to the carbocyclic 6FDA-CTBDA analogue. 6FDA-TBDA displayed lower gas permeabilities but moderately higher gas-pair permselectivities than 6FDA-CTBDA. The enhanced permselectivity of 6FDA-TBDA resulted exclusively from higher diffusion-based selectivity. Direct gas sorption measurements demonstrated that the basicity in the Tröger\\'s base bridge moiety enhanced the sorption capacity of CO2 only slightly and had no effect on the CO2/CH4 solubility selectivity in 6FDA-TBDA vs. 6FDA-CTBDA.

  8. Synthesis of nucleoside and nucleotide conjugates of bile acids, and polymerase construction of bile acid-functionalized DNA

    Czech Academy of Sciences Publication Activity Database

    Ikonen, Satu; Macíčková-Cahová, Hana; Pohl, Radek; Šanda, Miloslav; Hocek, Michal

    2010-01-01

    Roč. 8, č. 5 (2010), s. 1194-1201 ISSN 1477-0520 R&D Projects: GA MŠk LC512; GA ČR GA203/09/0317 Institutional research plan: CEZ:AV0Z40550506 Keywords : steroids * nucleosides * nucleoside triphosphates * DNA polymerase Subject RIV: CC - Organic Chemistry Impact factor: 3.451, year: 2010

  9. Structural and Enzymatic Characterization of a Nucleoside Diphosphate Sugar Hydrolase from Bdellovibrio bacteriovorus.

    Directory of Open Access Journals (Sweden)

    Andres H de la Peña

    Full Text Available Given the broad range of substrates hydrolyzed by Nudix (nucleoside diphosphate linked to X enzymes, identification of sequence and structural elements that correctly predict a Nudix substrate or characterize a family is key to correctly annotate the myriad of Nudix enzymes. Here, we present the structure determination and characterization of Bd3179 -- a Nudix hydrolase from Bdellovibrio bacteriovorus-that we show localized in the periplasmic space of this obligate Gram-negative predator. We demonstrate that the enzyme is a nucleoside diphosphate sugar hydrolase (NDPSase and has a high degree of sequence and structural similarity to a canonical ADP-ribose hydrolase and to a nucleoside diphosphate sugar hydrolase (1.4 and 1.3 Å Cα RMSD respectively. Examination of the structural elements conserved in both types of enzymes confirms that an aspartate-X-lysine motif on the C-terminal helix of the α-β-α NDPSase fold differentiates NDPSases from ADPRases.

  10. Highly regioselective synthesis of undecylenic acid esters of purine nucleosides catalyzed by Candida antarctica lipase B.

    Science.gov (United States)

    Gao, Wen-Li; Li, Ning; Zong, Min-Hua

    2011-11-01

    Regioselective undecylenoylation of purine nucleosides as potential dual prodrugs was achieved by Candida antarctica lipase B using adenosine as a model reactant. The optimum organic solvent, molar ratio of vinyl ester to nucleoside, enzyme dosage, reaction temperature and molecular sieve amount were anhydrous THF, 5:1, 20 U/ml, 45°C and 75 mg/ml, respectively. Under the optimum conditions, the initial reaction rate, yield and 5'-regioselectivity were 1.1 mM/h, 90% and >99%, respectively. The enzymatic acylation of various nucleosides furnished the desired 5'-ester derivatives with the yields of 60-95% and 5'-regioselectivities of >99%. In addition, the lipase displayed excellent operational stability in THF, and retained 96% of its initial activity after reused for five batches.

  11. Structure of purine nucleoside phosphorylase (DeoD) from Bacillus anthracis

    International Nuclear Information System (INIS)

    Grenha, Rosa; Levdikov, Vladimir M.; Fogg, Mark J.; Blagova, Elena V.; Brannigan, James A.; Wilkinson, Anthony J.; Wilson, Keith S.

    2005-01-01

    The crystal structure of purine nucleoside phosphorylase (DeoD) from B. anthracis was solved by X-ray crystallography using molecular replacement and refined at a resolution of 2.24 Å. Protein structures from the causative agent of anthrax (Bacillus anthracis) are being determined as part of a structural genomics programme. Amongst initial candidates for crystallographic analysis are enzymes involved in nucleotide biosynthesis, since these are recognized as potential targets in antibacterial therapy. Purine nucleoside phosphorylase is a key enzyme in the purine-salvage pathway. The crystal structure of purine nucleoside phosphorylase (DeoD) from B. anthracis has been solved by molecular replacement at 2.24 Å resolution and refined to an R factor of 18.4%. This is the first report of a DeoD structure from a Gram-positive bacterium

  12. Training Residential Staff to Conduct Trial-Based Functional Analyses

    Science.gov (United States)

    Lambert, Joseph M.; Bloom, Sarah E.; Kunnavatana, S. Shanun; Collins, Shawnee D.; Clay, Casey J.

    2013-01-01

    We taught 6 supervisors of a residential service provider for adults with developmental disabilities to train 9 house managers to conduct trial-based functional analyses. Effects of the training were evaluated with a nonconcurrent multiple baseline. Results suggest that house managers can be trained to conduct trial-based functional analyses with…

  13. In situ enzymatic removal of orthophosphate by the nucleoside phosphorylase catalyzed phosphorolysis of nicotinamide riboside.

    Science.gov (United States)

    Shriver, J W; Sykes, B D

    1982-09-01

    An enzymatic orthophosphate removal system is described which can be effectively used to continuously remove orthophosphate from biochemical samples. The phosphorolysis of nicotinamide riboside is catalyzed by calf spleen nucleoside phosphorylase to give ribose-1-PO4 and nicotinamide along with a proton. At pH 8 the production of ribose-1-PO4 from orthophosphate is essentially quantitative. This reaction can be monitored optically or by 31P nuclear magnetic resonance (NMR). Equations are given for determining the time required to remove a given amount of phosphate from a typical NMR sample with a known amount of nucleoside phosphorylase. The effects of a competing orthophosphate-producing reaction are considered.

  14. Versatile synthesis of amino acid functionalized nucleosides via a domino carboxamidation reaction

    Directory of Open Access Journals (Sweden)

    Vicky Gheerardijn

    2014-11-01

    Full Text Available Functionalized oligonucleotides have recently gained increased attention for incorporation in modified nucleic acid structures both for the design of aptamers with enhanced binding properties as well as the construction of catalytic DNA and RNA. As a shortcut alternative to the incorporation of multiple modified residues, each bearing one extra functional group, we present here a straightforward method for direct linking of functionalized amino acids to the nucleoside base, thus equipping the nucleoside with two extra functionalities at once. As a proof of principle, we have introduced three amino acids with functional groups frequently used as key-intermediates in DNA- and RNAzymes via an efficient and straightforward domino carboxamidation reaction.

  15. Use of nucleoside (tide) analogues in patients with hepatitis B-related acute liver failure

    DEFF Research Database (Denmark)

    Dao, Doan Y; Seremba, Emmanuel; Ajmera, Veeral

    2012-01-01

    The efficacy of nucleoside(tide) analogues (NA) in the treatment of acute liver failure due to hepatitis B virus (HBV-ALF) remains controversial. We determined retrospectively the impact of NAs in a large cohort of patients with HBV-ALF.......The efficacy of nucleoside(tide) analogues (NA) in the treatment of acute liver failure due to hepatitis B virus (HBV-ALF) remains controversial. We determined retrospectively the impact of NAs in a large cohort of patients with HBV-ALF....

  16. Mutagenicity of irradiated solutions of nuclei acid bases and nucleosides in Salmonella typhimurium

    International Nuclear Information System (INIS)

    Wilmer, J.; Schubert, J.

    1981-01-01

    Solutions of nucleic acid bases, nucleosides and a nucleotide, saturated with either N 2 , N 2 O or O 2 , were irradiated and tested for mutagenicity towards Salmonella typhimurium, with and without pre-incubation. Irradiated solutions of the nuclei acid bases were all non-mutagenic. Irradiated solutions of the nucleosides showed mutagenicity in S. typhimurium TA100 (pre-incubation assay). Generally, the mutagenicity followed the order: N 2 O > N 2 > O 2 . The results show that the formation of mutagenic radiolytic products is initiated by attack of mainly solutions of the nucleotide thymidine-5'-monophosphate, no mutagenicity could be detected. (orig.)

  17. Anopheles gambiae Purine Nucleoside Phosphorylase: Catalysis, Structure, and Inhibition

    Energy Technology Data Exchange (ETDEWEB)

    Taylor,E.; Rinaldo-Matthis, A.; Li, L.; Ghanem, M.; Hazleton, K.; Cassera, M.; Almo, S.; Schramm, V.

    2007-01-01

    The purine salvage pathway of Anopheles gambiae, a mosquito that transmits malaria, has been identified in genome searches on the basis of sequence homology with characterized enzymes. Purine nucleoside phosphorylase (PNP) is a target for the development of therapeutic agents in humans and purine auxotrophs, including malarial parasites. The PNP from Anopheles gambiae (AgPNP) was expressed in Escherichia coli and compared to the PNPs from Homo sapiens (HsPNP) and Plasmodium falciparum (PfPNP). AgPNP has kcat values of 54 and 41 s-1 for 2'-deoxyinosine and inosine, its preferred substrates, and 1.0 s-1 for guanosine. However, the chemical step is fast for AgPNP at 226 s-1 for guanosine in pre-steady-state studies. 5'-Deaza-1'-aza-2'-deoxy-1'-(9-methylene)-Immucillin-H (DADMe-ImmH) is a transition-state mimic for a 2'-deoxyinosine ribocation with a fully dissociated N-ribosidic bond and is a slow-onset, tight-binding inhibitor with a dissociation constant of 3.5 pM. This is the tightest-binding inhibitor known for any PNP, with a remarkable Km/Ki* of 5.4 x 107, and is consistent with enzymatic transition state predictions of enhanced transition-state analogue binding in enzymes with enhanced catalytic efficiency. Deoxyguanosine is a weaker substrate than deoxyinosine, and DADMe-Immucillin-G is less tightly bound than DADMe-ImmH, with a dissociation constant of 23 pM for AgPNP as compared to 7 pM for HsPNP. The crystal structure of AgPNP was determined in complex with DADMe-ImmH and phosphate to a resolution of 2.2 Angstroms to reveal the differences in substrate and inhibitor specificity. The distance from the N1' cation to the phosphate O4 anion is shorter in the AgPNP{center_dot}DADMe-ImmH{center_dot}PO4 complex than in HsPNP{center_dot}DADMe-ImmH{center_dot}SO4, offering one explanation for the stronger inhibitory effect of DADMe-ImmH for AgPNP.

  18. Selective Phosphorylation of South and North-Cytidine and Adenosine Methanocarba-Nucleosides by Human Nucleoside and Nucleotide Kinases Correlates with Their Growth Inhibitory Effects on Cultured Cells.

    Science.gov (United States)

    Sjuvarsson, Elena; Marquez, Victor E; Eriksson, Staffan

    2015-01-01

    Here bicyclo[3.1.0]hexane locked deoxycytidine (S-MCdC, N-MCdC), and deoxyadenosine analogs (S-MCdA and N-MCdA) were examined as substrates for purified preparations of human deoxynucleoside kinases: dCK, dGK, TK2, TK1, the ribonucleoside kinase UCK2, two NMP kinases (CMPK1, TMPK) and a NDP kinase. dCK can be important for the first step of phosphorylation of S-MCdC in cells, but S-MCdCMP was not a substrate for CMPK1, TMPK, or NDPK. dCK and dGK had a preference for the S-MCdA whereas N-MCdA was not a substrate for dCK, TK1, UCK2, TK2, dGK nucleoside kinases. The cell growth experiments suggested that N-MCdC and S-MCdA could be activated in cells by cellular kinases so that a triphosphate metabolite was formed. List of abbreviations: ddC, 2', 3'-didioxycytosine, Zalcitabine; 3TC, β-L-(-)-2',3'-dideoxy-3'-thiacytidine, Lamivudine; CdA, 2-cloro-2'-deoxyadenosine, Cladribine; AraA, 9-β-D-arabinofuranosyladenine; hCNT 1-3, human Concentrative Nucleoside Transporter type 1, 2 and 3; hENT 1-4, human Equilibrative Nucleoside Transporter type 1, 2, 3, and 4.

  19. Synthesis of 2'-deoxyuridine and 2'-deoxycytidine nucleosides bearing bipyridine and terpyridine ligands in position 5

    Czech Academy of Sciences Publication Activity Database

    Kalachová, Lubica; Pohl, Radek; Hocek, Michal

    -, č. 1 (2009), s. 105-112 ISSN 0039-7881 R&D Projects: GA MŠk LC512 Institutional research plan: CEZ:AV0Z40550506 Keywords : nucleosides * pyrimidines * cross-coupling * bipyridine s Subject RIV: CC - Organic Chemistry Impact factor: 2.572, year: 2009

  20. Inhibition of the Escherichia coli 6-oxopurine phosphoribosyltransferases by nucleoside phosphonates: potential for new antibacterial agents

    Czech Academy of Sciences Publication Activity Database

    Keough, D. T.; Hocková, Dana; Rejman, Dominik; Špaček, Petr; Vrbková, Silvie; Krečmerová, Marcela; Eng, W. S.; Jans, H.; West, N. P.; Naesens, L. M. J.; de Jersey, J.; Guddat, L. W.

    2013-01-01

    Roč. 56, č. 17 (2013), s. 6967-6984 ISSN 0022-2623 R&D Projects: GA ČR GAP207/11/0108 Institutional support: RVO:61388963 Keywords : nucleoside phosphonates * antibacterial agents * hypoxanthine-guanine phosphoribosyltransferase * state analog inhibitor * antimalarial chemotherapy Subject RIV: CC - Organic Chemistry Impact factor: 5.480, year: 2013

  1. Expression and purification of human and Saccharomyces cerevisiae equilibrative nucleoside transporters.

    Science.gov (United States)

    Boswell-Casteel, Rebba C; Johnson, Jennifer M; Roe-Žurž, Zygy; Duggan, Kelli D; Schmitz, Hannah; Hays, Franklin A

    2018-02-01

    Nucleosides play an essential role in the physiology of eukaryotes by acting as metabolic precursors in de novo nucleic acid synthesis and energy metabolism. Nucleosides also act as ligands for purinergic receptors. Equilibrative nucleoside transporters (ENTs) are polytopic integral membrane proteins that aid in regulating plasmalemmal flux of purine and pyrimidine nucleosides and nucleobases. ENTs exhibit broad substrate selectivity across different isoforms and utilize diverse mechanisms to drive substrate flux across membranes. However, the molecular mechanisms and chemical determinants of ENT-mediated substrate recognition, binding, inhibition, and transport are poorly understood. To determine how ENT-mediated transport occurs at the molecular level, greater chemical insight and assays employing purified protein are essential. This article focuses on the expression and purification of human ENT1, human ENT2, and Saccharomyces cerevisiae ScENT1 using novel expression and purification strategies to isolate recombinant ENTs. ScENT1, hENT1, and hENT2 were expressed in W303 Saccharomyces cerevisiae cells and detergent solubilized from the membrane. After detergent extraction, these ENTs were further purified using immobilized metal affinity chromatography and size exclusion chromatography. This effort resulted in obtaining quantities of purified protein sufficient for future biophysical analysis. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Design and Synthesis of Fluorescent Acyclic Nucleoside Phosphonates as Potent Inhibitors of Bacterial Adenylate Cyclases

    Czech Academy of Sciences Publication Activity Database

    Břehová, Petra; Šmídková, Markéta; Skácel, Jan; Dračínský, Martin; Mertlíková-Kaiserová, Helena; Velasquez, M. P. S.; Watts, V. J.; Janeba, Zlatko

    2016-01-01

    Roč. 11, č. 22 (2016), s. 2534-2546 ISSN 1860-7179 R&D Projects: GA MV VG20102015046; GA MŠk LO1302 Institutional support: RVO:61388963 Keywords : adenylate cyclase toxin * acyclic nucleoside phosphonates * anthranilic acid Subject RIV: CC - Organic Chemistry Impact factor: 3.225, year: 2016

  3. Secretion of antiretroviral chemokines by human cells cultured with acyclic nucleoside phosphonates

    Czech Academy of Sciences Publication Activity Database

    Zídek, Zdeněk; Kmoníčková, Eva; Holý, Antonín

    2007-01-01

    Roč. 574, - (2007), s. 77-84 ISSN 0014-2999 R&D Projects: GA MŠk 1M0508 Institutional research plan: CEZ:AV0Z50390512; CEZ:AV0Z40550506 Keywords : Acyclic nucleoside phosphonate * Chemokine * Cytokine Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.376, year: 2007

  4. Cu(I)-catalyzed efficient synthesis of 2′-Triazolo-nucleoside conjugates

    DEFF Research Database (Denmark)

    Mathur, D.; Rana, N.; Olsen, Carl Erik

    2015-01-01

    -nucleoside conjugates, which can be evaluated for different biological activity for suitable drug development, were unambiguously identified on the basis of 1H NMR, 13C NMR, IR, and HRMS data analysis. These compounds have been synthesized for the first time and have not been reported in the literature earlier....

  5. The Synthesis and Conformation of Dihydroxypiperidinyl Derivates of Nucleobases as Novel Iminosugar Nucleoside Analogs

    Czech Academy of Sciences Publication Activity Database

    Rejman, Dominik; Pohl, Radek; Dračínský, Martin

    -, č. 11 (2011), s. 2172-2187 ISSN 1434-193X R&D Projects: GA MŠk(CZ) LC06077; GA MŠk 2B06065; GA AV ČR KJB400550903 Institutional research plan: CEZ:AV0Z40550506 Keywords : iminosugars * azanucleosides * nucleosides Subject RIV: CC - Organic Chemistry Impact factor: 3.329, year: 2011

  6. Acyclic nucleoside phosphonate antivirals activate gene expression of monocyte chemotactic protein 1 and 3.

    Czech Academy of Sciences Publication Activity Database

    Potměšil, Petr; Holý, Antonín; Kmoníčková, Eva; Křížková, Jana; Zídek, Zdeněk

    2007-01-01

    Roč. 14, č. 1 (2007), s. 59-66 ISSN 1021-7770 R&D Projects: GA MŠk 1M0508 Institutional research plan: CEZ:AV0Z50390512; CEZ:AV0Z40550506 Keywords : Acyclic nucleoside phosponate * HIV * Monocyte chemotactic protein Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.024, year: 2007

  7. Novel and Efficient Synthesis of gem-Difluorinated Derivatives of Acyclic Nucleoside Phosphonates (ANPs)

    Czech Academy of Sciences Publication Activity Database

    Pomeisl, Karel; Beier, Petr; Pohl, Radek; Krečmerová, Marcela

    2016-01-01

    Roč. 1, č. 10 (2016), s. 2102-2106 ISSN 2365-6549 R&D Projects: GA ČR(CZ) GA14-00522S Institutional support: RVO:61388963 Keywords : difluoromethylation * difluoromethylphosphonate * acyclic nucleoside phosphonate * phosphonate ester * microwave reaction Subject RIV: CC - Organic Chemistry

  8. beta-1,2,3-Triazolyl-Nucleosides as Nicotinamide Riboside Mimics

    Czech Academy of Sciences Publication Activity Database

    Amigues, E.J.; Armstrong, E.; Dvořáková, Marcela; Migaud, M.E.; Huang, M.

    2009-01-01

    Roč. 28, č. 3 (2009), s. 238-259 ISSN 1525-7770 Institutional research plan: CEZ:AV0Z50380511 Keywords : Nucleoside * nucleotide * sirtuin Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.768, year: 2009

  9. A novel type of acyclic nucleoside phosphonates derived from 2-(phosphonomethoxy)propanoic acid

    Czech Academy of Sciences Publication Activity Database

    Kaiser, Martin Maxmilian; Jansa, Petr; Dračínský, Martin; Janeba, Zlatko

    2012-01-01

    Roč. 68, č. 21 (2012), s. 4003-4012 ISSN 0040-4020 R&D Projects: GA MV VG20102015046 Institutional support: RVO:61388963 Keywords : acyclic nucleoside phosphonates * CPMEA * HPMPA * PME A * oxidation * TEMPO * microwave * antiviral Subject RIV: CC - Organic Chemistry Impact factor: 2.803, year: 2012

  10. N-Branched acyclic nucleoside phosphonates as monomers for the synthesis of modified oligonucleotides

    Czech Academy of Sciences Publication Activity Database

    Hocková, Dana; Rosenbergová, Šárka; Ménová, Petra; Páv, Ondřej; Pohl, Radek; Novák, Pavel; Rosenberg, Ivan

    2015-01-01

    Roč. 13, č. 15 (2015), s. 4449-4458 ISSN 1477-0520 R&D Projects: GA ČR GAP207/11/0108; GA ČR GA13-26526S Institutional support: RVO:61388963 Keywords : acyclic nucleoside phosphonates * oligonucleotides * solid phase synthesis Subject RIV: CC - Organic Chemistry Impact factor: 3.559, year: 2015

  11. Inhibition of hypoxanthine-guanine phosphoribosyltransferase by acyclic nucleoside phosphonates: A new class of antimalarial therapeutics

    Czech Academy of Sciences Publication Activity Database

    Keough, D. T.; Hocková, Dana; Holý, Antonín; Naesens, L.; Skinner-Adams, T. S.; de Jersey, J.; Guddat, L. W.

    2009-01-01

    Roč. 52, č. 14 (2009), s. 4391-4399 ISSN 0022-2623 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * phosphoribosyltransferase * enzyme inhibitors * Plasmodium falciparum Subject RIV: CC - Organic Chemistry Impact factor: 4.802, year: 2009

  12. Synthesis and properties of novel types of chiral open-ring acyclic nucleoside phosphonates

    Czech Academy of Sciences Publication Activity Database

    Holý, Antonín; Doláková, Petra

    2005-01-01

    Roč. 65, č. 3 (2005), A31 ISSN 0166-3542. [International Conference on Antiviral Research /8./. Barcelona, 11.04.2005-14.04.2005] Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonate * pyrimidine * antiviral activity Subject RIV: CC - Organic Chemistry Impact factor: 3.406, year: 2005

  13. Detergent inhibited, heat labile nucleoside triphosphatase in cores of avian myeloblastosis virus

    DEFF Research Database (Denmark)

    Jensen, Kaj Frank

    1978-01-01

    Endogenous DNA synthesis was studied in isolated core particles of avian myeloblastosis virus. It was found that cores contained an enzymatic activity which rapidly converted the added nucleoside triphosphates to diphosphates (but not further) at 0 degrees C, thus inhibiting DNA synthesis. This t...

  14. Activities of Different Classes of Acyclic Nucleoside Phosphonates against BK Virus in Primary Human Renal Cells

    Czech Academy of Sciences Publication Activity Database

    Topalis, D.; Lebeau, I.; Krečmerová, Marcela; Andrei, G.; Snoeck, R.

    2011-01-01

    Roč. 55, č. 5 (2011), s. 1961-1967 ISSN 0066-4804 Institutional research plan: CEZ:AV0Z40550506 Keywords : polyomavirus * BK virus * nephropathy * acyclic nucleoside phosphonates * HPMP-5-azaC Subject RIV: CC - Organic Chemistry Impact factor: 4.841, year: 2011

  15. Synthesis of Novel Uracil Non-Nucleoside Derivatives as Potential Reverse Transcriptase Inhibitors of HIV-1

    DEFF Research Database (Denmark)

    El-Brollosy, Nasser R.; Al-Deeb, Omar. A.; El-Emam, Ali A.

    2009-01-01

    Novel emivirine and TNK-651 analogues 5a-d were synthesized by reaction of chloromethyl ethyl ether and / or benzyl chloromethyl ether, respectively, with uracils having 5-ethyl and 6-(4-methylbenzyl) or 6-(3,4-dimethoxybenzyl) substituents. A series of new uracil non-nucleosides substituted at N-1...

  16. Organometallic nucleoside analogues with ferrocenyl linker groups: synthesis and cancer cell line studies.

    Science.gov (United States)

    Nguyen, Huy V; Sallustrau, Antoine; Balzarini, Jan; Bedford, Matthew R; Eden, John C; Georgousi, Niki; Hodges, Nikolas J; Kedge, Jonathan; Mehellou, Youcef; Tselepis, Chris; Tucker, James H R

    2014-07-10

    Examples of organometallic compounds as nucleoside analogues are rare within the field of medicinal bioorganometallic chemistry. We report on the synthesis and properties of two chiral ferrocene derivatives containing a nucleobase and a hydroxyalkyl group. These so-called ferronucleosides show promising anticancer activity, with cytostatic studies on five different cancer cell lines indicating that both functional groups are required for optimal activity.

  17. Cross-coupling reactions of unprotected halopurine bases, nucleosides, nucleotides and nucleoside triphosphates with 4-boronophenylalanine in water. Synthesis of (purin-8-yl)- and (purin-6-yl)phenylalanines.

    Science.gov (United States)

    Capek, Petr; Pohl, Radek; Hocek, Michal

    2006-06-07

    An expeditious and highly efficient single-step methodology for the introduction of a phenylalanine moiety into position 8 and 6 of the purine scaffold was developed based on aqueous-phase Pd-catalysed Suzuki-Miyaura cross-coupling reactions of unprotected 4-boronophenylalanine with 8-bromo- or 6-chloropurines. The scope of the methodology was demonstrated by syntheses of unprotected (adenin-8-yl)phenylalanine base, nucleosides, nucleotides and nucleoside triphosphates as well as (purin-6-yl)phenylalanine base and nucleosides. All these products were obtained in high yields and in optically pure form.

  18. Structural and thermodynamic analysis of modified nucleosides in self-assembled DNA cross-tiles.

    Science.gov (United States)

    Hakker, Lauren; Marchi, Alexandria N; Harris, Kimberly A; LaBean, Thomas H; Agris, Paul F

    2014-01-01

    DNA Holliday junctions are important natural strand-exchange structures that form during homologous recombination. Immobile four-arm junctions, analogs to Holliday junctions, have been designed to self-assemble into cross-tile structures by maximizing Watson-Crick base pairing and fixed crossover points. The cross-tiles, self-assembled from base pair recognition between designed single-stranded DNAs, form higher order lattice structures through cohesion of self-associating sticky ends. These cross-tiles have 16 unpaired nucleosides in the central loop at the junction of the four duplex stems. The importance of the centralized unpaired nucleosides to the structure's thermodynamic stability and self-assembly is unknown. Cross-tile DNA nanostructures were designed and constructed from nine single-stranded DNAs with four shell strands, four arms, and a central loop containing 16 unpaired bases. The 16 unpaired bases were either 2'-deoxyribothymidines, 2'-O-methylribouridines, or abasic 1',2'-dideoxyribonucleosides. Thermodynamic profiles and structural base-stacking contributions were assessed using UV absorption spectroscopy during thermal denaturation and circular dichroism spectroscopy, respectively, and the resulting structures were observed by atomic force microscopy. There were surprisingly significant changes in the thermodynamic and structural properties of lattice formation as a result of altering only the 16 unpaired, centralized nucleosides. The 16 unpaired 2'-O-methyluridines were stabilizing and produced uniform tubular structures. In contrast, the abasic nucleosides were destabilizing producing a mixture of structures. These results strongly indicate the importance of a small number of centrally located unpaired nucleosides within the structures. Since minor modifications lead to palpable changes in lattice formation, DNA cross-tiles present an easily manipulated structure convenient for applications in biomedical and biosensing devices.

  19. Kinetic α-deuterium isotope effect as a probe of transition state structure and reaction mechanism in nucleoside hydrolysis

    International Nuclear Information System (INIS)

    Stein, R.L.

    1978-01-01

    Theoretical equilibrium α-deuterium isotope effects were calculated for systems modeling nucleoside and glycoside hydrolyses using a computer program (Burton, G.W., Sims, L.B., Wilson, J.C., and Fry, A.J., J. Amer. Chem. Soc., 99, 3374(1977)) which computes isotope effects directly from the expression of Biegeleisen and Mayer (Biegeleisen, J. and Mayer, M.G., J. Chem. Phys., 17, 675(1949)). For nucleoside hydrolysis proceeding through an oxocarbonium ion intermediate, KH/KD = 1.21 to 1.25; while for nucleoside hydrolysis proceeding through an oxocarbonium ion intermediate KH/KD = 1.15 to 1.19. The models used in the calculations were generated systematically and involved a minimum of subjectivity in the selection of molecular parameters. The isotope effects calculated formed the basis for the interpretation of experimental kinetic α-deuterium isotope effects for nucleoside and glycoside hydrolysis

  20. Bifunctional acyclic nucleoside phosphonates: 2. Symmetrical 2-{[bis(phosphono)methoxy]methyl}ethyl derivatives of purines and pyrimidines

    Czech Academy of Sciences Publication Activity Database

    Vrbková, Silvie; Dračínský, Martin; Holý, Antonín

    2007-01-01

    Roč. 72, č. 7 (2007), s. 965-983 ISSN 0010-0765 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Grant - others:Gilead Sciences(US) HPAW-2002-10096 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * ANPs * acyclic nucleoside bisphosphonates * ANbPs * phosphonomethyl ethers Subject RIV: CC - Organic Chemistry Impact factor: 0.879, year: 2007

  1. Novel regulation of equlibrative nucleoside transporter 1 (ENT1) by receptor-stimulated Ca2+-dependent calmodulin binding

    Science.gov (United States)

    Bicket, Alex; Mehrabi, Pedram; Naydenova, Zlatina; Wong, Victoria; Donaldson, Logan; Stagljar, Igor

    2016-01-01

    Equilibrative nucleoside transporters (ENTs) facilitate the flux of nucleosides, such as adenosine, and nucleoside analog (NA) drugs across cell membranes. A correlation between adenosine flux and calcium-dependent signaling has been previously reported; however, the mechanistic basis of these observations is not known. Here we report the identification of the calcium signaling transducer calmodulin (CaM) as an ENT1-interacting protein, via a conserved classic 1-5-10 motif in ENT1. Calcium-dependent human ENT1-CaM protein interactions were confirmed in human cell lines (HEK293, RT4, U-87 MG) using biochemical assays (HEK293) and the functional assays (HEK293, RT4), which confirmed modified nucleoside uptake that occurred in the presence of pharmacological manipulations of calcium levels and CaM function. Nucleoside and NA drug uptake was significantly decreased (∼12% and ∼39%, respectively) by chelating calcium (EGTA, 50 μM; BAPTA-AM, 25 μM), whereas increasing intracellular calcium (thapsigargin, 1.5 μM) led to increased nucleoside uptake (∼26%). Activation of N-methyl-d-aspartate (NMDA) receptors (in U-87 MG) by glutamate (1 mM) and glycine (100 μM) significantly increased nucleoside uptake (∼38%) except in the presence of the NMDA receptor antagonist, MK-801 (50 μM), or CaM antagonist, W7 (50 μM). These data support the existence of a previously unidentified novel receptor-dependent regulatory mechanism, whereby intracellular calcium modulates nucleoside and NA drug uptake via CaM-dependent interaction of ENT1. These findings suggest that ENT1 is regulated via receptor-dependent calcium-linked pathways resulting in an alteration of purine flux, which may modulate purinergic signaling and influence NA drug efficacy. PMID:27009875

  2. N-Phosphonocarbonylpyrrolidine Derivatives of Guanine: A New Class of Bi-Substrate Inhibitors of Human Purine Nucleoside Phosphorylase

    Czech Academy of Sciences Publication Activity Database

    Rejman, Dominik; Panova, Natalya; Klener, P.; Maswabi, B.; Pohl, Radek; Rosenberg, Ivan

    2012-01-01

    Roč. 55, č. 4 (2012), s. 1612-1621 ISSN 0022-2623 R&D Projects: GA MŠk 2B06065; GA MŠk(CZ) LC06077; GA AV ČR KAN200520801 Institutional research plan: CEZ:AV0Z40550506 Keywords : nucleoside * phosphonic acid * pyrrolidine * purine nucleoside Subject RIV: CC - Organic Chemistry Impact factor: 5.614, year: 2012

  3. Reactivity Studies of 2,6-Bis-(1,2,3-triazolyl)purine Nucleosides with Hydrazines and Amino Acids

    OpenAIRE

    Laķis, E; Novosjolova, I; Bizdēna, Ē; Turks, M

    2014-01-01

    Reactions of 2,6-bis-(1,2,3-triazolyl)purine nucleosides with nucleophiles is efficient route to C(6)-derivatization of purine base. To extend the field of application of the method, we studied reactivity of bis-(1,2,3-triazolyl)purine derivatives toward hydrazines and amino acids and obtained nucleosides substituted at C(6) with hydrazine and amino acid moieties.

  4. Can Crystal Symmetry and Packing Influence the Active Site Conformation of Homohexameric Purine Nucleoside Phosphorylases?

    Directory of Open Access Journals (Sweden)

    Marija Luić

    2016-06-01

    Full Text Available It is generaly believed that enzymes retain most of their functionality in the crystal form due to the large solvent content of protein crystals. This is facilitated by the fact that their natural environment in solution is not too far from the one found in the crystal form. Nevertheless, if the nature of the enzyme is such to require conformational changes, overcoming of the crystal packing constraints may prove to be too difficult. Such conformational change is present in one class of enzymes (purine nucleoside phosphorylases, that is the subject of our scientific interest for many years. The influence of crystal symmetry and crystal packing on the conformation of the active sites in the case of homohexameric purine nucleoside phosphorylases is presented and analysed. This work is licensed under a Creative Commons Attribution 4.0 International License.

  5. Synthesis and anti-cancer activity of some novel 5-azacytosine nucleosides.

    Science.gov (United States)

    Tiwari, Kamal N; Cappellacci, Loredana; Montgomery, John A; Secrist, John A

    2003-12-01

    1-O-Acetyl-2-deoxy-3,5-di-O-toluoyl-4-thio-D-erythro-pentofuranose and 2-deoxy-1,3,5-tri-O-acetyl-4-thio-L-threo-pentofuranose were coupled with 5-azacytosine to obtain alpha and beta anomers of nucleosides. All four nucleosides were reduced to the corresponding dihydro derivatives and deblocked to give target compounds. All eight target compounds were evaluated in a series of human cancer cell lines in culture. Only 2'-deoxy-4'-thio-5-azacytidine (3beta) was found to be cytotoxic in all the cell lines and was further evaluated in vivo. Details of the synthesis and biological activity are reported.

  6. Three-dimensional structure of E. Coli purine nucleoside phosphorylase at 0.99 Å resolution

    Energy Technology Data Exchange (ETDEWEB)

    Timofeev, V. I., E-mail: tostars@mail.ru [Russian Academy of Sciences, Shubnikov Institute of Crystallography (Russian Federation); Abramchik, Yu. A., E-mail: ugama@yandex.ru [Russian Academy of Sciences, Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry (Russian Federation); Zhukhlistova, N. E., E-mail: inna@ns.crys.ras.ru [Russian Academy of Sciences, Shubnikov Institute of Crystallography (Russian Federation); Muravieva, T. I.; Esipov, R. S. [Russian Academy of Sciences, Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry (Russian Federation); Kuranova, I. P. [Russian Academy of Sciences, Shubnikov Institute of Crystallography (Russian Federation)

    2016-03-15

    Purine nucleoside phosphorylases (PNPs) catalyze the reversible phosphorolysis of nucleosides and are key enzymes involved in nucleotide metabolism. They are essential for normal cell function and can catalyze the transglycosylation. Crystals of E. coli PNP were grown in microgravity by the capillary counterdiffusion method through a gel layer. The three-dimensional structure of the enzyme was determined by the molecular-replacement method at 0.99 Å resolution. The structural features are considered, and the structure of E. coli PNP is compared with the structures of the free enzyme and its complexes with purine base derivatives established earlier. A comparison of the environment of the purine base in the complex of PNP with formycin A and of the pyrimidine base in the complex of uridine phosphorylase with thymidine revealed the main structural features of the base-binding sites. Coordinates of the atomic model determined with high accuracy were deposited in the Protein Data Bank (PDB-ID: 4RJ2).

  7. Synthesis and antiviral properties of spirocyclic [1,2,3]-triazolooxazine nucleosides.

    Science.gov (United States)

    Dell'Isola, Antonio; McLachlan, Matthew M W; Neuman, Benjamin W; Al-Mullah, Hawaa M N; Binks, Alexander W D; Elvidge, Warren; Shankland, Kenneth; Cobb, Alexander J A

    2014-09-08

    An efficient synthesis of spirocyclic triazolooxazine nucleosides is described. This was achieved by the conversion of β-D-psicofuranose to the corresponding azido-derivative, followed by alkylation of the primary alcohol with a range of propargyl bromides, obtained by Sonogashira chemistry. The products of these reactions underwent 1,3-dipolar addition smoothly to generate the protected spirocyclic adducts. These were easily deprotected to give the corresponding ribose nucleosides. The library of compounds obtained was investigated for its antiviral activity using MHV (mouse hepatitis virus) as a model wherein derivative 3 f showed the most promising activity and tolerability. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Nucleoside Phosphate-Conjugates Come of Age: Catalytic Transformation, Polymerase Recognition and Antiviral Properties.

    Science.gov (United States)

    Groaz, Elisabetta; Herdewijn, Piet

    2015-01-01

    Over the past few decades, different types of nucleoside phosphate-conjugates have been under extensive investigation due to their favorable molecular lability with interesting catalytic hydrolysis mechanisms, recognition as polymerase substrates, and especially for their development as antiviral/ anticancer protide therapeutics. The antiviral conjugates such as nucleoside phosphoesters and phosphoramidates that were discovered and developed in the initial years have been well reviewed by the pioneers in the field. In the present review, we will discuss the basic chemical and biological principles behind consideration of some representative structural classes. We will also summarize the chemical and biological properties of some of the more recent analogues that were synthesized and evaluated in our laboratory and by others. This includes new principles for their application as direct substrates of polymerases, nucleobasedependent catalytic and antiviral activity, and a plausible 'prodrug of a prodrug' strategy for tissue/organ-specific targeted drug delivery.

  9. Metabolism, Biochemical Actions, and Chemical Synthesis of Anticancer Nucleosides, Nucleotides, and Base Analogs.

    Science.gov (United States)

    Shelton, Jadd; Lu, Xiao; Hollenbaugh, Joseph A; Cho, Jong Hyun; Amblard, Franck; Schinazi, Raymond F

    2016-12-14

    Nucleoside, nucleotide, and base analogs have been in the clinic for decades to treat both viral pathogens and neoplasms. More than 20% of patients on anticancer chemotherapy have been treated with one or more of these analogs. This review focuses on the chemical synthesis and biology of anticancer nucleoside, nucleotide, and base analogs that are FDA-approved and in clinical development since 2000. We highlight the cellular biology and clinical biology of analogs, drug resistance mechanisms, and compound specificity towards different cancer types. Furthermore, we explore analog syntheses as well as improved and scale-up syntheses. We conclude with a discussion on what might lie ahead for medicinal chemists, biologists, and physicians as they try to improve analog efficacy through prodrug strategies and drug combinations.

  10. Crystal structures of HIV-1 reverse transcriptase complexes with thiocarbamate non-nucleoside inhibitors

    International Nuclear Information System (INIS)

    Spallarossa, Andrea; Cesarini, Sara; Ranise, Angelo; Ponassi, Marco; Unge, Torsten; Bolognesi, Martino

    2008-01-01

    O-Phthalimidoethyl-N-arylthiocarbamates (TCs) have been recently identified as a new class of potent HIV-1 reverse transcriptase (RT) non-nucleoside inhibitors (NNRTIs), by means of computer-aided drug design techniques [Ranise A. Spallarossa, S. Cesarini, F. Bondavalli, S. Schenone, O. Bruno, G. Menozzi, P. Fossa, L. Mosti, M. La Colla, et al., Structure-based design, parallel synthesis, structure-activity relationship, and molecular modeling studies of thiocarbamates, new potent non-nucleoside HIV-1 reverse transcriptase inhibitor isosteres of phenethylthiazolylthiourea derivatives, J. Med. Chem. 48 (2005) 3858-3873]. To elucidate the atomic details of RT/TC interaction and validate an earlier TC docking model, the structures of three RT/TC complexes were determined at 2.8-3.0 A resolution by X-ray crystallography. The conformations adopted by the enzyme-bound TCs were analyzed and compared with those of bioisosterically related NNRTIs

  11. Purification, crystallization, and preliminary X-ray diffraction study of purine nucleoside phosphorylase from E. coli

    Energy Technology Data Exchange (ETDEWEB)

    Abramchik, Yu. A., E-mail: inna@ns.crys.ras.ru; Timofeev, V. I., E-mail: espiov@ibch.ru; Zhukhlistova, N. E., E-mail: tostars@mail.ru [Russian Academy of Sciences, Shubnikov Institute of Crystallography (Russian Federation); Muravieva, T. I.; Esipov, R. S. [Russian Academy of Sciences, Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry (Russian Federation); Kuranova, I. P. [Russian Academy of Sciences, Shubnikov Institute of Crystallography (Russian Federation)

    2015-07-15

    Crystals of E. coli purine nucleoside phosphorylase were grown in microgravity by the capillary counter-diffusion method through a gel layer. The X-ray diffraction data set suitable for the determination of the three-dimensional structure at atomic resolution was collected from one crystal at the Spring-8 synchrotron facility to 0.99 Å resolution. The crystals belong to sp. gr. P2{sub 1} and have the following unit-cell parameters: a = 74.1 Å, b = 110.2 Å, c = 88.2 Å, α = γ = 90°, β = 111.08°. The crystal contains six subunits of the enzyme comprising a hexamer per asymmetric unit. The hexamer is the biological active form of E. coli. purine nucleoside phosphorylase.

  12. Mechanistic Exploration of the Competition Relationship between a Ketone and C═C, C═N, or C═S Bond in the Rh(III)-Catalyzed Carbocyclization Reactions.

    Science.gov (United States)

    Xing, Yang-Yang; Liu, Jian-Biao; Sun, Chuan-Zhi; Huang, Fang; Chen, De-Zhan

    2018-04-02

    The introduction of a C═O, C═C, C═S, or C═N bond has emerged as an effective strategy for carbocycle synthesis. A computational mechanistic study of Rh(III)-catalyzed coupling of alkynes with enaminones, sulfoxonium ylides, or α-carbonyl-nitrones was carried out. Our results uncover the roles of dual directing groups in the three substrates and confirm that the ketone acts as the role of the directing group while the C═C, C═N, or C═S bond serves as the cyclization site. By comparing the coordination of the ketone versus the C═C, C═N, or C═S bond, as well as the chemoselectivity concerning the six- versus five-membered formation, a competition relationship is revealed within the dual directing groups. Furthermore, after the alkyne insertion, instead of the originally proposed direct reductive elimination mechanism, the ketone enolization is found to be essential prior to the reductive elimination. The following C(sp 2 )-C(sp 2 ) reductive elimination is more favorable than the C(sp 3 )-C(sp 2 ) formation, which can be explained by the aromaticity difference in the corresponding transition states. The substituent effect on controlling the selectivity was also discussed.

  13. Evidence for purine nucleoside phosphorylase (PNP) release from rat C6 glioma cells.

    Science.gov (United States)

    Giuliani, Patricia; Zuccarini, Mariachiara; Buccella, Silvana; Peña-Altamira, Luis Emiliano; Polazzi, Elisabetta; Virgili, Marco; Monti, Barbara; Poli, Alessandro; Rathbone, Michel P; Di Iorio, Patrizia; Ciccarelli, Renata; Caciagli, Francesco

    2017-04-01

    Intracellular purine turnover is mainly oriented to preserving the level of triphosphate nucleotides, fundamental molecules in vital cell functions that, when released outside cells, act as receptor signals. Conversely, high levels of purine bases and uric acid are found in the extracellular milieu, even in resting conditions. These compounds could derive from nucleosides/bases that, having escaped to cell reuptake, are metabolized by extracellular enzymes similar to the cytosolic ones. Focusing on purine nucleoside phosphorylase (PNP) that catalyzes the reversible phosphorolysis of purine (deoxy)-nucleosides/bases, we found that it is constitutively released from cultured rat C6 glioma cells into the medium, and has a molecular weight and enzyme activity similar to the cytosolic enzyme. Cell exposure to 10 μM ATP or guanosine triphosphate (GTP) increased the extracellular amount of all corresponding purines without modifying the levels/activity of released PNP, whereas selective activation of ATP P2Y 1 or adenosine A 2A metabotropic receptors increased PNP release and purine base formation. The reduction to 1% in oxygen supply (2 h) to cells decreased the levels of released PNP, leading to an increased presence of extracellular nucleosides and to a reduced formation of xanthine and uric acid. Conversely, 2 h cell re-oxygenation enhanced the extracellular amounts of both PNP and purine bases. Thus, hypoxia and re-oxygenation modulated in opposite manner the PNP release/activity and, thereby, the extracellular formation of purine metabolism end-products. In conclusion, extracellular PNP and likely other enzymes deputed to purine base metabolism are released from cells, contributing to the purinergic system homeostasis and exhibiting an important pathophysiological role. © 2017 International Society for Neurochemistry.

  14. Structure-activity relationships of nucleoside analogues for inhibition of tick-borne encephalitis virus

    Czech Academy of Sciences Publication Activity Database

    Eyer, L.; Šmídková, Markéta; Nencka, Radim; Neča, J.; Kastl, T.; Palus, Martin; De Clercq, E.; Růžek, Daniel

    2016-01-01

    Roč. 133, Sep (2016), s. 119-129 ISSN 0166-3542 R&D Projects: GA MZd(CZ) NV16-34238A; GA ČR(CZ) GA16-20054S Institutional support: RVO:61388963 ; RVO:60077344 Keywords : structure-activity relationship * tick-borne encephalitis * nucleoside inhibitor * antiviral activity * cytotoxicity Subject RIV: CC - Organic Chemistry; EE - Microbiology, Virology (BC-A) Impact factor: 4.271, year: 2016

  15. New prodrugs of two pyrimidine acyclic nucleoside phosphonates: Synthesis and antiviral activity

    Czech Academy of Sciences Publication Activity Database

    Krečmerová, Marcela; Dračínský, Martin; Snoeck, R.; Balzarini, J.; Pomeisl, Karel; Andrei, G.

    2017-01-01

    Roč. 25, č. 17 (2017), s. 4637-4648 ISSN 0968-0896 R&D Projects: GA ČR(CZ) GA14-00522S Institutional support: RVO:61388963 Keywords : acyclic nucleoside phosphonates * open-ring * PMEO-DAPy * 5-azacytosine * PME-azaC * HPMP-5-azaC Subject RIV: CC - Organic Chemistry OBOR OECD: Organic chemistry Impact factor: 2.930, year: 2016

  16. Cloning, purification and characterisation of a recombinant purine nucleoside phosphorylase from Bacillus halodurans Alk36

    CSIR Research Space (South Africa)

    Visser, Daniel F

    2010-03-01

    Full Text Available . 2008; Rocchietti et al. 2004). Enzymes provide regio- and stereoselectivity, and hence are an ideal option for nucleoside transglyco- sylation (Prasad et al. 1999; Utagawa 1999). The hydro- lysis reaction for the ribose decoupling reaction can... B. halodurans Alk36. Methods Materials All restriction enzymes and the T4-DNA ligase were pur- chased from Fermentas (Lithuania). The Roche high- fidelity PCR mix was used for all polymerase chain reac- tions. SDS-PAGE markers were purchased...

  17. Nucleoside 5'-C-phosphonates: reactivity of the alpha-hydroxyphosphonate moiety

    Czech Academy of Sciences Publication Activity Database

    Králíková, Šárka; Buděšínský, Miloš; Masojídková, Milena; Rosenberg, Ivan

    2006-01-01

    Roč. 62, č. 20 (2006), s. 4917-4932 ISSN 0040-4020 R&D Projects: GA ČR(CZ) GP203/04/P273; GA ČR(CZ) GA203/05/0832; GA AV ČR(CZ) IAA4055101 Institutional research plan: CEZ:AV0Z40550506 Keywords : nucleoside 5'-aldehydes * oxidation * phosphonates Subject RIV: CC - Organic Chemistry Impact factor: 2.817, year: 2006

  18. Discovery of triazolinone non-nucleoside inhibitors of HIV reverse transcriptase.

    Science.gov (United States)

    Sweeney, Zachary K; Acharya, Sahaja; Briggs, Andrew; Dunn, James P; Elworthy, Todd R; Fretland, Jennifer; Giannetti, Anthony M; Heilek, Gabrielle; Li, Yu; Kaiser, Ann C; Martin, Michael; Saito, Y David; Smith, Mark; Suh, Judy M; Swallow, Steven; Wu, Jeffrey; Hang, Julie Q; Zhou, Amy S; Klumpp, Klaus

    2008-08-01

    Novel non-nucleoside inhibitors of HIV-RT that contain pyridazinone isosteres were prepared, and a series of triazolinones were found to be potent inhibitors of HIV replication. These compounds were active against several NNRTI-resistant virus strains. Pharmacokinetic studies indicated that inhibitor 7e has good bioavailability in rats. Several fragments of inhibitor 7c were prepared, and the binding of these compounds to HIV-RT was analyzed by surface plasmon resonance spectroscopy.

  19. Copper-mediated arylsulfanylations and arylselanylations of pyrimidine or 7-deazapurine nucleosides and nucleotides

    Czech Academy of Sciences Publication Activity Database

    Botha, Filip; Slavíčková, Michaela; Pohl, Radek; Hocek, Michal

    2016-01-01

    Roč. 14, č. 42 (2016), s. 10018-10022 ISSN 1477-0520 R&D Projects: GA ČR GA14-04289S Institutional support: RVO:61388963 Keywords : cross-link formation * unprotected nucleosides * photochemical control Subject RIV: CC - Organic Chemistry Impact factor: 3.564, year: 2016 http://pubs.rsc.org/en/content/articlepdf/2016/ob/c6ob01917j

  20. Mechanisms by Which Human DNA Primase Chooses To Polymerize a Nucleoside Triphosphate

    Czech Academy of Sciences Publication Activity Database

    Urban, M.; Joubert, Nicolas; Purse, B. W.; Hocek, Michal; Kuchta, R. D.

    2010-01-01

    Roč. 49, č. 4 (2010), s. 727-735 ISSN 0006-2960 R&D Projects: GA MŠk LC512; GA AV ČR IAA400550902 Grant - others:NIH(US) GM54194 Institutional research plan: CEZ:AV0Z40550506 Keywords : C-nucleosides * DNA polymerase * primase * mechanism Subject RIV: CC - Organic Chemistry Impact factor: 3.226, year: 2010

  1. New Cerebroside and Nucleoside Derivatives from a Red Sea Strain of the Marine Cyanobacterium Moorea producens

    OpenAIRE

    Diaa T.A. Youssef; Sabrin R.M. Ibrahim; Lamiaa A. Shaala; Gamal A. Mohamed; Zainy M. Banjar

    2016-01-01

    In the course of our ongoing efforts to identify marine-derived bioactive compounds, the marine cyanobacterium Moorea producens was investigated. The organic extract of the Red Sea cyanobacterium afforded one new cerebroside, mooreaside A (1), two new nucleoside derivatives, 3-acetyl-2′-deoxyuridine (2) and 3-phenylethyl-2′-deoxyuridine (3), along with the previously reported compounds thymidine (4) and 2,3-dihydroxypropyl heptacosanoate (5). The structures of the compounds were determined by...

  2. Differential effects of acyclic nucleoside phosphonates on nitric oxide and cytokines in rat hepatocytes and macrophages

    Czech Academy of Sciences Publication Activity Database

    Kostecká, Petra; Holý, Antonín; Farghali, H.; Zídek, Zdeněk; Kmoníčková, Eva

    2012-01-01

    Roč. 12, č. 2 (2012), s. 342-349 ISSN 1567-5769 R&D Projects: GA MŠk 1M0508 Institutional research plan: CEZ:AV0Z50390512; CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * cytokines * nitric oxide Subject RIV: FR - Pharmacology ; Medidal Chemistry; CC - Organic Chemistry (UOCHB-X) Impact factor: 2.417, year: 2012

  3. Enhanced Topical and Transdermal Delivery of Antineoplastic and Antiviral Acyclic Nucleoside Phosphonate cPr-PMEDAP

    Czech Academy of Sciences Publication Activity Database

    Vávrová, K.; Kovaříková, P.; Školová, B.; Líbalová, M.; Roh, J.; Čáp, R.; Holý, Antonín; Hrabálek, A.

    2011-01-01

    Roč. 28, č. 12 (2011), s. 3105-3115 ISSN 0724-8741 R&D Projects: GA MŠk 1M0508 Grant - others:GA ČR(CZ) GAP207/11/0365 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * antiviral s * antineoplastics * permeation enhancer * topical skin application * transdermal delivery Subject RIV: CC - Organic Chemistry Impact factor: 4.093, year: 2011

  4. Comparison of Clostridium difficile detection by monolayer and by inhibition of nucleoside uptake

    Energy Technology Data Exchange (ETDEWEB)

    Fuhr, J.E.; Trent, D.J.; Collmann, I.R.

    1987-02-01

    Detection and identification of Clostridium difficile toxin by traditional monolayer assay were compared with results obtained by a new procedure based on toxin-dependent inhibition of target cell uptake of a radioactive nucleoside. A high degree of correlation was noted between the two determinations. Although the new procedure was quantitative and objective, its value is seen at present as a rapid screen that may support results obtained in monolayers and as a potential assay for other, currently unidentified, toxins.

  5. Novel modified purine bases and nucleosides: new methodologies of synthesis and biological activity

    Czech Academy of Sciences Publication Activity Database

    Hocek, Michal

    -, č. 49 (2005), s. 29-30 ISSN 0261-3166. [International Symposium on Nucleic Acids Chemistry /4./. Fukuoka, 20.09.2005-22.09.2005] R&D Projects: GA ČR(CZ) GA203/03/0035; GA MŠk(CZ) 1M0508 Institutional research plan: CEZ:AV0Z40550506 Keywords : purines * nucleosides * synthesis Subject RIV: CC - Organic Chemistry

  6. Botrytis cinerea can import and utilize nucleosides in salvage and catabolism and BcENT functions as high affinity nucleoside transporter.

    Science.gov (United States)

    Daumann, Manuel; Golfier, Philippe; Knüppel, Nathalie; Hahn, Matthias; Möhlmann, Torsten

    2016-08-01

    Nucleotide de novo synthesis is an essential pathway in nearly all organisms. Transport processes as well as salvage and catabolism of nucleotides and pathway intermediates are required to balance nucleotide pools. We have analysed the genome of the fungal plant pathogen Botrytis cinerea for genes involved in nucleotide metabolism and found a complete set of genes necessary for purine and pyrimidine uptake and salvage based on homology of the gene products to corresponding proteins from Aspergillus nidulans. Candidate genes required for a complete purine catabolic sequence were identified in addition. These analyses were complemented by growth tests showing functional transport and salvage activity for pyrimidines. Growth of B. cinerea mycelium in nitrogen free medium could be restored by addition of purines, indicating the presence of a functional purine catabolism, whereas pyrimidines did not support growth. Bcin07g05490 (BcENT) was identified as sole member of the equilibrative nucleoside transporter (ENT) family. The protein synthesized in Saccharomyces cerevisiae revealed high affinity transport of adenosine (KM = 6.81 μM) and uridine (KM=9.04 μM). Furthermore, a BcENT knockout mutant was generated and tested in a range of growth and infection assays. These results provide detailed insight in the use of externally supplied nucleobases and nucleosides by B. cinerea. Copyright © 2016 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  7. Preliminary crystallographic studies of purine nucleoside phosphorylase from the cariogenic pathogen Streptococcus mutans

    International Nuclear Information System (INIS)

    Hou, Qiao-Ming; Liu, Xiang; Brostromer, Erik; Li, Lan-Fen; Su, Xiao-Dong

    2009-01-01

    Purine nucleoside phosphorylase (PNP), which is a pivotal enzyme in the nucleotide-salvage pathway, has been expressed in Escherichia coli strain BL21 (DE3) in a soluble form at a high level. After purification of the PNP enzyme, the protein was crystallized using the sitting-drop vapour-diffusion technique. The punA gene of the cariogenic pathogen Streptococcus mutans encodes purine nucleoside phosphorylase (PNP), which is a pivotal enzyme in the nucleotide-salvage pathway, catalyzing the phosphorolysis of purine nucleosides to generate purine bases and α-ribose 1-phosphate. In the present work, the PNP protein was expressed in Escherichia coli strain BL21 (DE3) in a soluble form at a high level. After purification of the PNP enzyme, the protein was crystallized using the sitting-drop vapour-diffusion technique; the crystals diffracted to 1.6 Å resolution at best. The crystals belonged to space group H3, with unit-cell parameters a = b = 113.0, c = 60.1 Å

  8. The chemistry of nicotinamide adenine dinucleotide (NAD) analogues containing C-nucleosides related to nicotinamide riboside.

    Science.gov (United States)

    Pankiewicz, Krzysztof W; Watanabe, Kyoichi A; Lesiak-Watanabe, Krystyna; Goldstein, Barry M; Jayaram, Hiremagalur N

    2002-04-01

    Oncolytic C-nucleosides, tiazofurin (2-beta-D-ribofuranosylthiazole-4-carboxamide) and benzamide riboside (3-beta-D-ribofuranosylbenzamide) are converted in cell into active metabolites thiazole-4-carboxamide- and benzamide adenine dinucleotide, TAD and BAD, respectively. TAD and BAD as NAD analogues were found to bind at the nicotinamide adenine dinucleotide (cofactor NAD) site of inosine monophosphate dehydrogenase (IMPDH), an important target in cancer treatment. The synthesis and evaluation of anticancer activity of a number of C-nucleosides related to tiazofurin and nicotinamide riboside then followed and are reviewed herein. Interestingly, pyridine C-nucleosides (such as C-nicotinamide riboside) are not metabolized into the corresponding NAD analogues in cell. Their conversion by chemical methods is described. As dinucleotides these compounds show inhibition of IMPDH in low micromolar level. Also, the synthesis of BAD in metabolically stable bis(phosphonate) form is discussed indicating the usefulness of such preformed inhibitors in drug development. Among tiazofurin analogues, Franchetti and Grifantini found, that the replacement of the sulfur by oxygen (as in oxazafurin) but not the removal of nitrogen (tiophenfurin) of the thiazole ring resulted in inactive compounds. The anti cancer activity of their synthetic dinucleotide analogues indicate that inactive compounds are not only poorly metabolized in cell but also are weak inhibitors of IMPDH as dinucleotides.

  9. In Silico Investigation of Flavonoids as Potential Trypanosomal Nucleoside Hydrolase Inhibitors

    Directory of Open Access Journals (Sweden)

    Christina Hung Hung Ha

    2015-01-01

    Full Text Available Human African Trypanosomiasis is endemic to 37 countries of sub-Saharan Africa. It is caused by two related species of Trypanosoma brucei. Current therapies suffer from resistance and public accessibility of expensive medicines. Finding safer and effective therapies of natural origin is being extensively explored worldwide. Pentamidine is the only available therapy for inhibiting the P2 adenosine transporter involved in the purine salvage pathway of the trypanosomatids. The objective of the present study is to use computational studies for the investigation of the probable trypanocidal mechanism of flavonoids. Docking experiments were carried out on eight flavonoids of varying level of hydroxylation, namely, flavone, 5-hydroxyflavone, 7-hydroxyflavone, chrysin, apigenin, kaempferol, fisetin, and quercetin. Using AutoDock 4.2, these compounds were tested for their affinity towards inosine-adenosine-guanosine nucleoside hydrolase and the inosine-guanosine nucleoside hydrolase, the major enzymes of the purine salvage pathway. Our results showed that all of the eight tested flavonoids showed high affinities for both hydrolases (lowest free binding energy ranging from −10.23 to −7.14 kcal/mol. These compounds, especially the hydroxylated derivatives, could be further studied as potential inhibitors of the nucleoside hydrolases.

  10. Zika virus protection by a single low-dose nucleoside-modified mRNA vaccination.

    Science.gov (United States)

    Pardi, Norbert; Hogan, Michael J; Pelc, Rebecca S; Muramatsu, Hiromi; Andersen, Hanne; DeMaso, Christina R; Dowd, Kimberly A; Sutherland, Laura L; Scearce, Richard M; Parks, Robert; Wagner, Wendeline; Granados, Alex; Greenhouse, Jack; Walker, Michelle; Willis, Elinor; Yu, Jae-Sung; McGee, Charles E; Sempowski, Gregory D; Mui, Barbara L; Tam, Ying K; Huang, Yan-Jang; Vanlandingham, Dana; Holmes, Veronica M; Balachandran, Harikrishnan; Sahu, Sujata; Lifton, Michelle; Higgs, Stephen; Hensley, Scott E; Madden, Thomas D; Hope, Michael J; Karikó, Katalin; Santra, Sampa; Graham, Barney S; Lewis, Mark G; Pierson, Theodore C; Haynes, Barton F; Weissman, Drew

    2017-03-09

    Zika virus (ZIKV) has recently emerged as a pandemic associated with severe neuropathology in newborns and adults. There are no ZIKV-specific treatments or preventatives. Therefore, the development of a safe and effective vaccine is a high priority. Messenger RNA (mRNA) has emerged as a versatile and highly effective platform to deliver vaccine antigens and therapeutic proteins. Here we demonstrate that a single low-dose intradermal immunization with lipid-nanoparticle-encapsulated nucleoside-modified mRNA (mRNA-LNP) encoding the pre-membrane and envelope glycoproteins of a strain from the ZIKV outbreak in 2013 elicited potent and durable neutralizing antibody responses in mice and non-human primates. Immunization with 30 μg of nucleoside-modified ZIKV mRNA-LNP protected mice against ZIKV challenges at 2 weeks or 5 months after vaccination, and a single dose of 50 μg was sufficient to protect non-human primates against a challenge at 5 weeks after vaccination. These data demonstrate that nucleoside-modified mRNA-LNP elicits rapid and durable protective immunity and therefore represents a new and promising vaccine candidate for the global fight against ZIKV.

  11. An unusual UMP C-5 methylase in nucleoside antibiotic polyoxin biosynthesis

    Directory of Open Access Journals (Sweden)

    Wenqing Chen

    2016-07-01

    Full Text Available ABSTRACT Polyoxin is a group of structurally-related peptidyl nucleoside antibiotics bearing C-5 modifications on the nucleoside skeleton. Although the structural diversity and bioactivity preference of polyoxin are, to some extent, affected by such modifications, the biosynthetic logic for their occurence remains obscure. Here we report the identification of PolB in polyoxin pathway as an unusual UMP C-5 methylase with thymidylate synthase activity which is responsible for the C-5 methylation of the nucleoside skeleton. To probe its molecular mechanism, we determined the crystal structures of PolB alone and in complexes with 5-Br UMP and 5-Br dUMP at 2.15 Å, 1.76 Å and 2.28 Å resolutions, respectively. Loop 1 (residues 117–131, Loop 2 (residues 192–201 and the substrate recognition peptide (residues 94–102 of PolB exhibit considerable conformational flexibility and adopt distinct structures upon binding to different substrate analogs. Consistent with the structural findings, a PolB homolog that harbors an identical function from Streptomyces viridochromogenes DSM 40736 was identified. The discovery of UMP C5-methylase opens the way to rational pathway engineering for polyoxin component optimization, and will also enrich the toolbox for natural nucleotide chemistry.

  12. Synthesis and anti-cancer activities of new sulfonamides 4-substituted-triazolyl nucleosides.

    Science.gov (United States)

    Alaoui, Soukaina; Dufies, Maeva; Driowya, Mohsine; Demange, Luc; Bougrin, Khalid; Robert, Guillaume; Auberger, Patrick; Pagès, Gilles; Benhida, Rachid

    2017-05-01

    Nucleoside analogues are among the most known drugs commonly used in antiviral and anticancer chemotherapies. Among them, those featuring a five-membered ring nucleobase are of utmost interest such as the anti-cancer agent AICAR or the anti-viral drug ribavirin. Despite its low activity in vitro in different cell lines, AICAR is under clinical development for several pathologies, thanks to its original mode of action. Indeed, AICAR induced autophagy cell death and is able, following this mechanism, to circumvent resistance to apoptotic drugs including kinase inhibitors currently on the market. To improve the activity of AICAR, we report herein an efficient synthesis of new series of sulfonamide-4-substituted-1,2,3-triazolyl nucleosides using a Cu-catalyzed 1,3-dipolar cycloaddition. All these molecules have been fully characterized and evaluated against two aggressive tumor cell lines, RCC4 and MDA-MB-231. Among them, nucleoside analogue 5i belonging to the ribose series was found to be 19 to 66-fold more active than AICAR. Western blot analyses on RCC4 cells showed that 5i displayed an interesting mode of action by inducing both apoptosis and autophagy cell death, making therefore this class of molecules highly promising for further hit-to-lead optimization. Copyright © 2017 Elsevier Ltd. All rights reserved.

  13. Training Teachers to Conduct Trial-Based Functional Analyses

    Science.gov (United States)

    Kunnavatana, S. Shanun; Bloom, Sarah E.; Samaha, Andrew L.; Dayton, Elizabeth

    2013-01-01

    The trial-based functional analysis (FA) is a promising approach to identification of behavioral function and is especially suited for use in educational settings. Not all studies on trial-based FA have included teachers as therapists, and those studies that have, included minimal information on teacher training. The purpose of this study was to…

  14. Novel base-functionalized DNA. Efficient methodology for construction and bioanalytical applications

    Czech Academy of Sciences Publication Activity Database

    Hocek, Michal; Vrábel, Milan; Cahová, Hana; Riedl, Jan; Kalachová, Lubica; Pivoňková, Hana; Horáková Brázdilová, Petra; Havran, Luděk; Fojta, Miroslav

    -, č. 52 (2008), s. 53-54 ISSN 0261-3166. [Joint Symposium of the International Roundtable on Nucleosides, Nucleotides and Nucleic Acids /18./ and the International Symposium on Nucleic Acid Chemistry /35./. Kyoto, 08.09.2008-12.09.2008] R&D Projects: GA ČR GA203/07/1195; GA MŠk LC512; GA MŠk(CZ) LC06035 Institutional research plan: CEZ:AV0Z40550506; CEZ:AV0Z50040507 Keywords : cross-coupling reactions * nucleoside triphosphates * DNA Subject RIV: CC - Organic Chemistry

  15. The chemoenzymatic synthesis of clofarabine and related 2′-deoxyfluoroarabinosyl nucleosides: the electronic and stereochemical factors determining substrate recognition by E. coli nucleoside phosphorylases

    Directory of Open Access Journals (Sweden)

    Ilja V. Fateev

    2014-07-01

    Full Text Available Two approaches to the synthesis of 2-chloro-9-(2-deoxy-2-fluoro-β-D-arabinofuranosyladenine (1, clofarabine were studied. The first approach consists in the chemical synthesis of 2-deoxy-2-fluoro-α-D-arabinofuranose-1-phosphate (12a, 2FAra-1P via three step conversion of 1,3,5-tri-O-benzoyl-2-deoxy-2-fluoro-α-D-arabinofuranose (9 into the phosphate 12a without isolation of intermediary products. Condensation of 12a with 2-chloroadenine catalyzed by the recombinant E. coli purine nucleoside phosphorylase (PNP resulted in the formation of clofarabine in 67% yield. The reaction was also studied with a number of purine bases (2-aminoadenine and hypoxanthine, their analogues (5-aza-7-deazaguanine and 8-aza-7-deazahypoxanthine and thymine. The results were compared with those of a similar reaction with α-D-arabinofuranose-1-phosphate (13a, Ara-1P. Differences of the reactivity of various substrates were analyzed by ab initio calculations in terms of the electronic structure (natural purines vs analogues and stereochemical features (2FAra-1P vs Ara-1P of the studied compounds to determine the substrate recognition by E. coli nucleoside phosphorylases. The second approach starts with the cascade one-pot enzymatic transformation of 2-deoxy-2-fluoro-D-arabinose into the phosphate 12a, followed by its condensation with 2-chloroadenine thereby affording clofarabine in ca. 48% yield in 24 h. The following recombinant E. coli enzymes catalyze the sequential conversion of 2-deoxy-2-fluoro-D-arabinose into the phosphate 12a: ribokinase (2-deoxy-2-fluoro-D-arabinofuranose-5-phosphate, phosphopentomutase (PPN; no 1,6-diphosphates of D-hexoses as co-factors required (12a, and finally PNP. The substrate activities of D-arabinose, D-ribose and D-xylose in the similar cascade syntheses of the relevant 2-chloroadenine nucleosides were studied and compared with the activities of 2-deoxy-2-fluoro-D-arabinose. As expected, D-ribose exhibited the best substrate

  16. The chemoenzymatic synthesis of clofarabine and related 2'-deoxyfluoroarabinosyl nucleosides: the electronic and stereochemical factors determining substrate recognition by E. coli nucleoside phosphorylases.

    Science.gov (United States)

    Fateev, Ilja V; Antonov, Konstantin V; Konstantinova, Irina D; Muravyova, Tatyana I; Seela, Frank; Esipov, Roman S; Miroshnikov, Anatoly I; Mikhailopulo, Igor A

    2014-01-01

    Two approaches to the synthesis of 2-chloro-9-(2-deoxy-2-fluoro-β-D-arabinofuranosyl)adenine (1, clofarabine) were studied. The first approach consists in the chemical synthesis of 2-deoxy-2-fluoro-α-D-arabinofuranose-1-phosphate (12a, (2F)Ara-1P) via three step conversion of 1,3,5-tri-O-benzoyl-2-deoxy-2-fluoro-α-D-arabinofuranose (9) into the phosphate 12a without isolation of intermediary products. Condensation of 12a with 2-chloroadenine catalyzed by the recombinant E. coli purine nucleoside phosphorylase (PNP) resulted in the formation of clofarabine in 67% yield. The reaction was also studied with a number of purine bases (2-aminoadenine and hypoxanthine), their analogues (5-aza-7-deazaguanine and 8-aza-7-deazahypoxanthine) and thymine. The results were compared with those of a similar reaction with α-D-arabinofuranose-1-phosphate (13a, Ara-1P). Differences of the reactivity of various substrates were analyzed by ab initio calculations in terms of the electronic structure (natural purines vs analogues) and stereochemical features ((2F)Ara-1P vs Ara-1P) of the studied compounds to determine the substrate recognition by E. coli nucleoside phosphorylases. The second approach starts with the cascade one-pot enzymatic transformation of 2-deoxy-2-fluoro-D-arabinose into the phosphate 12a, followed by its condensation with 2-chloroadenine thereby affording clofarabine in ca. 48% yield in 24 h. The following recombinant E. coli enzymes catalyze the sequential conversion of 2-deoxy-2-fluoro-D-arabinose into the phosphate 12a: ribokinase (2-deoxy-2-fluoro-D-arabinofuranose-5-phosphate), phosphopentomutase (PPN; no 1,6-diphosphates of D-hexoses as co-factors required) (12a), and finally PNP. The substrate activities of D-arabinose, D-ribose and D-xylose in the similar cascade syntheses of the relevant 2-chloroadenine nucleosides were studied and compared with the activities of 2-deoxy-2-fluoro-D-arabinose. As expected, D-ribose exhibited the best substrate activity

  17. Inhibitory effect of extracellular purine nucleotide and nucleoside concentrations on T cell proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Weiler, Monica [Department of Medicine III and Transfusion Medicine, University Hospital Grosshadern, Ludwig-Maximilians-University, Munich (Germany); Schmetzer, Helga [Helmholtz Center Munich (Germany); German Research Center for Environmental Health, Munich (Germany); Braeu, Marion; Buhmann, Raymund [Helmholtz Center Munich (Germany); German Research Center for Environmental Health, Munich (Germany); Department of Medicine III and Transfusion Medicine, University Hospital Grosshadern, Ludwig-Maximilians-University, Munich (Germany)

    2016-11-15

    The release of nucleic acids and derivatives after tissue-injury may affect cellular immune-response. We studied the impact of extracellular ribo-, desoxyribonucleotides and nucleosides on T-cell immunity. Peripheral-blood-mononuclear-cells (PBMCs) or isolated CD3{sup +}T-cells obtained from 6 healthy donors were stimulated via CD3/CD28 Dynabeads or dendritic cells (DCs) in the presence or absence of pyrimidine-, purine-nucleotides and -nucleosides (range 2–200 µM). Addition of deoxy-, guanosine-triphosphate (dGTP, GTP) and guanosine resulted concentration dependent in a complete, adenosine-triphosphate (ATP) in a partial inhibition of the induced T-cell-proliferation. Deoxyadenosine-triphosphate (dATP), adenosine and the pyrimidine-ribo- and -deoxyribonucleotides displayed no inhibitory capacity. Inhibitory effects of dGTP and GTP, but not of guanosine and ATP were culture-media-dependent and could be almost abrogated by use of the serum-free lymphocyte-culture-media X-Vivo15 instead of RPMI1640 with standard-supplementation. In contrast to RPMI1640, X-Vivo15 resulted in a significant down-regulation of the cell-surface-located ectonucleotidases CD39 (Ecto-Apyrase) and CD73 (Ecto-5′-Nucleotidase), critical for the extracellular nucleotides-hydrolysis to nucleosides, explaining the loss of inhibition mediated by dGTP and GTP, but not Guanosine. In line with previous findings ATP was found to exert immunosuppressive effects on T-cell-proliferation. Purine-nucleotides, dGTP and GTP displayed a higher inhibitory capacity, but seem to be strictly dependent on the microenvironmental conditions modulating the responsiveness of the respective T-lymphocytes. Further evaluation of experimental and respective clinical settings should anticipate these findings.

  18. Pharmacogenetic characterization of naturally occurring germline NT5C1A variants to chemotherapeutic nucleoside analogs

    Science.gov (United States)

    Saliba, Jason; Zabriskie, Ryan; Ghosh, Rajarshi; Powell, Bradford C; Hicks, Stephanie; Kimmel, Marek; Meng, Qingchang; Ritter, Deborah I; Wheeler, David A; Gibbs, Richard A; Tsai, Francis T F; Plon, Sharon E

    2016-01-01

    Background Mutations or alteration in expression of the 5’ nucleotidase gene family can confer altered responses to treatment with nucleoside analogs. While investigating leukemia susceptibility genes, we discovered a very rare p.L254P NT5C1A missense variant in the substrate recognition motif. Given the paucity of cellular drug response data from NT5C1A germline variation, we characterized p.L254P and eight rare variants of NT5C1A from genomic databases. Methods Through lentiviral infection, we created HEK293 cell lines that stably overexpress wildtype NT5C1A, p.L254P, or eight NT5C1A variants reported in the NHLBI Exome Variant server (one truncating and seven missense). IC50 values were determined by cytotoxicity assays after exposure to chemotherapeutic nucleoside analogs (Cladribine, Gemcitabine, 5-Fluorouracil). In addition, we used structure-based homology modeling to generate a 3D model for the C-terminal region of NT5C1A. Results The p.R180X (truncating), p.A214T, and p.L254P missense changes were the only variants that significantly impaired protein function across all nucleotide analogs tested (>5-fold difference versus WT; p<.05). Several of the remaining variants individually displayed differential effects (both more and less resistant) across the analogs tested. The homology model provided a structural framework to understand the impact of NT5C1A mutants on catalysis and drug processing. The model predicted active site residues within NT5C1A motif III and we experimentally confirmed that p.K314 (not p.K320) is required for NT5C1A activity. Conclusion We characterized germline variation and predicted protein structures of NT5C1A. Individual missense changes showed substantial variation in response to the different nucleoside analogs tested, which may impact patients’ responses to treatment. PMID:26906009

  19. Inhibitory effect of extracellular purine nucleotide and nucleoside concentrations on T cell proliferation

    International Nuclear Information System (INIS)

    Weiler, Monica; Schmetzer, Helga; Braeu, Marion; Buhmann, Raymund

    2016-01-01

    The release of nucleic acids and derivatives after tissue-injury may affect cellular immune-response. We studied the impact of extracellular ribo-, desoxyribonucleotides and nucleosides on T-cell immunity. Peripheral-blood-mononuclear-cells (PBMCs) or isolated CD3 + T-cells obtained from 6 healthy donors were stimulated via CD3/CD28 Dynabeads or dendritic cells (DCs) in the presence or absence of pyrimidine-, purine-nucleotides and -nucleosides (range 2–200 µM). Addition of deoxy-, guanosine-triphosphate (dGTP, GTP) and guanosine resulted concentration dependent in a complete, adenosine-triphosphate (ATP) in a partial inhibition of the induced T-cell-proliferation. Deoxyadenosine-triphosphate (dATP), adenosine and the pyrimidine-ribo- and -deoxyribonucleotides displayed no inhibitory capacity. Inhibitory effects of dGTP and GTP, but not of guanosine and ATP were culture-media-dependent and could be almost abrogated by use of the serum-free lymphocyte-culture-media X-Vivo15 instead of RPMI1640 with standard-supplementation. In contrast to RPMI1640, X-Vivo15 resulted in a significant down-regulation of the cell-surface-located ectonucleotidases CD39 (Ecto-Apyrase) and CD73 (Ecto-5′-Nucleotidase), critical for the extracellular nucleotides-hydrolysis to nucleosides, explaining the loss of inhibition mediated by dGTP and GTP, but not Guanosine. In line with previous findings ATP was found to exert immunosuppressive effects on T-cell-proliferation. Purine-nucleotides, dGTP and GTP displayed a higher inhibitory capacity, but seem to be strictly dependent on the microenvironmental conditions modulating the responsiveness of the respective T-lymphocytes. Further evaluation of experimental and respective clinical settings should anticipate these findings.

  20. Structural and Dynamical Impact of a Universal Fluorescent Nucleoside Analogue Inserted Into a DNA Duplex.

    Science.gov (United States)

    Zargarian, Loussiné; Ben Imeddourene, Akli; Gavvala, Krishna; Barthes, Nicolas P F; Michel, Benoit Y; Kenfack, Cyril A; Morellet, Nelly; René, Brigitte; Fossé, Philippe; Burger, Alain; Mély, Yves; Mauffret, Olivier

    2017-12-21

    Recently, a 3-hydroxychromone based nucleoside 3HCnt has been developed as a highly environment-sensitive nucleoside surrogate to investigate protein-DNA interactions. When it is incorporated in DNA, the probe is up to 50-fold brighter than 2-aminopurine, the reference fluorescent nucleoside. Although the insertion of 3HCnt in DNA was previously shown to not alter the overall DNA structure, the possibility of the probe inducing local effects cannot be ruled out. Hence, a systematic structural and dynamic study is required to unveil the 3HCnt's limitations and to properly interpret the data obtained with this universal probe. Here, we investigated by NMR a 12-mer duplex, in which a central adenine was replaced by 3HCnt. The chemical shifts variations and nOe contacts revealed that the 3HCnt is well inserted in the DNA double helix with extensive stacking interactions with the neighbor base pairs. These observations are in excellent agreement with the steady-state and time-resolved fluorescence properties indicating that the 3HCnt fluorophore is protected from the solvent and does not exhibit rotational motion. The 3HCnt insertion in DNA is accompanied by the extrusion of the opposite nucleobase from the double helix. Molecular dynamics simulations using NMR-restraints demonstrated that 3HCnt fluorophore exhibits only translational dynamics. Taken together, our data showed an excellent intercalation of 3HCnt in the DNA double helix, which is accompanied by localized perturbations. This confirms 3HCnt as a highly promising tool for nucleic acid labeling and sensing.

  1. The Crystal Structure of Streptococcus pyogenes Uridine Phosphorylase Reveals a Distinct Subfamily of Nucleoside Phosphorylases

    Energy Technology Data Exchange (ETDEWEB)

    Tran, Timothy H.; Christoffersen, S.; Allan, Paula W.; Parker, William B.; Piskur, Jure; Serra, I.; Terreni, M.; Ealick, Steven E. (Cornell); (Pavia); (Lund); (Southern Research)

    2011-09-20

    Uridine phosphorylase (UP), a key enzyme in the pyrimidine salvage pathway, catalyzes the reversible phosphorolysis of uridine or 2'-deoxyuridine to uracil and ribose 1-phosphate or 2'-deoxyribose 1-phosphate. This enzyme belongs to the nucleoside phosphorylase I superfamily whose members show diverse specificity for nucleoside substrates. Phylogenetic analysis shows Streptococcus pyogenes uridine phosphorylase (SpUP) is found in a distinct branch of the pyrimidine subfamily of nucleoside phosphorylases. To further characterize SpUP, we determined the crystal structure in complex with the products, ribose 1-phosphate and uracil, at 1.8 {angstrom} resolution. Like Escherichia coli UP (EcUP), the biological unit of SpUP is a hexamer with an ?/? monomeric fold. A novel feature of the active site is the presence of His169, which structurally aligns with Arg168 of the EcUP structure. A second active site residue, Lys162, is not present in previously determined UP structures and interacts with O2 of uracil. Biochemical studies of wild-type SpUP showed that its substrate specificity is similar to that of EcUP, while EcUP is {approx}7-fold more efficient than SpUP. Biochemical studies of SpUP mutants showed that mutations of His169 reduced activity, while mutation of Lys162 abolished all activity, suggesting that the negative charge in the transition state resides mostly on uracil O2. This is in contrast to EcUP for which transition state stabilization occurs mostly at O4.

  2. Analysis of the Main Nucleosides in Cordyceps Sinensis by LC/ESI-MS

    Directory of Open Access Journals (Sweden)

    Yun-Biao He

    2010-01-01

    Full Text Available A sensitive, selective and reliable liquid chromatography-mass spectrometry coupled with electrospray ionization interface method for simultaneous separation and determination of thymine, adenine, adenosine and cordycepin in Cordyceps sinensis has been established. The optimum separation for these analytes was achieved using a gradient elution system and a 2.0 × 150 mm Shimadzu VP-ODS column. 2-Chloroadenosine was used as internal standard for this assay. [M+H]+ions at m/z 127, 136, 268, 252 and 302 were chosen and selective ion monitoring (SIM mode was used for quantitative analysis of the four main nucleosides. The regression equations were linear in the range of 1.0–117.5 μg·mL-1 for thymine, 1.8-127.0 μg·mL-1 for adenine, 0.6-114.0 μg·mL-1 for adenosine and 0.5-107.5 μg·mL-1 for cordycepin. The limits of quantitation (LOQ and detection (LOD were 1.0 and 0.2 μg·mL-1 for thymine, 1.8 and 0.6 μg·mL-1 for adenine, 0.6 and 0.1 μg·mL-1 for adenosine and 0.5 and 0.1 μg·mL-1 for cordycepin, respectively. The recoveries of the four nucleosides ranged from 98.47 to 99.32%. The developed method was successfully used to determine nucleosides in Cordyceps sinensis from different sources.

  3. The MONET trial: darunavir/ritonavir with or without nucleoside analogues, for patients with HIV RNA below 50 copies/ml

    DEFF Research Database (Denmark)

    Arribas, Jose R; Horban, Andrzej; Gerstoft, Jan

    2010-01-01

    In virologically suppressed patients, darunavir-ritonavir (DRV/r) monotherapy could maintain virological suppression similarly to DRV/r and two nucleosides.......In virologically suppressed patients, darunavir-ritonavir (DRV/r) monotherapy could maintain virological suppression similarly to DRV/r and two nucleosides....

  4. The identification and characterization of non-coding and coding RNAs and their modified nucleosides by mass spectrometry

    Science.gov (United States)

    Gaston, Kirk W; Limbach, Patrick A

    2014-01-01

    The analysis of ribonucleic acids (RNA) by mass spectrometry has been a valuable analytical approach for more than 25 years. In fact, mass spectrometry has become a method of choice for the analysis of modified nucleosides from RNA isolated out of biological samples. This review summarizes recent progress that has been made in both nucleoside and oligonucleotide mass spectral analysis. Applications of mass spectrometry in the identification, characterization and quantification of modified nucleosides are discussed. At the oligonucleotide level, advances in modern mass spectrometry approaches combined with the standard RNA modification mapping protocol enable the characterization of RNAs of varying lengths ranging from low molecular weight short interfering RNAs (siRNAs) to the extremely large 23 S rRNAs. New variations and improvements to this protocol are reviewed, including top-down strategies, as these developments now enable qualitative and quantitative measurements of RNA modification patterns in a variety of biological systems. PMID:25616408

  5. Enhancement of radiation-induced base release from nucleosides in alkaline solution: essential role of the O.- radical

    International Nuclear Information System (INIS)

    Scholes, M.L.; Schuchmann, M.N.; Sonntag, C. von

    1992-01-01

    The effect of pH on base release in the γ-radiolysis of N 2 O-saturated solutions of a number of nucleosides (including uridine, 3-methyluridine, 2', 3' -O-isopropylidene-uridine, and adenosine) has been investigated. For all these nucleosides, independent of the base or sugar moiety, base release is very low at pH below 10 (G∼(0.3-0.7) x 10 -7 mol J -1 ), but increases drastically to G∼(3-4) x 10 -7 mol J -1 at pH ≥ 13. It is concluded that the increase in base release at high pH is caused by the increasing participation of O .- , which, unlike . OH, attacks the nucleosides preferentially at their sugar moieties, and is not due to an OH - -induced radical transfer from the base to the sugar moiety. (author)

  6. Wet-dry cycles enable the parallel origin of canonical and non-canonical nucleosides by continuous synthesis.

    Science.gov (United States)

    Becker, Sidney; Schneider, Christina; Okamura, Hidenori; Crisp, Antony; Amatov, Tynchtyk; Dejmek, Milan; Carell, Thomas

    2018-01-11

    The molecules of life were created by a continuous physicochemical process on an early Earth. In this hadean environment, chemical transformations were driven by fluctuations of the naturally given physical parameters established for example by wet-dry cycles. These conditions might have allowed for the formation of (self)-replicating RNA as the fundamental biopolymer during chemical evolution. The question of how a complex multistep chemical synthesis of RNA building blocks was possible in such an environment remains unanswered. Here we report that geothermal fields could provide the right setup for establishing wet-dry cycles that allow for the synthesis of RNA nucleosides by continuous synthesis. Our model provides both the canonical and many ubiquitous non-canonical purine nucleosides in parallel by simple changes of physical parameters such as temperature, pH and concentration. The data show that modified nucleosides were potentially formed as competitor molecules. They could in this sense be considered as molecular fossils.

  7. Conformationally constrained nucleoside phosphonic acids - potent inhibitors of human mitochondrial and cytosolic 5'(3')-nucleotidases

    Czech Academy of Sciences Publication Activity Database

    Šimák, Ondřej; Pachl, Petr; Fábry, Milan; Buděšínský, Miloš; Jandušík, T.; Hnízda, Aleš; Skleničková, Radka; Petrová, Magdalena; Veverka, Václav; Řezáčová, Pavlína; Brynda, Jiří; Rosenberg, Ivan

    2014-01-01

    Roč. 12, č. 40 (2014), s. 7971-7982 ISSN 1477-0520 R&D Projects: GA ČR GA203/09/0820; GA ČR GA13-24880S; GA ČR GA13-26526S; GA MŠk(CZ) LK11205; GA AV ČR KAN200520801 Institutional support: RVO:61388963 ; RVO:68378050 Keywords : 5'(3')-nucleotidase * structure * inhibition * cdN * mdN * nucleoside * SAR * phosphonic acid Subject RIV: CC - Organic Chemistry Impact factor: 3.562, year: 2014

  8. Synthesis and in vitro growth inhibitory activity of novel silyl- and trityl-modified nucleosides

    CSIR Research Space (South Africa)

    Panayides, Jenny-Lee

    2016-06-01

    Full Text Available & Medicinal Chemistry 24 (2016) 2716–2724 Synthesis and in vitro growth inhibitory activity of novel silyl- and trityl-modified nucleosides Jenny-Lee Panayides a,b, Véronique Mathieu c, Laetitia Moreno Y. Banuls c, Helen Apostolellis d, Nurit Dahan...-Farkas d, Hajierah Davids d,e , Leonie Harmse d , M. E. Christine Rey f , Ivan R. Green g, Stephen C. Pelly g, Robert Kiss c, Alexander Kornienko h, Willem A. L. van Otterlo a,g,⇑ a Molecular Sciences Institute, School of Chemistry, University...

  9. Molecular Mechanism of Distinct Salt-Dependent Enzyme Activity of Two Halophilic Nucleoside Diphosphate Kinases

    OpenAIRE

    Yamamura, Akihiro; Ichimura, Takefumi; Kamekura, Masahiro; Mizuki, Toru; Usami, Ron; Makino, Tsukasa; Ohtsuka, Jun; Miyazono, Ken-ichi; Okai, Masahiko; Nagata, Koji; Tanokura, Masaru

    2009-01-01

    Nucleoside diphosphate kinases from haloarchaea Haloarcula quadrata (NDK-q) and H. sinaiiensis (NDK-s) are identical except for one out of 154 residues, i.e., Arg31 in NDK-q and Cys31 in NDK-s. However, the salt-dependent activity profiles of NDK-q and NDK-s are quite different: the optimal NaCl concentrations of NDK-q and NDK-s are 1 M and 2 M, respectively. We analyzed the relationships of the secondary, tertiary, and quaternary structures and NDK activity of these NDKs at various salt conc...

  10. Ethenoguanines undergo glycosylation by nucleoside 2'-deoxyribosyltransferases at non-natural sites.

    Directory of Open Access Journals (Sweden)

    Wenjie Ye

    Full Text Available Deoxyribosyl transferases and functionally related purine nucleoside phosphorylases are used extensively for synthesis of non-natural deoxynucleosides as pharmaceuticals or standards for characterizing and quantitating DNA adducts. Hence exploring the conformational tolerance of the active sites of these enzymes is of considerable practical interest. We have determined the crystal structure at 2.1 Å resolution of Lactobacillus helveticus purine deoxyribosyl transferase (PDT with the tricyclic purine 8,9-dihydro-9-oxoimidazo[2,1-b]purine (N2,3-ethenoguanine at the active site. The active site electron density map was compatible with four orientations, two consistent with sites for deoxyribosylation and two appearing to be unproductive. In accord with the crystal structure, Lactobacillus helveticus PDT glycosylates the 8,9-dihydro-9-oxoimidazo[2,1-b]purine at N7 and N1, with a marked preference for N7. The activity of Lactobacillus helveticus PDT was compared with that of the nucleoside 2'-deoxyribosyltransferase enzymes (DRT Type II from Lactobacillus leichmannii and Lactobacillus fermentum, which were somewhat more effective in the deoxyribosylation than Lactobacillus helveticus PDT, glycosylating the substrate with product profiles dependent on the pH of the incubation. The purine nucleoside phosphorylase of Escherichia coli, also commonly used in ribosylation of non-natural bases, was an order of magnitude less efficient than the transferase enzymes. Modeling based on published active-site structures as templates suggests that in all cases, an active site Phe is critical in orienting the molecular plane of the purine derivative. Adventitious hydrogen bonding with additional active site residues appears to result in presentation of multiple nucleophilic sites on the periphery of the acceptor base for ribosylation to give a distribution of nucleosides. Chemical glycosylation of O9-benzylated 8,9-dihydro-9-oxoimidazo[2,1-b]purine also resulted

  11. Cloning of human purine-nucleoside phosphorylase cDNA sequences by complementation in Escherichia coli.

    OpenAIRE

    Goddard, J M; Caput, D; Williams, S R; Martin, D W

    1983-01-01

    We have obtained cDNA clones that contain the entire coding region of the human purine-nucleoside phosphorylase (PNP; EC 2.4.2.1) mRNA. The cDNA sequences were generated by reverse transcription of PNP-enriched mRNA obtained by immunoadsorption of HeLa cell polyribosomes with monospecific antibody to human PNP. cDNA molecules that were close in length to PNP mRNA were separated by agarose gel electrophoresis and inserted into the Pst I site of the plasmid pBR322. Plasmid DNA from the pooled c...

  12. Focus on Chirality of HIV-1 Non-Nucleoside Reverse Transcriptase Inhibitors

    Directory of Open Access Journals (Sweden)

    Valeria Famiglini

    2016-02-01

    Full Text Available Chiral HIV-1 non-nucleoside reverse transcriptase inhibitors (NNRTIs are of great interest since one enantiomer is often more potent than the corresponding counterpart against the HIV-1 wild type (WT and the HIV-1 drug resistant mutant strains. This review exemplifies the various studies made to investigate the effect of chirality on the antiretroviral activity of top HIV-1 NNRTI compounds, such as nevirapine (NVP, efavirenz (EFV, alkynyl- and alkenylquinazolinone DuPont compounds (DPC, diarylpyrimidine (DAPY, dihydroalkyloxybenzyloxopyrimidine (DABO, phenethylthiazolylthiourea (PETT, indolylarylsulfone (IAS, arylphosphoindole (API and trifluoromethylated indole (TFMI The chiral separation, the enantiosynthesis, along with the biological properties of these HIV-1 NNRTIs, are discussed.

  13. Human cytomegalovirus resistance to deoxyribosylindole nucleosides maps to a transversion mutation in the terminase subunit-encoding gene UL89.

    Science.gov (United States)

    Gentry, Brian G; Phan, Quang; Hall, Ellie D; Breitenbach, Julie M; Borysko, Katherine Z; Kamil, Jeremy P; Townsend, Leroy B; Drach, John C

    2015-01-01

    Human cytomegalovirus (HCMV) infection can cause severe illnesses, including encephalopathy and mental retardation, in immunocompromised and immunologically immature patients. Current pharmacotherapies for treating systemic HCMV infections include ganciclovir, cidofovir, and foscarnet. However, long-term administration of these agents can result in serious adverse effects (myelosuppression and/or nephrotoxicity) and the development of viral strains with reduced susceptibility to drugs. The deoxyribosylindole (indole) nucleosides demonstrate a 20-fold greater activity in vitro (the drug concentration at which 50% of the number of plaques was reduced with the presence of drug compared to the number in the absence of drug [EC50] = 0.34 μM) than ganciclovir (EC50 = 7.4 μM) without any observed increase in cytotoxicity. Based on structural similarity to the benzimidazole nucleosides, we hypothesize that the indole nucleosides target the HCMV terminase, an enzyme responsible for packaging viral DNA into capsids and cleaving the DNA into genome-length units. To test this hypothesis, an indole nucleoside-resistant HCMV strain was isolated, the open reading frames of the genes that encode the viral terminase were sequenced, and a G766C mutation in exon 1 of UL89 was identified; this mutation resulted in an E256Q change in the amino acid sequence of the corresponding protein. An HCMV wild-type strain, engineered with this mutation to confirm resistance, demonstrated an 18-fold decrease in susceptibility to the indole nucleosides (EC50 = 3.1 ± 0.7 μM) compared to that of wild-type virus (EC50 = 0.17 ± 0.04 μM). Interestingly, this mutation did not confer resistance to the benzimidazole nucleosides (EC50 for wild-type HCMV = 0.25 ± 0.04 μM, EC50 for HCMV pUL89 E256Q = 0.23 ± 0.04 μM). We conclude, therefore, that the G766C mutation that results in the E256Q substitution is unique for indole nucleoside resistance and distinct from previously discovered substitutions

  14. Determination of the Main Nucleosides and Nucleobases in Natural and Cultured Ophiocordyceps xuefengensis

    Directory of Open Access Journals (Sweden)

    Juan Zou

    2017-09-01

    Full Text Available Ophiocordyceps xuefengensis, a recently described species of Ophiocordyceps that is associated with the larvae of Phassus nodus (Hepialidae in the living root or trunk of the medicinal plant Clerodendrum cyrtophyllum, is the largest known Cordyceps species and is recognized as a desirable alternative for natural Ophiocordyceps sinensis. This study investigated the main nucleosides and nucleobases in natural and cultured Ophiocordyceps xuefengensis. The contents of the nucleosides and nucleobases in the natural and cultured samples were determined by reverse phase HPLC. The highest concentration of adenosine was found in the natural fruit body and the cultured stroma, with almost no adenosine in the cadaver of Phassus nodus. The contents of adenine, guanosine, uridine and uracil in the cultured mycelium were significantly higher than those in the natural sample. Inosine was only detected in the natural samples. Thymidine and 2-deoxyadenosine were only found in the cadaver of Phassus nodus. Cordycepin was not detected in the five samples examined. These results suggested that the cultured mycelium and cultured stroma of Ophiocordyceps xuefengensis might be a promising substitute for natural O. xuefengensis.

  15. Modeling of Plasmodium falciparum Telomerase Reverse Transcriptase Ternary Complex: Repurposing of Nucleoside Analog Inhibitors.

    Science.gov (United States)

    Mohanty, Pallavi; Gupta, Akanksha; Bhatnagar, Sonika

    2015-12-01

    The Plasmodium falciparum telomerase reverse transcriptase (PfTERT) is a ribonucleoprotein that assists the maintenance of the telomeric ends of chromosomes by reverse transcription of its own RNA subunit. It represents an attractive therapeutic target for eradication of the plasmodial parasite at the asexual liver stage. Automated modeling using MUSTER and knowledge-based techniques were used to obtain a three-dimensional model of the active site of reverse transcriptase domain of PfTERT, which is responsible for catalyzing the addition of incoming dNTPs to the growing DNA strand in presence of divalent magnesium ions. Further, the ternary complex of the active site of PfTERT bound to a DNA-RNA duplex was also modeled using Haddock server and represents the functional form of the enzyme. Initially, established nucleoside analog inhibitors of PfTERT, AZTTP, and ddGTP were docked in the modeled binding site of the PfTERT ternary complex using AutoDock v4.2. Subsequently, docking studies were carried out with 14 approved nucleoside analog inhibitors. Docking studies predicted that floxuridine, gemcitabine, stavudine, and vidarabine have high affinity for the PfTERT ternary complex. Further analysis on the basis of known side effects led us to propose repositioning of vidarabine as a suitable drug candidate for inhibition of PfTERT.

  16. Investigation and conformational analysis of fluorinated nucleoside antibiotics targeting siderophore biosynthesis.

    Science.gov (United States)

    Dawadi, Surendra; Viswanathan, Kishore; Boshoff, Helena I; Barry, Clifton E; Aldrich, Courtney C

    2015-05-15

    Antibiotic resistance represents one of the greatest threats to public health. The adenylation inhibitor 5'-O-[N-(salicyl)sulfamoyl]adenosine (SAL-AMS) is the archetype for a new class of nucleoside antibiotics that target iron acquisition in pathogenic microorganisms and is especially effective against Mycobacterium tuberculosis, the causative agent of tuberculosis. Strategic incorporation of fluorine at the 2' and 3' positions of the nucleoside was performed by direct fluorination to enhance activity and improve drug disposition properties. The resulting SAL-AMS analogues were comprehensively assessed for biochemical potency, whole-cell antitubercular activity, and in vivo pharmacokinetic parameters. Conformational analysis suggested a strong preference of fluorinated sugar rings for either a 2'-endo, 3'-exo (South), or a 3'-endo,2'-exo (North) conformation. The structure-activity relationships revealed a strong conformational bias for the C3'-endo conformation to maintain potent biochemical and whole-cell activity, whereas improved pharmacokinetic properties were associated with the C2'-endo conformation.

  17. Synthesis and Biological Evaluation of Triazolyl 13α-Estrone–Nucleoside Bioconjugates

    Directory of Open Access Journals (Sweden)

    Brigitta Bodnár

    2016-09-01

    Full Text Available 2′-Deoxynucleoside conjugates of 13α-estrone were synthesized by applying the copper-catalyzed alkyne–azide click reaction (CuAAC. For the introduction of the azido group the 5′-position of the nucleosides and a propargyl ether functional group on the 3-hydroxy group of 13α-estrone were chosen. The best yields were realized in our hands when the 3′-hydroxy groups of the nucleosides were protected by acetyl groups and the 5′-hydroxy groups were modified by the tosyl–azide exchange method. The commonly used conditions for click reaction between the protected-5′-azidonucleosides and the steroid alkyne was slightly modified by using 1.5 equivalent of Cu(I catalyst. All the prepared conjugates were evaluated in vitro by means of MTT assays for antiproliferative activity against a panel of human adherent cell lines (HeLa, MCF-7 and A2780 and the potential inhibitory activity of the new conjugates on human 17β-hydroxysteroid dehydrogenase 1 (17β-HSD1 was investigated via in vitro radiosubstrate incubation. Some protected conjugates displayed moderate antiproliferative properties against a panel of human adherent cancer cell lines (the protected cytidine conjugate proved to be the most potent with IC50 value of 9 μM. The thymidine conjugate displayed considerable 17β-HSD1 inhibitory activity (IC50 = 19 μM.

  18. Acanthamoeba polyphaga mimivirus NDK: preliminary crystallographic analysis of the first viral nucleoside diphosphate kinase

    International Nuclear Information System (INIS)

    Jeudy, Sandra; Coutard, Bruno; Lebrun, Régine; Abergel, Chantal

    2005-01-01

    A. polyphaga mimivirus, the largest known double-stranded DNA virus, is the first virus to exhibit a nucleoside diphosphate kinase gene. The expression and crystallization of the viral NDK are reported. The complete sequence of the largest known double-stranded DNA virus, Acanthamoeba polyphaga mimivirus, has recently been determined [Raoult et al. (2004 ▶), Science, 306, 1344–1350] and revealed numerous genes not expected to be found in a virus. A comprehensive structural and functional study of these gene products was initiated [Abergel et al. (2005 ▶), Acta Cryst. F61, 212–215] both to better understand their role in the virus physiology and to obtain some clues to the origin of DNA viruses. Here, the preliminary crystallographic analysis of the viral nucleoside diphosphate kinase protein is reported. The crystal belongs to the cubic space group P2 1 3, with unit-cell parameter 99.425 Å. The self-rotation function confirms that there are two monomers per asymmetric unit related by a twofold non-crystallographic axis and that the unit cell thus contains four biological entities

  19. Acanthamoeba polyphaga mimivirus NDK: preliminary crystallographic analysis of the first viral nucleoside diphosphate kinase

    Energy Technology Data Exchange (ETDEWEB)

    Jeudy, Sandra [Information Génomique et Structurale, CNRS UPR 2589, 31 Chemin Joseph Aiguier, 13402 Marseille CEDEX 20 (France); Coutard, Bruno [Architecture et Fonction des Macromolecules Biologiques, CNRS UMR 6098, 31 Chemin Joseph Aiguier, 13402 Marseille CEDEX 20 (France); Lebrun, Régine [IBSM, 31 Chemin Joseph Aiguier, 13402 Marseille CEDEX 20 (France); Abergel, Chantal, E-mail: chantal.abergel@igs.cnrs-mrs.fr [Information Génomique et Structurale, CNRS UPR 2589, 31 Chemin Joseph Aiguier, 13402 Marseille CEDEX 20 (France)

    2005-06-01

    A. polyphaga mimivirus, the largest known double-stranded DNA virus, is the first virus to exhibit a nucleoside diphosphate kinase gene. The expression and crystallization of the viral NDK are reported. The complete sequence of the largest known double-stranded DNA virus, Acanthamoeba polyphaga mimivirus, has recently been determined [Raoult et al. (2004 ▶), Science, 306, 1344–1350] and revealed numerous genes not expected to be found in a virus. A comprehensive structural and functional study of these gene products was initiated [Abergel et al. (2005 ▶), Acta Cryst. F61, 212–215] both to better understand their role in the virus physiology and to obtain some clues to the origin of DNA viruses. Here, the preliminary crystallographic analysis of the viral nucleoside diphosphate kinase protein is reported. The crystal belongs to the cubic space group P2{sub 1}3, with unit-cell parameter 99.425 Å. The self-rotation function confirms that there are two monomers per asymmetric unit related by a twofold non-crystallographic axis and that the unit cell thus contains four biological entities.

  20. Magnetic resonance imaging based functional imaging in paediatric oncology.

    Science.gov (United States)

    Manias, Karen A; Gill, Simrandip K; MacPherson, Lesley; Foster, Katharine; Oates, Adam; Peet, Andrew C

    2017-02-01

    Imaging is central to management of solid tumours in children. Conventional magnetic resonance imaging (MRI) is the standard imaging modality for tumours of the central nervous system (CNS) and limbs and is increasingly used in the abdomen. It provides excellent structural detail, but imparts limited information about tumour type, aggressiveness, metastatic potential or early treatment response. MRI based functional imaging techniques, such as magnetic resonance spectroscopy, diffusion and perfusion weighted imaging, probe tissue properties to provide clinically important information about metabolites, structure and blood flow. This review describes the role of and evidence behind these functional imaging techniques in paediatric oncology and implications for integrating them into routine clinical practice. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Photon Harvesting in Conjugated Polymer-Based Functional Nanoparticles.

    Science.gov (United States)

    Jana, Bikash; Ghosh, Arnab; Patra, Amitava

    2017-09-21

    The design of new generation light-harvesting systems based on conjugated polymer nanoparticles (PNPs) is an emerging field of research to convert solar energy into renewable energy. In this Perspective, we focus on the understanding of the light harvesting processes like exciton dynamics, energy transfer, antenna effect, charge carrier dynamics, and other related processes of conjugated polymer-based functional nanomaterials. Spectroscopic investigations unveil the rotational dynamics of the dye molecules inside of PNPs and exciton dynamics of the self-assembled structures. A detailed understanding of the cascade energy transfer for white light and singlet oxygen generation in multiple fluorophores containing a PNP system by time-resolved spectroscopy is highlighted. Finally, ultrafast spectroscopic investigations provide direct insight into the impacts of electron and hole transfer at the interface in the hybrid materials for photocatalysis and photocurrent generation to construct efficient light-harvesting systems.

  2. Acyclic Nucleoside Phosphonates Containing 9-Deazahypoxanthine and a Five-Membered Heterocycle as Selective Inhibitors of Plasmodial 6-Oxopurine Phosphoribosyltransferases

    Czech Academy of Sciences Publication Activity Database

    Kaiser, Martin Maxmilian; Baszczyňski, Ondřej; Hocková, Dana; Poštová Slavětínská, Lenka; Dračínský, Martin; Keough, D. T.; Guddat, L. W.; Janeba, Zlatko

    2017-01-01

    Roč. 12, č. 14 (2017), s. 1133-1141 ISSN 1860-7179 R&D Projects: GA ČR(CZ) GA16-06049S Institutional support: RVO:61388963 Keywords : inhibitors * nucleosides * malaria * phosphonates * purine salvage Subject RIV: CC - Organic Chemistry OBOR OECD: Organic chemistry Impact factor: 3.225, year: 2016

  3. Cytostatic 6-arylpurine nucleosides. 6. SAR in anti-HCV and cytostatic activity of extended series of 6-hetarylpurine ribonucleosides

    Czech Academy of Sciences Publication Activity Database

    Hocek, Michal; Nauš, Petr; Pohl, Radek; Votruba, Ivan; Furman, P. A.; Tharnish, P. M.; Otto, M. J.

    2005-01-01

    Roč. 48, č. 18 (2005), s. 5869-5873 ISSN 0022-2623 R&D Projects: GA MŠk(CZ) 1M0508 Institutional research plan: CEZ:AV0Z40550506 Keywords : purines * nucleosides * antiviral activity Subject RIV: CC - Organic Chemistry Impact factor: 4.926, year: 2005

  4. Syntheses of N3-substituted thymine acyclic nucleoside phosphonates and a comparison of their inhibitory effect towards thymidine phosphorylase

    Czech Academy of Sciences Publication Activity Database

    Pomeisl, Karel; Holý, Antonín; Votruba, Ivan; Pohl, Radek

    2008-01-01

    Roč. 18, č. 4 (2008), s. 1364-1367 ISSN 0960-894X Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * thymidine phosphorylase * fluorination * pyrimidine Subject RIV: CC - Organic Chemistry Impact factor: 2.531, year: 2008

  5. Hypoxanthine production by ischemic heart demonstrated by high pressure liquid chromatography of blood purine nucleosides and oxypurines

    NARCIS (Netherlands)

    E. Harmsen; J.W. de Jong (Jan Willem); P.W.J.C. Serruys (Patrick)

    1981-01-01

    textabstractAn isocratic high pressure liquid chromatographic system was developed for the estimation of purine nucleosides and oxypurines in blood. Use was made of a reversed-phase column. Nucleotides derived from erythrocytes affected the separation; these compounds were removed with A12O3. The

  6. Radioimmunoassays for the modified nucleosides N[9-(beta-D-ribofuranosyl)purin-6-ylcarbamoyl]-L-threonine and 2-methylthioadenosine.

    Science.gov (United States)

    Vold, B S

    1979-01-01

    Radioimmunoassays were established for the modified nucleosides N-[9-(beta-D-ribofuranosyl)purin-6-ylcarbamoyl]-L-threonine, t6A, and 2-methylthioadenosine, ms2A. The assays depended on the production of antisera specific for t6A and ms2A that have not been previously reported. The nitrocellulose membrane filtration and saturated ammonium sulfate RIA techniques were compared for efficiency. Various radioactive antigens were employed to establish which type of antigen would give the best binding. The tritium post-labeling procedure of Randerath and Randerath was used to obtain labeled nucleosides of high enough specific activity to be useful for RIAs when the labeled nucleoside was not available commerically. The specificity of the antibodies toward nucleosides and purified tRNAs is reported. Although the titer of the t6A antiserum was low, the specificity was very sharp. An interesting finding was that threonine, a major structural component of the side-chain modification of t6A, was completely infective as an inhibitor. PMID:493139

  7. Synthesis and Evaluation of Novel Acyclic Nucleoside Phosphonates as Inhibitors of Plasmodium falciparum and Human 6-Oxopurine Phosphoribosyltransferases

    Czech Academy of Sciences Publication Activity Database

    Kaiser, Martin Maxmilian; Hocková, Dana; Wang, T. H.; Dračínský, Martin; Poštová Slavětínská, Lenka; Procházková, Eliška; Edstein, M. D.; Chavchich, M.; Keough, D. T.; Guddat, L. W.; Janeba, Zlatko

    2015-01-01

    Roč. 10, č. 10 (2015), s. 1707-1723 ISSN 1860-7179 R&D Projects: GA MV VG20102015046; GA ČR GAP207/11/0108 Institutional support: RVO:61388963 Keywords : 6-oxopurine * acyclic nucleoside phosphonates * phosphoribosyltransferases * malaria * phosphoramidates Subject RIV: CC - Organic Chemistry Impact factor: 2.980, year: 2015

  8. Liquid chromatography-tandem mass spectrometric assay for the nucleoside reverse transcriptase inhibitor emtricitabine in human plasma

    NARCIS (Netherlands)

    Sparidans, Rolf W.; Prins, Jan M.; Schellens, Jan H. M.; Beijnen, Jos H.

    2007-01-01

    A liquid chromatography-tandem mass spectrometric assay for the determination of the antiretroviral nucleoside emtricitabine in human plasma was developed and validated using a simple sample pre-treatment procedure. After addition of 5'-deoxy-5-fluorocytidine as the internal standard and protein

  9. Inhibition of nucleoside transport and synergistic potentiation of methotrexate cytotoxicity by cimicifugoside, a triterpenoid from Cimicifuga simplex.

    Science.gov (United States)

    Yawata, Ayako; Matsuhashi, Yuko; Kato, Hanako; Uemura, Keiko; Kusano, Genjiro; Ito, Junko; Chikuma, Toshiyuki; Hojo, Hiroshi

    2009-11-05

    Cimicifugoside, a triterpenoid isolated from Cimicifuga simplex, which has been used as a traditional Chinese medicine due to its anti-inflammatory, analgesic or anti-pyretic action, was examined for inhibition of nucleoside transport and synergistic potentiation of methotrexate cytotoxicity. Cimicifugoside inhibited uptake of uridine, thymidine and adenosine in human leukemia U937 cells with the low nanomolar IC(50) values, but did not affect that of uracil, leucine or 2-deoxyglucose at cimicifugenin (aglycon of cimicifugoside)>bugbanoside B>cimicifugenin A, O-methyl cimicifugenin and bugbanoside A. Cimicifugoside had less affinity for the binding site of nitrobenzylthioinosine (typical high-affinity inhibitor of equilibrative nucleoside transporter-1) in U937 cells, K562 cells and human erythrocyte membranes compared with the prototype nucleoside transport inhibitor dipyridamole. Cimicifugoside markedly potentiated methotrexate cytotoxicity in a culture of U937 cells and human carcinoma KB cells. Potentiation of methotrexate cytotoxicity by cimicifugoside analogs in U937 cells was in proportion to their inhibitory activity against uridine uptake. The present study demonstrates that cimicifugoside is a novel specific nucleoside transport inhibitor that displays synergistic potentiation of methotrexate cytotoxicity.

  10. Targeted Delivery of Deoxycytidine Kinase to Her2-Positive Cells Enhances the Efficacy of the Nucleoside Analog Fludarabine.

    Directory of Open Access Journals (Sweden)

    Sujatha P Koduvayur

    Full Text Available Cytotoxic drugs, such as nucleoside analogs and toxins, commonly suffer from off-target effects. One approach to mitigate this problem is to deliver the cytotoxic drug selectively to the intended site. While for toxins this can be achieved by conjugating the cell-killing moiety to a targeting moiety, it is not an option for nucleoside analogs, which rely on intracellular enzymes to convert them to their active triphosphorylated form. To overcome this limitation, and achieve site-targeted activation of nucleoside analogs, we fused the coding region of a prodrug-activating enzyme, deoxycytidine kinase (dCK, to affinity reagents that bind to the Her2 cell surface protein. We evaluated dCK fusions to an anti-Her2 affibody and Designed Ankyrin Repeat Protein (DARPin for their ability to kill cancer cells by promoting the activation of the nucleoside analog fludarabine. Cell staining and flow cytometry experiments with three Her2 positive cancer cell lines (BT-474-JB, JIMT-1 and SK-OV-3 indicate dCK fusions binding and cellular internalization. In contrast, these reagents bind only weakly to the Her2 negative cell line, MCF-7. Cell proliferation assays indicate that SK-OV-3 and BT-474-JB cell lines exhibit significantly reduced proliferation rates when treated with targeting-module fused dCK and fludarabine, compared to fludarabine alone. These findings demonstrate that we have succeeded in delivering active dCK into the Her2-positive cells, thereby increasing the activation of fludarabine, which ultimately reduces the dose of nucleoside analog needed for cell killing. This strategy may help establish the therapeutic index required to differentiate between healthy tissues and cancer cells.

  11. A System for Dosage-Based Functional Genomics in Poplar.

    Science.gov (United States)

    Henry, Isabelle M; Zinkgraf, Matthew S; Groover, Andrew T; Comai, Luca

    2015-09-01

    Altering gene dosage through variation in gene copy number is a powerful approach to addressing questions regarding gene regulation, quantitative trait loci, and heterosis, but one that is not easily applied to sexually transmitted species. Elite poplar (Populus spp) varieties are created through interspecific hybridization, followed by clonal propagation. Altered gene dosage relationships are believed to contribute to hybrid performance. Clonal propagation allows for replication and maintenance of meiotically unstable ploidy or structural variants and provides an alternative approach to investigating gene dosage effects not possible in sexually propagated species. Here, we built a genome-wide structural variation system for dosage-based functional genomics and breeding of poplar. We pollinated Populus deltoides with gamma-irradiated Populus nigra pollen to produce >500 F1 seedlings containing dosage lesions in the form of deletions and insertions of chromosomal segments (indel mutations). Using high-precision dosage analysis, we detected indel mutations in ∼55% of the progeny. These indels varied in length, position, and number per individual, cumulatively tiling >99% of the genome, with an average of 10 indels per gene. Combined with future phenotype and transcriptome data, this population will provide an excellent resource for creating and characterizing dosage-based variation in poplar, including the contribution of dosage to quantitative traits and heterosis. © 2015 American Society of Plant Biologists. All rights reserved.

  12. Abacavir-based triple nucleoside regimens for maintenance therapy in patients with HIV.

    Science.gov (United States)

    Cruciani, Mario; Mengoli, Carlo; Serpelloni, Giovanni; Parisi, Saverio G; Malena, Marina; Bosco, Oliviero

    2013-06-05

    Regimen simplification can be defined as a change in established effective therapy to reduce pill burden and dosing frequency, to enhance tolerability, or to decrease specific food and fluid requirements. Many patients on suppressive antiretroviral therapy may be considered candidates for a simplification strategy and, among them, those who have achieved virologic suppression. Several clinical trials have evaluated the efficacy of triple nucleoside combination as a simplification therapy in patients who achieved virologic suppression The aim of this review is to combine randomised, controlled trials to examine whether in patients with undetectable viraemia on a Protease inhibitor (PI) based regimen simplification treatment with abacavir (ABC)-based triple-nucleoside combinations has similar rates of efficacy and tolerability compared with a PI regimen or simplification with a NNRTIs (efavirenz-EFV- or nevirapine-NVP) containing regimen. Studies were included if they had at least two of the three interventions, including one 3NRTI arm. Electronic databases and conference proceedings were searched (1996-2012) with relevant search terms without limits to language. Randomised controlled trials (RCTs) only are included in this review. Patients population is represented by HIV-infected adult patients treated with a PI-containing regimen (PI or boosted PI),  with undetectable viral load. Patients on a PI-containing regimen had three possibilities: continue the PI regimen or switch to a simplification maintenance regimen, including switch to a NNRTI (EFV or NVP) containing regimen, or switch to a triple-NRTI regimen (ABC-zidovudine-lamivudine) The primary outcomes were: proportion of patients discontinuing or switching antiretroviral therapy due to virologic failure or to adverse events; death (all cause) and AIDS defining illness; occurrence of myocardial infarction and cardiovascular disease. Secondary outcomes  were: proportion of patients maintaining an undetectable

  13. Low liver stiffness among cirrhotic patients with hepatitis B after prolonged treatment with nucleoside analogs

    DEFF Research Database (Denmark)

    Andersen, Ellen Sloth; Weiland, Ola; Leutscher, Peter

    2011-01-01

    Abstract Objective. Case reports and short-term clinical trials have suggested that treatment for chronic hepatitis B (CHB) may lead to improvement of cirrhosis. The aim of the present study was to measure liver stiffness in patients diagnosed with advanced fibrosis or cirrhosis prior to prolonged...... treatment with nucleoside or nucleotide analogs (NUCs) for CHB. Materials and methods. Patients with CHB and advanced fibrosis or cirrhosis prior to treatment with NUCs for at least 1 year were offered inclusion in the study. We measured liver stiffness using transient elastography (TE) at follow-up. TE cut...... duration was 50.5 months. Among patients with cirrhosis prior to treatment, 26 (49%) had liver stiffness below 11.0 kPa at follow-up, suggesting regression of cirrhosis. Among patients with advanced fibrosis (F3) prior to treatment, 10 (77%) had liver stiffness below 8.1 kPa after treatment, suggesting...

  14. Structure-Activity Relationships of Acyclic Selenopurine Nucleosides as Antiviral Agents

    Directory of Open Access Journals (Sweden)

    Pramod K. Sahu

    2017-07-01

    Full Text Available A series of acyclic selenopurine nucleosides 3a–f and 4a–g were synthesized based on the bioisosteric rationale between oxygen and selenium, and then evaluated for antiviral activity. Among the compounds tested, seleno-acyclovir (4a exhibited the most potent anti-herpes simplex virus (HSV-1 (EC50 = 1.47 µM and HSV-2 (EC50 = 6.34 µM activities without cytotoxicity up to 100 µM, while 2,6-diaminopurine derivatives 4e–g exhibited significant anti-human cytomegalovirus (HCMV activity, which is slightly more potent than the guanine derivative 4d, indicating that they might act as prodrugs of seleno-ganciclovir (4d.

  15. [Fluorinated analogs of nucleosides and fluorinated tracers of gene expression for positron emission tomography].

    Science.gov (United States)

    Couturier, Olivier; Chatal, Jean-François; Hustinx, Roland

    2004-09-01

    18F-FDG is currently the only fluorinated tracer used in routine clinical positron emission tomography (PET). Fluorine 18 is considered as the ideal radioisotope for PET, thanks to a low positron energy, which not only limits the dose rate to the patients but also provides high-resolution images. Furthermore, the 110 min. physical half-life allows for high-yield radiosynthesis, transport from the production site to the imaging site, and imaging protocols that could span hours, which permits dynamic studies and assessing metabolic processes that may be fairly slow. Recently, synthesis of fluorinated tracers from prosthetic group precursors, which allows easier radiolabeling of biomolecules, has given a boost to the development of numerous fluorinated tracers. Given the wide availability of fluorine 18, such tracers may well develop into important routine tracers. This article is a review of the literature concerning fluorinated analogs of nucleosides and fluorinated radiotracers of gene expression recently developed and under investigation.

  16. Cytological localization of adenosine kinase, nucleoside phosphorylase-1, and esterase-10 genes on mouse chromosome 14

    International Nuclear Information System (INIS)

    Samuelson, L.C.; Farber, R.A.

    1985-01-01

    The authors have determined the regional locations on mouse chromosome 14 of the genes for mouse adenosine kinase (ADK), nucleoside phosphorylase- 1 (NP-1), and esterase-10 (ES-10) by analysis of rearranged mouse chromosomes in gamma-irradiated Chinese hamster X mouse hybrid cell lines. Irradiated clones were screened for expression of the murine forms of these enzymes; segregant clones that expressed only one or two of the three markers were karyotyped. The patterns of enzyme expression in these segregants were correlated with the presence of rearranged chromosomes. The Adk gene was localized to bands A2 to B, Np-1 to bands B to C1, and Es-10 to bands D2 to E2

  17. Nitrobenzylthioinosine (NBT), a nucleoside transport inhibitor, protects against Shiga toxin cytotoxicity in human microvascular endothelial cells.

    Science.gov (United States)

    Ohmi, K; Kiyokawa, N; Sekino, T; Suzuki, T; Mimori, K; Taguchi, T; Nakajima, H; Katagiri, Y U; Fujimoto, J; Nakao, H; Takeda, T

    2001-01-01

    Infections with Shiga toxin (Stx)-producing Escherichia coli (STEC) cause microvascular endothelial cell damage, resulting in hemorrhagic colitis and hemolytic uremic syndrome. The prevention of endothelial cell damage is therefore a crucial step in overcoming this disorder. Here, we report that nitrobenzylthioinosine (NBT), a nucleoside transport inhibitor, has a protective effect against the cytotoxicity of Stxs in human microvascular endothelial cells (HMVECs). The relative viability of cells treated with 1.5-15 pM of Stx1 was reduced to 10-20% of that without Stx1. However, the viability of cells treated with NBT (10-100 microM) remained higher than 80%, even in the presence of Stx1. NBT also protected against Stx1 cytotoxicity in sodium butyrate-treated hypersensitive HMVECs. The protective effect of NBT against Stx cytotoxicity may be due to the depletion of ATP in the cells, thereby inhibiting the entry of Stx1.

  18. Aquaporin 3 (AQP3) participates in the cytotoxic response to nucleoside-derived drugs

    International Nuclear Information System (INIS)

    Pérez-Torras, Sandra; Casado, F Javier; Pastor-Anglada, Marçal

    2012-01-01

    Nucleoside analogs used in the chemotherapy of solid tumors, such as the capecitabine catabolite 5 ′ -deoxy-5-fluorouridine (5 ′ -DFUR) trigger a transcriptomic response that involves the aquaglyceroporin aquaporin 3 along with other p53-dependent genes. Here, we examined whether up-regulation of aquaporin 3 (AQP3) mRNA in cancer cells treated with 5 ′ -DFUR represents a collateral transcriptomic effect of the drug, or conversely, AQP3 participates in the activity of genotoxic agents. The role of AQP3 in cell volume increase, cytotoxicity and cell cycle arrest was analyzed using loss-of-function approaches. 5 ′ -DFUR and gemcitabine, but not cisplatin, stimulated AQP3 expression and cell volume, which was partially and significantly blocked by knockdown of AQP3. Moreover, AQP3 siRNA significantly blocked other effects of nucleoside analogs, including G 1 /S cell cycle arrest, p21 and FAS up-regulation, and cell growth inhibition. Short incubations with 5-fluorouracil (5-FU) also induced AQP3 expression and increased cell volume, and the inhibition of AQP3 expression significantly blocked growth inhibition triggered by this drug. To further establish whether AQP3 induction is related to cell cycle arrest and apoptosis, cells were exposed to long incubations with escalating doses of 5-FU. AQP3 was highly up-regulated at doses associated with cell cycle arrest, whereas at doses promoting apoptosis induction of AQP3 mRNA expression was reduced. Based on the results, we propose that the aquaglyceroporin AQP3 is required for cytotoxic activity of 5’-DFUR and gemcitabine in the breast cancer cell line MCF7 and the colon adenocarcinoma cell line HT29, and is implicated in cell volume increase and cell cycle arrest

  19. Immuno-Northern Blotting: Detection of RNA Modifications by Using Antibodies against Modified Nucleosides.

    Science.gov (United States)

    Mishima, Eikan; Jinno, Daisuke; Akiyama, Yasutoshi; Itoh, Kunihiko; Nankumo, Shinnosuke; Shima, Hisato; Kikuchi, Koichi; Takeuchi, Yoichi; Elkordy, Alaa; Suzuki, Takehiro; Niizuma, Kuniyasu; Ito, Sadayoshi; Tomioka, Yoshihisa; Abe, Takaaki

    2015-01-01

    The biological roles of RNA modifications are still largely not understood. Thus, developing a method for detecting RNA modifications is important for further clarification. We developed a method for detecting RNA modifications called immuno-northern blotting (INB) analysis and herein introduce its various capabilities. This method involves the separation of RNAs using either polyacrylamide or agarose gel electrophoresis, followed by transfer onto a nylon membrane and subsequent immunoblotting using antibodies against modified nucleosides for the detection of specific modifications. We confirmed that INB with the antibodies for 1-methyladenosine (m1A), N6-methyladenosine (m6A), pseudouridine, and 5-methylcytidine (m5C) showed different modifications in a variety of RNAs from various species and organelles. INB with the anti-m5C antibody revealed that the antibody cross-reacted with another modification on DNA, suggesting the application of this method for characterization of the antibody for modified nucleosides. Additionally, using INB with the antibody for m1A, which is a highly specific modification in eukaryotic tRNA, we detected tRNA-derived fragments known as tiRNAs under the cellular stress response, suggesting the application for tracking target RNA containing specific modifications. INB with the anti-m6A antibody confirmed the demethylation of m6A by the specific demethylases fat mass and obesity-associated protein (FTO) and ALKBH5, suggesting its application for quantifying target modifications in separated RNAs. Furthermore, INB demonstrated that the knockdown of FTO and ALKBH5 increased the m6A modification in small RNAs as well as in mRNA. The INB method has high specificity, sensitivity, and quantitative capability, and it can be employed with conventional experimental apparatus. Therefore, this method would be useful for research on RNA modifications and metabolism.

  20. Immuno-Northern Blotting: Detection of RNA Modifications by Using Antibodies against Modified Nucleosides

    Science.gov (United States)

    Akiyama, Yasutoshi; Itoh, Kunihiko; Nankumo, Shinnosuke; Shima, Hisato; Kikuchi, Koichi; Takeuchi, Yoichi; Elkordy, Alaa; Suzuki, Takehiro; Niizuma, Kuniyasu; Ito, Sadayoshi; Tomioka, Yoshihisa; Abe, Takaaki

    2015-01-01

    The biological roles of RNA modifications are still largely not understood. Thus, developing a method for detecting RNA modifications is important for further clarification. We developed a method for detecting RNA modifications called immuno-northern blotting (INB) analysis and herein introduce its various capabilities. This method involves the separation of RNAs using either polyacrylamide or agarose gel electrophoresis, followed by transfer onto a nylon membrane and subsequent immunoblotting using antibodies against modified nucleosides for the detection of specific modifications. We confirmed that INB with the antibodies for 1-methyladenosine (m1A), N6-methyladenosine (m6A), pseudouridine, and 5-methylcytidine (m5C) showed different modifications in a variety of RNAs from various species and organelles. INB with the anti-m5C antibody revealed that the antibody cross-reacted with another modification on DNA, suggesting the application of this method for characterization of the antibody for modified nucleosides. Additionally, using INB with the antibody for m1A, which is a highly specific modification in eukaryotic tRNA, we detected tRNA-derived fragments known as tiRNAs under the cellular stress response, suggesting the application for tracking target RNA containing specific modifications. INB with the anti-m6A antibody confirmed the demethylation of m6A by the specific demethylases fat mass and obesity-associated protein (FTO) and ALKBH5, suggesting its application for quantifying target modifications in separated RNAs. Furthermore, INB demonstrated that the knockdown of FTO and ALKBH5 increased the m6A modification in small RNAs as well as in mRNA. The INB method has high specificity, sensitivity, and quantitative capability, and it can be employed with conventional experimental apparatus. Therefore, this method would be useful for research on RNA modifications and metabolism. PMID:26606401

  1. In vivo phenotypic characterisation of nucleoside label-retaining cells in mouse periosteum

    Directory of Open Access Journals (Sweden)

    HM Cherry

    2014-03-01

    Full Text Available Periosteum is known to contain cells that, after isolation and culture-expansion, display properties of mesenchymal stromal/stem cells (MSCs. However, the equivalent cells have not been identified in situ mainly due to the lack of specific markers. Postnatally, stem cells are slow-cycling, long-term nucleoside-label-retaining cells. This study aimed to identify and characterise label-retaining cells in mouse periosteum in vivo. Mice received iodo-deoxy-uridine (IdU via the drinking water for 30 days, followed by a 40-day washout period. IdU+ cells were identified by immunostaining in conjunction with MSC and lineage markers. IdU-labelled cells were detected throughout the periosteum with no apparent focal concentration, and were negative for the endothelial marker von Willebrand factor and the pan-haematopoietic marker CD45. Subsets of IdU+ cells were positive for the mesenchymal/stromal markers vimentin and cadherin-11. IdU+ cells expressed stem cell antigen-1, CD44, CD73, CD105, platelet-derived growth factor receptor-α and p75, thereby displaying an MSC-like phonotype. Co-localisation was not detectable between IdU and the pericyte markers CD146, alpha smooth muscle actin or NG2, nor did IdU co-localise with β-galactosidase in a transgenic mouse expressing this reporter gene in pericytes and smooth muscle cells. Subsets of IdU+ cells expressed the osteoblast-lineage markers Runx2 and osteocalcin. The IdU+ cells expressing osteocalcin were lining the bone and were negative for the MSC marker p75. In conclusion, mouse periosteum contains nucleoside-label-retaining cells with a phenotype compatible with MSCs that are distinct from pericytes and osteoblasts. Future studies characterising the MSC niche in vivo could reveal novel therapeutic targets for promoting bone regeneration/repair.

  2. The crystal structure of the hexameric purine nucleoside phosphorylase from Bacillus subtilis in complex with adenosine

    International Nuclear Information System (INIS)

    Giuseppe, P.O.; Meza, A.N.; Martins, N.H.; Santos, C.R.; Murakami, M.T.

    2012-01-01

    Full text: Purine nucleoside phosphorylases (PNPs) play a key role in the purine-salvage pathway in both prokaryotes and eukaryotes. Its ribosyltransferase activity is of great biotechnological interest due to potential application in the synthesis of nucleoside analogues used in the treatment of antiviral infections and in anticancer chemotherapy. Trimeric PNPs are found mainly in vertebrates and are specific for 6-oxo-purines whereas hexameric PNPs are prevalent in prokaryotes and exhibit a broad range of substrates including 6-oxo and 6-amino purines. BsPNP233, the hexameric PNP from B. subtilis, is able to catalyze the bioconversion of ribavirin, an anti-viral drug, and is relatively thermostable, being a good target for industrial use. Here we report the crystal structures of BsPNP233 in the apo form and in complex with adenosine solved at 2.65 and 1.91 resolution, respectively. The apo and ligand-bound BsPNP233 subunits superposed with an overall r.m.s. deviation of 0.31 for all Cα atoms, which suggests that no major conformational changes occur upon substrate binding. Based on the crystal structure of BsPNP233 in complex with adenosine we have defined the active site residues implicated in binding the ribose (H4 * , R43 * , M64, R87, E178, M179, E180) and the nitrogenous base (S90, C91, G92, S202, V177, F159). These residues are highly conserved among the bacterial hexameric PNPs, suggesting they share the same mode of interaction with the substrates. This work will probably contribute to a better understanding of the molecular basis for the broad substrate specificity of hexameric PNPs and to projects aiming the rational design of PNPs for industrial purposes. (author)

  3. Immuno-Northern Blotting: Detection of RNA Modifications by Using Antibodies against Modified Nucleosides.

    Directory of Open Access Journals (Sweden)

    Eikan Mishima

    Full Text Available The biological roles of RNA modifications are still largely not understood. Thus, developing a method for detecting RNA modifications is important for further clarification. We developed a method for detecting RNA modifications called immuno-northern blotting (INB analysis and herein introduce its various capabilities. This method involves the separation of RNAs using either polyacrylamide or agarose gel electrophoresis, followed by transfer onto a nylon membrane and subsequent immunoblotting using antibodies against modified nucleosides for the detection of specific modifications. We confirmed that INB with the antibodies for 1-methyladenosine (m1A, N6-methyladenosine (m6A, pseudouridine, and 5-methylcytidine (m5C showed different modifications in a variety of RNAs from various species and organelles. INB with the anti-m5C antibody revealed that the antibody cross-reacted with another modification on DNA, suggesting the application of this method for characterization of the antibody for modified nucleosides. Additionally, using INB with the antibody for m1A, which is a highly specific modification in eukaryotic tRNA, we detected tRNA-derived fragments known as tiRNAs under the cellular stress response, suggesting the application for tracking target RNA containing specific modifications. INB with the anti-m6A antibody confirmed the demethylation of m6A by the specific demethylases fat mass and obesity-associated protein (FTO and ALKBH5, suggesting its application for quantifying target modifications in separated RNAs. Furthermore, INB demonstrated that the knockdown of FTO and ALKBH5 increased the m6A modification in small RNAs as well as in mRNA. The INB method has high specificity, sensitivity, and quantitative capability, and it can be employed with conventional experimental apparatus. Therefore, this method would be useful for research on RNA modifications and metabolism.

  4. The thiopurine nucleoside analogue 6-methylmercaptopurine riboside (6MMPr) effectively blocks Zika virus replication.

    Science.gov (United States)

    de Carvalho, Otavio Valério; Félix, Daniele Mendes; de Mendonça, Leila Rodrigues; de Araújo, Catarina Maria Cataldi Sabino; de Oliveira Franca, Rafael Freitas; Cordeiro, Marli Tenório; Silva Júnior, Abelardo; Pena, Lindomar José

    2017-12-01

    Since the emergence of Zika virus (ZIKV) in Brazil in 2015, 48 countries and territories in the Americas have confirmed autochthonous cases of disease caused by the virus. ZIKV-associated neurological manifestations and congenital defects make the development of safe and effective antivirals against ZIKV of utmost importance. Here we evaluated the antiviral activity of 6-methylmercaptopurine riboside (6MMPr), a thiopurine nucleoside analogue derived from the prodrug azathioprine, against the epidemic ZIKV strain circulating in Brazil. In all of the assays, an epithelial (Vero) and a human neuronal (SH-SY5Y) cell line were used to evaluate the cytotoxicity and effective concentrations of 6MMPr against ZIKV. Levels of ZIKV-RNA, viral infectious titre and the percentage of infected cells in the presence or absence of 6MMPr were used to determine antiviral efficacy. 6MMPr decreased ZIKV production by >99% in both cell lines in a dose- and time-dependent manner. Interestingly, 6MMPr was 1.6 times less toxic to SH-SY5Y cells compared with Vero cells, presenting a 50% cytotoxic concentrations (CC 50 ) of 460.3 µM and 291 µM, respectively. The selectivity index of 6MMPr for Vero and SH-SY5Y cells was 11.9 and 22.7, respectively, highlighting the safety profile of the drug to neuronal cells. Taken together, these results identify, for the first time, the thiopurine nucleoside analogue 6MMPr as a promising antiviral candidate against ZIKV that warrants further in vivo evaluation. Copyright © 2017 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

  5. Vaccinia virions lacking the RNA helicase nucleoside triphosphate phosphohydrolase II are defective in early transcription.

    Science.gov (United States)

    Gross, C H; Shuman, S

    1996-12-01

    Temperature-sensitive mutations (ts10, ts18, and ts39) of the vaccinia virus RNA helicase nucleoside triphosphate phosphohydrolase II (NPH-II) result in the production of noninfectious progeny virions at the restrictive temperature. The noninfectious mutant particles contain the wild-type complement of virion core and envelope polypeptides, as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The results of Western blot (immunoblot) analysis indicate that these particles lack NPH-II, whereas other enzymatic components of the virus core are present. These components include the following: DNA-dependent RNA polymerase subunits rpo147, rpo132, rpo94, rpo35, rpo30, rpo22, and rpo18; early transcription initiation factor subunits A8 and D6; mRNA capping enzyme subunits D1 and D12; RNA cap 2'-O-methyltransferase; A18 DNA helicase; DNA-dependent ATPase NPH-I; and DNA topoisomerase. Although RNA polymerase is encapsidated by the mutant viruses, mRNA synthesis in vitro by permeabilized mutant virions is only 5 to 20% that of the wild-type virus, as judged by nucleoside monophosphate incorporation into acid-insoluble material. Moreover, the transcripts synthesized by the mutant particles are longer than normal and remain virion associated. Transcription initiation by mutant virions occurs accurately at an endogenous genomic promoter, albeit at reduced levels (1 to 7%) compared with that of wild-type virions. In contrast, extracts of the mutant virions catalyze the wild-type level of transcription from an exogenous template containing an early promoter. We conclude that NPH-II is required for early mRNA synthesis uniquely in the context of the virus particle. Possible roles in transcription termination and RNA transport are discussed.

  6. Silicon(IV) phthalocyanines substituted axially with different nucleoside moieties. Effects of nucleoside type on the photosensitizing efficiencies and in vitro photodynamic activities.

    Science.gov (United States)

    Zheng, Bi-Yuan; Shen, Xiao-Min; Zhao, Dong-Mei; Cai, Yi-Bin; Ke, Mei-Rong; Huang, Jian-Dong

    2016-06-01

    A series of new silicon(IV) phthalocyanines (SiPcs) di-substituted axially with different nucleoside moieties have been synthesized and evaluated for their singlet oxygen quantum yields (ΦΔ) and in vitro photodynamic activities. The adenosine-substituted SiPc shows a lower photosensitizing efficiency (ΦΔ=0.35) than the uridine- and cytidine-substituted analogs (ΦΔ=0.42-0.44), while the guanosine-substituted SiPc exhibits a weakest singlet oxygen generation efficiency with a ΦΔ value down to 0.03. On the other hand, replacing axial adenosines with chloro-modified adenosines and purines can result in the increase of photogenerating singlet oxygen efficiencies of SiPcs. The formed SiPcs 1 and 2, which contain monochloro-modified adenosines and dichloro-modified purines respectively, appear as efficient photosensitizers with ΦΔ of 0.42-0.44. Both compounds 1 and 2 present high photocytotoxicities against HepG2 and BGC823 cancer cells with IC50 values ranging from 9nM to 33nM. The photocytotoxicities of these two compounds are remarkably higher than the well-known anticancer photosensitizer, chlorin e6 (IC50=752nM against HepG2 cells) in the same condition. As revealed by confocal microscopy, for both cell lines, compound 1 can essentially bind to mitochondria, while compound 2 is just partially localized in mitochondria. In addition, the two compounds induce cell death of HepG2 cells likely through apoptosis. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Composition and functionality of whole jamun based functional confection.

    Science.gov (United States)

    Sehwag, Sneha; Das, Madhusweta

    2016-06-01

    Whole jamun based functional confection (WJFC) was developed from an optimized blend (through response surface methodology) containing 26.585 % paste of jamun pulp with adhering skin, 2 % jamun seed powder, hydrocolloid mixture (2.289 % agar, 1.890 % pectin and 27.236 % polydextrose), antimicrobials (0.022 % benzoic acid and 0.085 % sorbic acid), and 40 % added water. The confection also contained 0.08 % sucralose, 0.06 % citric acid and 100 mg CaCl2.2H2O/g pectin. The confection was found to be rich in minerals like Ca, Mg, K, Na and P, with prebiotic activity and low glycemic index (48.1). Additionally, WJFC had reduced calorie (1.48 kcal/g) and high dietary fiber content (15.49 ± 0.058 g/100 g (db)). The antioxidant potential measured as DPPH radical scavenging activity and FRAP with different extraction solvents was found to range between 0.26 ± 0.01 and 0.98 ± 0.04 mg BHA/g and 2.57 ± 0.97 and 18.17 ± 1.30 μM Fe(2+)/g, respectively, with highest yield obtained for 50 % aq. ethanolic extract. Moreover, the antioxidant potential was observed to be dose dependent with IC50 values as 9.89 and 2.75 mg (db) against DPPH and superoxide anion radicals, respectively. WJFC was found to suppress α-amylase activity and retard glucose dialysis depicting the antidiabetic effect.

  8. Biosynthesis of N-(purin-6-ylcarbamoyl)-l-threonine riboside. Incorporation of l-threonine in vivo into modified nucleoside of transfer ribonucleic acid

    Science.gov (United States)

    Chheda, Girish B.; Hong, Chung Il; Piskorz, Conrad F.; Harmon, G. A.

    1972-01-01

    l-[U-14C]Threonine is incorporated into N-(purin-6-ylcarbamoyl)-l-threonine riboside of rat liver and Escherichia coli tRNA. A pathway is suggested for the biosynthesis of this nucleoside. PMID:4561775

  9. Bifunctional acyclic nucleoside phosphonates: synthesis of chiral 9-{3-hydroxy[1,4-bis(phosphonomethoxy)]butan-2-yl} derivatives of purines

    Czech Academy of Sciences Publication Activity Database

    Vrbková, Silvie; Dračínský, Martin; Holý, Antonín

    2007-01-01

    Roč. 18, č. 18 (2007), s. 2233-2247 ISSN 0957-4166 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * acyclic nucleoside bisphosphonates * phosphonomethyl ethers * purines Subject RIV: CC - Organic Chemistry Impact factor: 2.634, year: 2007

  10. .I.Ribo./I.-, .I.Xylo./I.-, and .I.Arabino./I.-configured adenine-based nucleoside phosphonates: synthesis of monomers for solid-phase oligonucleotide assembly

    Czech Academy of Sciences Publication Activity Database

    Páv, Ondřej; Buděšínský, Miloš; Rosenberg, Ivan

    2003-01-01

    Roč. 22, 5/8 (2003), s. 1053-1056 ISSN 1525-7770. [International Roundtable Nucleosides, Nucleotides and Nucleic Acids /15./. Leuven, 10.09.2002-14.09.2002] R&D Projects: GA ČR GA203/01/1166; GA AV ČR IAA4055101 Institutional research plan: CEZ:AV0Z4055905 Keywords : nucleoside phosphonic acids * adenosine * phosphonylation Subject RIV: CC - Organic Chemistry Impact factor: 0.813, year: 2003

  11. Bone mineral density changes in protease inhibitor-sparing vs. nucleoside reverse transcriptase inhibitor-sparing highly active antiretroviral therapy: data from a randomized trial

    DEFF Research Database (Denmark)

    Hansen, Ann-Brit Eg; Obel, N; Nielsen, H

    2011-01-01

    The aim of the study was to compare changes in bone mineral density (BMD) over 144 weeks in HIV-infected patients initiating nucleoside reverse transcriptase inhibitor (NRTI)-sparing or protease inhibitor-sparing highly active antiretroviral therapy (HAART).......The aim of the study was to compare changes in bone mineral density (BMD) over 144 weeks in HIV-infected patients initiating nucleoside reverse transcriptase inhibitor (NRTI)-sparing or protease inhibitor-sparing highly active antiretroviral therapy (HAART)....

  12. From norbornane-based nucleotide analogs locked in South conformation to novel inhibitors of feline herpes virus

    Czech Academy of Sciences Publication Activity Database

    Dejmek, Milan; Hřebabecký, Hubert; Šála, Michal; Dračínský, Martin; Procházková, Eliška; Leyssen, P.; Neyts, J.; Balzarini, J.; Nencka, Radim

    2014-01-01

    Roč. 22, č. 11 (2014), s. 2974-2983 ISSN 0968-0896 R&D Projects: GA MV VG20102015046 Institutional support: RVO:61388963 Keywords : carbocyclic nucleosides * nucleoside phosphonates * purine * norbornane * feline herpes virus Subject RIV: CC - Organic Chemistry Impact factor: 2.793, year: 2014

  13. Cross-coupling reactions of unprotected halopurine bases, nucleosides, nucleotides and nucleoside triphosphates with 4-boronophenylalanine in water. Synthesis of (purin-8-yl)- and (purin-6-yl)phenylalanines

    Czech Academy of Sciences Publication Activity Database

    Čapek, Petr; Pohl, Radek; Hocek, Michal

    2006-01-01

    Roč. 4, č. 11 (2006), s. 2278-2284 ISSN 1477-0520 R&D Projects: GA AV ČR(CZ) 1QS400550501; GA MŠk(CZ) 1M0508 Institutional research plan: CEZ:AV0Z40550506 Keywords : amino acids * purines * nucleosides * cross-coupling reactions Subject RIV: CC - Organic Chemistry Impact factor: 2.874, year: 2006

  14. Nonenantioselectivity Property of Human Deoxycytidine Kinase Explained by Structures of the Enzyme in Complex with [subscript L]- and [subscript D]-Nucleosides

    Energy Technology Data Exchange (ETDEWEB)

    Sabini, Elisabetta; Hazra, Saugata; Konrad, Manfred; Lavie, Arnon (UIC); (MXPL-G)

    2008-07-31

    Biological molecules are predominantly enantioselective. Yet currently two nucleoside analogue prodrugs (3TC and FTC) with opposite chirality compared to physiological nucleosides are clinically approved for the treatment of HIV infections. These prodrugs require conversion to their triphosphorylated forms to achieve pharmacological activity. The first step in the activation of these agents is catalyzed by human deoxycytidine kinase (dCK). This enzyme possesses the ability to phosphorylate nucleosides of the unnatural L-chirality. To understand the molecular basis for the nonenantioselectivity of dCK, we solved the crystal structures of the enzyme in complex with the L-enantiomer and of its physiological substrate deoxycytidine and with the L-nucleoside analogue FTC. These were compared to a structure solved with D-dC. Our results highlight structural adjustments imposed on the L-nucleosides and properties of the enzyme endowing it with the ability to phosphorylate substrates with nonphysiological chirality. This work reveals the molecular basis for the activation of L-nucleosides by dCK.

  15. Kinetic properties of Cellulomonas sp. purine nucleoside phosphorylase with typical and non-typical substrates: implications for the reaction mechanism.

    Science.gov (United States)

    Wielgus-Kutrowska, Beata; Bzowska, Agnieszka

    2005-01-01

    Phosphorolysis catalyzed by Cellulomonas sp. PNP with typical nucleoside substrate, inosine (Ino), and non-typical 7-methylguanosine (m7Guo), with either nucleoside or phosphate (Pd) as the varied substrate, kinetics of the reverse synthetic reaction with guanine (Gua) and ribose-1-phosphate (R1P) as the varied substrates, and product inhibition patterns of synthetic and phosphorolytic reaction pathways were studied by steady-state kinetic methods. It is concluded that, like for mammalian trimeric PNP, complex kinetic characteristics observed for Cellulomonas enzyme results from simultaneous occurrence of three phenomena. These are sequential but random, not ordered binding of substrates, tight binding of one substrate purine bases, leading to the circumstances that for such substrates (products) rapid-equilibrium assumptions do not hold, and a dual role of Pi, a substrate, and also a reaction modifier that helps to release a tightly bound purine base.

  16. Searching for novel N1-substituted benzimidazol-2-ones as non-nucleoside HIV-1 RT inhibitors.

    Science.gov (United States)

    Ferro, Stefania; Buemi, Maria Rosa; De Luca, Laura; Agharbaoui, Fatima E; Pannecouque, Christophe; Monforte, Anna-Maria

    2017-07-15

    Non-nucleoside reverse transcriptase inhibitors (NNRTIs) represent an integral part of the currently available combination antiretroviral therapy (cART) contributing to reduce the AIDS-mortality and turned the disease from lethal to chronic. In this context we recently reported a series of 6-chloro-1-(3-methylphenylsulfonyl)-1,3-dihydro-2H-benzimidazol-2-ones as potent non-nucleoside HIV-1 reverse transcriptase inhibitors. In this paper, we describe the design and the synthesis of two novel series of benzimidazolone analogues in which the linker moiety between the phenyl ring and the sulfonyl group was modified and new small lipophilic groups on the benzyl sulfonyl pendant were introduced. All the new obtained compounds were evaluated as RT inhibitors and were also tested against RTs containing single amino acid mutations. Finally, molecular docking studies were performed in order to rationalize the observed activity of the most promising compound. Copyright © 2017. Published by Elsevier Ltd.

  17. Comparative study of purine and pyrimidine nucleoside analogues acting on the thymidylate kinases of Mycobacterium tuberculosis and of humans.

    OpenAIRE

    Pochet , Sylvie; Dugué , Laurence; Labesse , Gilles; Delepierre , Muriel; Munier-Lehmann , Hélène

    2003-01-01

    Thymidine monophosphate kinase (TMPK) from Mycobacterium tuberculosis (TMPKmt) is an attractive target for the design of specific inhibitors. This fact is the result of its key role in the thymidine pathway and of unique structural features in the active site observed by X-ray crystallography, especially in comparison to its human counterpart (TMPKh). Different 5-modified thymidine derivatives, as well as purine and pyrimidine analogues or C-nucleosides were tested on TMPKmt and TMPKh, and th...

  18. Influence of Acyclic Nucleoside Phosphonate Antivirals on Gene Expression of Chemokine Receptors CCR5 and CXCR4

    Czech Academy of Sciences Publication Activity Database

    Potměšil, P.; Holý, Antonín; Zídek, Zdeněk

    2015-01-01

    Roč. 61, č. 1 (2015), s. 1-7 ISSN 0015-5500 R&D Projects: GA ČR GA305/03/1470; GA MŠk 1M0508 Institutional support: RVO:61388963 ; RVO:68378041 Keywords : acyclic nucleoside phosphonate * HIV * CCR5 * CXCR4 * cytokine * RT-PCR Subject RIV: CC - Organic Chemistry; FR - Pharmacology ; Medidal Chemistry (UEM-P) Impact factor: 0.833, year: 2015

  19. Estimation of apparent binding constant of complexes of selected acyclic nucleoside phosphonates with beta-cyclodextrin by affinity capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Šolínová, Veronika; Mikysková, Hana; Kaiser, Martin Maxmilian; Janeba, Zlatko; Holý, Antonín; Kašička, Václav

    2016-01-01

    Roč. 37, č. 2 (2016), s. 239-247 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GA13-17224S; GA ČR(CZ) GA15-01948S Institutional support: RVO:61388963 Keywords : acyclic nucleoside phosphonates * affinity capillary electrophoresis * binding constant * nucleotide analogs * beta-cyclodextrin Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.744, year: 2016

  20. Oxidative cleavage of ribofuranose 5-(alpha-hydroxyphosphonates): a route to erythrofuranose-based nucleoside phosphonic acids

    Czech Academy of Sciences Publication Activity Database

    Králíková, Šárka; Buděšínský, Miloš; Tomečková, Ivana; Rosenberg, Ivan

    2006-01-01

    Roč. 62, č. 41 (2006), s. 9742-9750 ISSN 0040-4020 R&D Projects: GA ČR GP203/04/P273; GA ČR GA203/05/0827; GA ČR GA202/05/0628 Institutional research plan: CEZ:AV0Z40550506 Keywords : AMP analogue * nucleoside phosphonic acid * sugar phosphonate Subject RIV: CC - Organic Chemistry Impact factor: 2.817, year: 2006

  1. Direct One-Pot Synthesis of Nucleosides from Unprotected or 5-O-Monoprotected D-Ribose

    Czech Academy of Sciences Publication Activity Database

    Downey, Alan Michael; Richter, C.; Pohl, Radek; Mahrwald, R.; Hocek, Michal

    2015-01-01

    Roč. 17, č. 18 (2015), s. 4604-4607 ISSN 1523-7060 R&D Projects: GA ČR GAP207/11/0344 Institutional support: RVO:61388963 Keywords : nucleosides * cytostatics * biological activity Subject RIV: CC - Organic Chemistry Impact factor: 6.732, year: 2015 http://pubs.acs.org/doi/pdf/10.1021/acs.orglett.5b02332

  2. Tetrazolylpyrene unnatural nucleoside as a human telomeric multimeric G-quadruplex selective switch-on fluorescent sensor.

    Science.gov (United States)

    Bag, Subhendu Sekhar; Pradhan, Manoj Kumar; Talukdar, Sangita

    2017-12-13

    We report herein the specific sensing of dimeric H45 G-quadruplex DNA via a fluorescence light-up response using fluorescent tetrazolylpyrene nucleoside ( TzPy B Do ) as a probe. The strong binding of the probe via an intercalative stacking interaction inside the connecting loop of two G-quadruplex units of H45 and the discrimination to other monomeric and long DNA duplexes are accompanied by a drastic enhancement of the emission intensity without compromising the conformation and stability.

  3. Insights into Phosphate Cooperativity and Influence of Substrate Modifications on Binding and Catalysis of Hexameric Purine Nucleoside Phosphorylases

    Science.gov (United States)

    de Giuseppe, Priscila O.; Martins, Nadia H.; Meza, Andreia N.; dos Santos, Camila R.; Pereira, Humberto D’Muniz; Murakami, Mario T.

    2012-01-01

    The hexameric purine nucleoside phosphorylase from Bacillus subtilis (BsPNP233) displays great potential to produce nucleoside analogues in industry and can be exploited in the development of new anti-tumor gene therapies. In order to provide structural basis for enzyme and substrates rational optimization, aiming at those applications, the present work shows a thorough and detailed structural description of the binding mode of substrates and nucleoside analogues to the active site of the hexameric BsPNP233. Here we report the crystal structure of BsPNP233 in the apo form and in complex with 11 ligands, including clinically relevant compounds. The crystal structure of six ligands (adenine, 2′deoxyguanosine, aciclovir, ganciclovir, 8-bromoguanosine, 6-chloroguanosine) in complex with a hexameric PNP are presented for the first time. Our data showed that free bases adopt alternative conformations in the BsPNP233 active site and indicated that binding of the co-substrate (2′deoxy)ribose 1-phosphate might contribute for stabilizing the bases in a favorable orientation for catalysis. The BsPNP233-adenosine complex revealed that a hydrogen bond between the 5′ hydroxyl group of adenosine and Arg43* side chain contributes for the ribosyl radical to adopt an unusual C3’-endo conformation. The structures with 6-chloroguanosine and 8-bromoguanosine pointed out that the Cl6 and Br8 substrate modifications seem to be detrimental for catalysis and can be explored in the design of inhibitors for hexameric PNPs from pathogens. Our data also corroborated the competitive inhibition mechanism of hexameric PNPs by tubercidin and suggested that the acyclic nucleoside ganciclovir is a better inhibitor for hexameric PNPs than aciclovir. Furthermore, comparative structural analyses indicated that the replacement of Ser90 by a threonine in the B. cereus hexameric adenosine phosphorylase (Thr91) is responsible for the lack of negative cooperativity of phosphate binding in this

  4. Synthesis of phosphonomethoxyethyl or 1,3-bis(phosphonomethoxy)propan-2-yl lipophilic esters of acyclic nucleoside phosphonates

    Czech Academy of Sciences Publication Activity Database

    Vrbková, Silvie; Dračínský, Martin; Holý, Antonín

    2007-01-01

    Roč. 63, č. 46 (2007), s. 11391-11398 ISSN 0040-4020 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Grant - others:Descartes Prize(XE) HPAW-2002-10096 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * alkoxyalkyl phosphonates * hexadecyloxypropyl ester groups * bisphosphonates Subject RIV: CC - Organic Chemistry Impact factor: 2.869, year: 2007

  5. Acyclic nucleoside bisphosphonates: Synthesis and properties of chiral 2-amino-4,6-bis[(phosphonomethoxy)alkoxy]pyrimidines

    Czech Academy of Sciences Publication Activity Database

    Doláková, Petra; Dračínský, Martin; Masojídková, Milena; Šolínová, Veronika; Kašička, Václav; Holý, Antonín

    2009-01-01

    Roč. 44, č. 6 (2009), s. 2408-2424 ISSN 0223-5234 R&D Projects: GA MŠk 1M0508 Grant - others:NIH(US) 1UC1AIO62540-01 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * pyrimidine * bisphosphonates Subject RIV: CC - Organic Chemistry Impact factor: 3.269, year: 2009

  6. Simple method for fast deprotection of nucleosides by triethylamine-catalyzed methanolysis of acetates in aqueous medium

    International Nuclear Information System (INIS)

    Meier, Lidiane; Monteiro, Gustavo C.; Baldissera, Rodrigo A.M.; Sa, Marcus Mandolesi

    2010-01-01

    A straightforward methodology for deacetylation of protected ribonucleosides was developed based on triethylamine-catalyzed solvolysis in aqueous methanol. Reactions are completed in a few minutes under microwave irradiation and the free nucleosides are obtained in high yield after simple evaporation of volatiles. Other important features include the involvement of readily available reagents and the compatibility with diverse functional groups, which make this process very attractive for broad application. (author)

  7. Enantiopurity analysis of new types of acyclic nucleoside phosphonates by capillary electrophoresis with cyclodextrins as chiral selectors

    Czech Academy of Sciences Publication Activity Database

    Šolínová, Veronika; Kaiser, Martin Maxmilian; Lukáč, Miloš; Janeba, Zlatko; Kašička, Václav

    2014-01-01

    Roč. 37, č. 3 (2014), s. 295-303 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GAP206/12/0453; GA ČR(CZ) GA13-17224S; GA MV VG20102015046 Institutional support: RVO:61388963 Keywords : acyclic nucleoside phosphonates * CE * chiral analysis * cyclodextrins * nucleotide analogs Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.737, year: 2014

  8. Hydrogen/Deuterium Exchange Mass Spectrometry Reveals Mechanistic Details of Activation of Nucleoside Diphosphate Kinases by Oligomerization

    OpenAIRE

    Dautant , Alain; Meyer , Philippe; Georgescauld , Florian

    2017-01-01

    International audience; Most oligomeric proteins become active only after assembly, but why oligomerization is required to support function is not well understood. Here, we address this question using the WT and a destabilized mutant (D93N) of the hexameric nucleoside diphosphate kinase from the pathogen Mycobacterium tuberculosis (Mt-NDPK). The conformational dynamics and oligomeric states of each were analyzed during unfolding/folding by Hydrogen/Deuterium exchange mass spectrometry (HDX-MS...

  9. The synthesis of piperidine nucleoside analogs-a comparison of several methods to access the introduction of nucleobases

    Czech Academy of Sciences Publication Activity Database

    Kovačková, Soňa; Dračínský, Martin; Rejman, Dominik

    2011-01-01

    Roč. 67, č. 7 (2011), s. 1485-1500 ISSN 0040-4020 R&D Projects: GA MŠk 2B06065; GA MZd NR9138; GA MŠk(CZ) LC06077 Institutional research plan: CEZ:AV0Z40550506 Keywords : piperidine derivatives * nucleoside analogs * nucleobase * proline Subject RIV: CC - Organic Chemistry Impact factor: 3.025, year: 2011

  10. Discrimination between acid and alkali-labile phosphorylated residues on Immobilon: phosphorylation studies of nucleoside diphosphate kinase

    DEFF Research Database (Denmark)

    Biondi, R M; Walz, K; Issinger, O G

    1996-01-01

    in buffers containing 5% methanol allows unambiguous distinction between serine/threonine and histidine phosphorylation (O-phosphomonoesters and phosphoramide, respectively) since under these conditions only one type of residue is dephosphorylated. The addition of 5% methanol to all buffers was indispensable...... phosphate transfer activity of nucleoside diphosphate kinase (NDP kinase). Nonetheless, a significant degree of autophosphorylation on other residues has been reported by several laboratories, and the hypothesis has been advanced that this nonhistidine phosphorylation may play an important role in NDP...

  11. Anthraquinone as a Redox Label for DNA: Synthesis, Enzymatic Incorporation, and Electrochemistry of Anthraquinone-Modified Nucleosides, Nucleotides, and DNA

    Czech Academy of Sciences Publication Activity Database

    Balintová, Jana; Pohl, Radek; Horáková Brázdilová, Petra; Vidláková, Pavlína; Havran, Luděk; Fojta, Miroslav; Hocek, Michal

    2011-01-01

    Roč. 17, č. 50 (2011), s. 14063-14073 ISSN 0947-6539 R&D Projects: GA MŠk(CZ) LC06035; GA MŠk LC512; GA AV ČR(CZ) IAA400040901 Institutional research plan: CEZ:AV0Z40550506; CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : anthraquinone * DNA * electrochemistry * nucleosides * oligonucleotides Subject RIV: CC - Organic Chemistry Impact factor: 5.925, year: 2011

  12. Distinct modulation of telomere length in two T-lymphoblastic leukemia cell lines by cytotoxic nucleoside phosphonates PMEG and PMEDAP

    Czech Academy of Sciences Publication Activity Database

    Hájek, Miroslav; Cvilink, Viktor; Votruba, Ivan; Holý, Antonín; Mertlíková-Kaiserová, Helena

    2010-01-01

    Roč. 643, č. 1 (2010), s. 6-12 ISSN 0014-2999 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * PMEG * PMEDAP * telomere length * telomerase inhibition Subject RIV: CC - Organic Chemistry Impact factor: 2.737, year: 2010

  13. Insights into phosphate cooperativity and influence of substrate modifications on binding and catalysis of hexameric purine nucleoside phosphorylases.

    Directory of Open Access Journals (Sweden)

    Priscila O de Giuseppe

    Full Text Available The hexameric purine nucleoside phosphorylase from Bacillus subtilis (BsPNP233 displays great potential to produce nucleoside analogues in industry and can be exploited in the development of new anti-tumor gene therapies. In order to provide structural basis for enzyme and substrates rational optimization, aiming at those applications, the present work shows a thorough and detailed structural description of the binding mode of substrates and nucleoside analogues to the active site of the hexameric BsPNP233. Here we report the crystal structure of BsPNP233 in the apo form and in complex with 11 ligands, including clinically relevant compounds. The crystal structure of six ligands (adenine, 2'deoxyguanosine, aciclovir, ganciclovir, 8-bromoguanosine, 6-chloroguanosine in complex with a hexameric PNP are presented for the first time. Our data showed that free bases adopt alternative conformations in the BsPNP233 active site and indicated that binding of the co-substrate (2'deoxyribose 1-phosphate might contribute for stabilizing the bases in a favorable orientation for catalysis. The BsPNP233-adenosine complex revealed that a hydrogen bond between the 5' hydroxyl group of adenosine and Arg(43* side chain contributes for the ribosyl radical to adopt an unusual C3'-endo conformation. The structures with 6-chloroguanosine and 8-bromoguanosine pointed out that the Cl(6 and Br(8 substrate modifications seem to be detrimental for catalysis and can be explored in the design of inhibitors for hexameric PNPs from pathogens. Our data also corroborated the competitive inhibition mechanism of hexameric PNPs by tubercidin and suggested that the acyclic nucleoside ganciclovir is a better inhibitor for hexameric PNPs than aciclovir. Furthermore, comparative structural analyses indicated that the replacement of Ser(90 by a threonine in the B. cereus hexameric adenosine phosphorylase (Thr(91 is responsible for the lack of negative cooperativity of phosphate binding

  14. Synthesis of Chromone, Quinolone, and Benzoxazinone Sulfonamide Nucleosides as Conformationally Constrained Inhibitors of Adenylating Enzymes Required for Siderophore Biosynthesis

    OpenAIRE

    Engelhart, Curtis A.; Aldrich, Courtney C.

    2013-01-01

    MbtA catalyzes the first committed step of mycobactin biosynthesis in Mycobacterium tuberculosis (Mtb) and is responsible for the incorporation of salicylic acid into the mycobactin siderophores. 5′-O-[N-(Salicyl)sulfamoyl]adenosine (Sal-AMS) is an extremely potent nucleoside inhibitor of MbtA that possesses excellent activity against whole-cell Mtb, but suffers from poor bioavailability. In an effort to improve the bioavailability, we have designed four conformationally constrained analogues...

  15. Activities of several classes of acyclic nucleoside phosphonates against camelpox virus replication in different cell culture models

    Czech Academy of Sciences Publication Activity Database

    Duraffour, S.; Snoeck, R.; Krečmerová, Marcela; Van Den Oord, J.; De Vos, D.; Holý, Antonín; Crance, J. M.; Garin, D.; De Clercq, E.; Andrei, G.

    2007-01-01

    Roč. 51, č. 12 (2007), s. 4410-4419 ISSN 0066-4804 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Grant - others:FWO(BE) G.0267.04; NIH(US) AI06540-01 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * antivirals * HPMP-5-azacytosine * camelpox virus Subject RIV: CC - Organic Chemistry Impact factor: 4.390, year: 2007

  16. Syntheses of pyrimidine acyclic nucleoside phosphonates as potent inhibitors of thymidine phosphorylase (PD-ECGF) from SD-lymphoma

    Czech Academy of Sciences Publication Activity Database

    Pomeisl, Karel; Votruba, Ivan; Holý, Antonín; Pohl, Radek

    2007-01-01

    Roč. 26, 8/9 (2007), s. 1025-1028 ISSN 1525-7770. [International Roundtable /17./. Bern, 03.09.2006-07.09.2006] R&D Projects: GA MŠk 1M0508 Grant - others:Descartes Prize(XE) HPAW-CT-2002-9001 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * thymidine phosphorylase * pyrimidines * FPMP derivatives * fluor ination Subject RIV: CC - Organic Chemistry Impact factor: 0.723, year: 2007

  17. Amalgamation of nucleosides and amino acids in antibiotic biosynthesis: discovery of an L-threonine:uridine-5'-aldehyde transaldolase.

    Science.gov (United States)

    Barnard-Britson, Sandra; Chi, Xiuling; Nonaka, Koichi; Spork, Anatol P; Tibrewal, Nidhi; Goswami, Anwesha; Pahari, Pallab; Ducho, Christian; Rohr, Jurgen; Van Lanen, Steven G

    2012-11-14

    The lipopeptidyl nucleoside antibiotics represented by A-90289, caprazamycin, and muraymycin are structurally highlighted by a nucleoside core that contains a nonproteinogenic β-hydroxy-α-amino acid named 5'-C-glycyluridine (GlyU). Bioinformatic analysis of the biosynthetic gene clusters revealed a shared open reading frame encoding a protein with sequence similarity to serine hydroxymethyltransferases, resulting in the proposal that this shared enzyme catalyzes an aldol-type condensation with glycine and uridine-5'-aldehyde to furnish GlyU. Using LipK involved in A-90289 biosynthesis as a model, we now functionally assign and characterize the enzyme responsible for the C-C bond-forming event during GlyU biosynthesis as an l-threonine:uridine-5'-aldehyde transaldolase. Biochemical analysis revealed this transformation is dependent upon pyridoxal-5'-phosphate, the enzyme has no activity with alternative amino acids, such as glycine or serine, as aldol donors, and acetaldehyde is a coproduct. Structural characterization of the enzyme product is consistent with stereochemical assignment as the threo diastereomer (5'S,6'S)-GlyU. Thus this enzyme orchestrates C-C bond breaking and formation with concomitant installation of two stereocenters to make a new l-α-amino acid with a nucleoside side chain.

  18. Immobilization of Neutral Protease from Bacillus subtilis for Regioselective Hydrolysis of Acetylated Nucleosides: Application to Capecitabine Synthesis

    Directory of Open Access Journals (Sweden)

    Teodora Bavaro

    2016-11-01

    Full Text Available This paper describes the immobilization of the neutral protease from Bacillus subtilis and its application in the regioselective hydrolysis of acetylated nucleosides, including building blocks useful for the preparation of anticancer products. Regarding the immobilization study, different results have been obtained depending on the immobilization procedure. Epoxy hydrophobic carriers gave a poorly stable derivative that released almost 50% of the immobilized protein under the required reaction conditions. On the contrary, covalent immobilization on a differently activated hydrophilic carrier (agarose resulted in very stable enzyme derivatives. In an attempt to explain the obtained enzyme immobilization results, the hypothetical localization of lysines on the enzyme surface was predicted in a 3D structure model of B. subtilis protease N built in silico by using the structure of Staphylococcus aureus metalloproteinase as the template. The immobilized enzyme shown a high regioselectivity in the hydrolysis of different peracetylated nucleosides. A stable enzyme derivative was obtained and successfully used in the development of efficient preparative bioprocesses for the hydrolysis of acetylated nucleosides, giving new intermediates for the synthesis of capecitabine in high yield.

  19. Lack of the nucleoside transporter ENT1 results in the Augustine-null blood type and ectopic mineralization.

    Science.gov (United States)

    Daniels, Geoff; Ballif, Bryan A; Helias, Virginie; Saison, Carole; Grimsley, Shane; Mannessier, Lucienne; Hustinx, Hein; Lee, Edmond; Cartron, Jean-Pierre; Peyrard, Thierry; Arnaud, Lionel

    2015-06-04

    The Augustine-negative alias At(a-) blood type, which seems to be restricted to people of African ancestry, was identified half a century ago but remains one of the last blood types with no known genetic basis. Here we report that a nonsynonymous single nucleotide polymorphism in SLC29A1 (rs45458701) is responsible for the At(a-) blood type. The resulting p.Glu391Lys variation in the last extracellular loop of the equilibrative nucleoside transporter 1 (ENT1; also called SLC29a1) is known not to alter its ability to transport nucleosides and nucleoside analog drugs. Furthermore, we identified 3 individuals of European ancestry who are homozygous for a null mutation in SLC29A1 (c.589+1G>C) and thus have the Augustine-null blood type. These individuals lacking ENT1 exhibit periarticular and ectopic mineralization, which confirms an important role for ENT1/SLC29A1 in human bone homeostasis as recently suggested by the skeletal phenotype of aging Slc29a1(-/-) mice. Our results establish Augustine as a new blood group system and place SLC29A1 as a new candidate gene for idiopathic disorders characterized with ectopic calcification/mineralization. © 2015 by The American Society of Hematology.

  20. Nucleoside adducts are formed by cooperative reaction of acetaldehyde and alcohols: Possible mechanism for the role of ethanol in carcinogenesis

    International Nuclear Information System (INIS)

    Fraenkel-Conrat, H.; Singer, B.

    1988-01-01

    The exocyclic amino groups of ribonucleosides and deoxyribonucleosides react rapidly at ambient temperature with acetaldehyde and alcohols to yield mixed acetals [-NH-CH(CH 3 )OR]. Nucleotides and nucleoside di- and triphosphates also react. Depending on the nucleoside used and on the relative amounts of aldehyde, alcohol, and water, preparative reactions reach equilibrium with yields up to 75% in a few house. The structures have been confirmed by fast atom bombardment MS and proton NMR. Half-lives at 37 degree C have been determined, and maximum stability is in the pH range of 7.5-9.5. In the absence of alcohol, acetaldehyde-nucleoside adducts could be isolated at 4 degree C, but these were too unstable to characterize except for their UV spectra, also at 4 degree C. Ethanol is often present in human blood and tissues, and acetaldehyde is its initial metabolic product, as well as being formed by many other metabolic processes. Both chemicals have separately been implicated in carcinogenic and other cytophathologic processes, but no cooperative mechanism has been proposed. The reactions reported here are of biological concern because they also occur in dilute aqueous solution. These finding supply a mechanism by which ethanol can be covalently bound to nucleic acids under physiological conditions

  1. Dolutegravir Plus Two Nucleoside Reverse Transcriptase Inhibitors versus Efavirenz Plus Two Nucleoside Reverse Transcriptase Inhibitors As Initial Antiretroviral Therapy for People with HIV: A Systematic Review.

    Science.gov (United States)

    Rutherford, George W; Horvath, Hacsi

    2016-01-01

    Dolutegravir (DTG) is a once-daily unboosted second-generation integrase-inhibitor that along with two nucleoside reverse transcriptase inhibitors is one of several regimens recommended by the United States, United Kingdom and European Union for first-line antiretroviral treatment of people with HIV infection. Our objective was to review the evidence for the efficacy and safety of DTG-based first-line regimens compared to efavirenz (EFV)-based regimens. We conducted a systematic review. We comprehensively searched a range of databases as well as conference abstracts and a trials registry. We used Cochrane methods in screening and data collection and assessed each study's risk of bias with the Cochrane tool. We meta-analyzed data using a fixed-effects model. We used GRADE to assess evidence quality. From 492 search results, we identified two randomized controlled trials, reported in five peer-reviewed articles and one conference abstract. One trial tested two DTG-based regimens (DTG + abacavir (ABC) + lamivudine (3TC) or DTG + tenofovir + emtricitabine) against an EFV-based regimen (EFV+ ABC+3TC). The other trial tested DTG+ABC+3TC against EFV+ABC+3TC. In meta-analysis, DTG-containing regimens were superior to EFV-containing regimens at 48 weeks and at 96 weeks (RR = 1.10, 95% CI 1.04-1.16; and RR = 1.12, 95% CI 1.04-1.21, respectively). In one trial, the DTG-containing regimen was superior at 144 weeks (RR = 1.13, 95% CI 1.02-1.24). DTG-containing regimens were superior in reducing treatment discontinuation compared to those containing EFV at 96 weeks and at 144 weeks (RR = 0.27, 95% CI 0.15-0.50; and RR = 0.28, 95% CI 0.16-0.48, respectively). Risk of serious adverse events was similar in each regimen at 96 weeks (RR = 1.15, 95% CI 0.80-1.63) and 144 weeks (RR = 0.93, 95% CI 0.68-1.29). Risk of bias was moderate overall, as was GRADE evidence quality. DTG-based regimens should be considered in future World Health Organization guidelines for initial HIV treatment.

  2. Determination of nucleosides in Cordyceps sinensis and Ganoderma lucidum by high performance liquid chromatography method

    Directory of Open Access Journals (Sweden)

    Masood Shah Khan

    2015-01-01

    Full Text Available Background: Nucleosides are supportive in the regulation and modulation of various physiological processes in body, they acts as precursors in nucleic acid synthesis, enhance immune response, help in absorption of iron and influence the metabolism of fatty acids. Cordyceps sinensis and Ganoderma lucidum are well-known for its use in traditional medicine of China, Nepal and India. They are rich in nucleosides such as adenine, adenosine, cordycepin, etc. Hence, a simple, economic and accurate high-performance liquid chromatography (HPLC analytical method was proposed for determination of adenine and adenosine for the quality control of plants. Materials and Methods: Chromatographic experiments were conducted on YL9100 HPLC system (South Korea. Reversed-phase chromatography was performed on a C18 column with methanol and dihydrogen phosphate as the mobile phase in isocratic elution method at a flow rate of 1.0 mL/min. Detection was carried out at 254 nm, which gives a sharp peak of adenine and adenosine at a retention time of 6.53 ± 0.02 min and 12.41 ± 0.02, respectively. Results and Discussion: Linear regression analysis data for the calibration plot showed a good linear relationship between response and concentration in the range of 25–200 µg/mL for adenosine and 100–800 µg/mL for adenine with regression coefficient of 0.999 and 0.996, respectively. The adenine was found 0.16% and 0.71% w/w in G. lucidum and in C. sinensis, respectively, and adenosine was found to be 0.14% w/w in G. lucidum whereas absent in C. sinensis. Conclusion: The developed HPLC method for the quantification of adenosine and adenine can be used for the quality control and standardization of crude drug and for the different herbal formulations, in which adenine and adenosine are present as major constituents. The wide linearity range, sensitivity, accuracy, and simple mobile phase imply the method is suitable for routine quantification of adenosine and adenine with

  3. Leishmania donovani Nucleoside Hydrolase terminal domains in cross-protective immunotherapy against Leishmania amazonensis murine infection

    Directory of Open Access Journals (Sweden)

    Dirlei eNico

    2014-06-01

    Full Text Available Nucleoside hydrolases of the Leishmania genus are vital enzymes for the replication of the DNA and conserved phylogenetic markers of the parasites. Leishmania donovani Nucleoside hydrolase (NH36 induced a main CD4+ T cell driven protective response against Leishmania chagasi infection in mice which is directed against its C-terminal domain. In this study, we used the three recombinant domains of NH36: N-terminal domain (F1, amino acids 1-103, central domain (F2 aminoacids 104-198 and C-terminal domain (F3 amino acids 199-314 in combination with saponin and assayed their immunotherapeutic effect on Balb/c mice previously infected with L. amazonensis. We identified that the F1 and F3 peptides determined strong cross-immunotherapeutic effects, reducing the size of footpad lesions to 48% and 64%, and the parasite load in footpads to 82.6% and 81%, respectively. The F3 peptide induced the strongest anti-NH36 antibody response and intradermal response (IDR against L. amazonenis and a high secretion of IFN-γ and TNF-α with reduced levels of IL-10. The F1 vaccine, induced similar increases of IgG2b antibodies and IFN-γ and TNF-α levels, but no IDR and no reduction of IL-10. The multiparameter flow cytometry analysis was used to assess the immune response after immunotherapy and disclosed that the degree of the immunotherapeutic effect is predicted by the frequencies of the CD4+ and CD8+ T cells producing IL-2 or TNF-α or both. Total frequencies and frequencies of double-cytokine CD4 T cell producers were enhanced by F1 and F3 vaccines. Collectively, our multifunctional analysis disclosed that immunotherapeutic protection improved as the CD4 responses progressed from 1+ to 2+, in the case of the F1 and F3 vaccines, and as the CD8 responses changed qualitatively from 1+ to 3+, mainly in the case of the F1 vaccine, providing new correlates of immunotherapeutic protection against cutaneous leishmaniasis in mice based on T-helper TH1 and CD8+ mediated

  4. Rigid Adenine Nucleoside Derivatives as Novel Modulators of the Human Sodium Symporters for Dopamine and Norepinephrine

    Science.gov (United States)

    Tosh, Dilip K.; Eshleman, Amy J.; Jacobson, Kenneth A.

    2016-01-01

    Thirty-two congeneric rigid adenine nucleoside derivatives containing a North (N)-methanocarba ribose substitution and a 2-arylethynyl group either enhanced (up to 760% of control) or inhibited [125I] methyl (1R,2S,3S)-3-(4-iodophenyl)-8-methyl-8-azabicyclo[3.2.1]octane-2-carboxylate (RTI-55) binding at the human dopamine (DA) transporter (DAT) and inhibited DA uptake. Several nucleosides also enhanced [3H]mazindol [(±)-5-(4-chlorophenyl)-3,5-dihydro-2H-imidazo[2,1-a]isoindol-5-ol] binding to the DAT. The combination of binding enhancement and functional inhibition suggests possible allosteric interaction with the tropanes. The structure-activity relationship of this novel class of DAT ligands was explored: small N6-substition (methyl or ethyl) was favored, while the N1 of the adenine ring was essential. Effective terminal aryl groups include thien-2-yl (compounds 9 and 16), with EC50 values of 35.1 and 9.1 nM, respectively, in [125I]RTI-55 binding enhancement, and 3,4-difluorophenyl as in the most potent DA uptake inhibitor (compound 6) with an IC50 value of 92 nM (3-fold more potent than cocaine), but not nitrogen heterocycles. Several compounds inhibited or enhanced binding at the norepinephrine transporter (NET) and serotonin transporter (SERT) and inhibited function in the micromolar range; truncation at the 4′-position in compound 23 allowed for weak inhibition of the SERT. We have not yet eliminated adenosine receptor affinity from this class of DAT modulators, but we identified modifications that remove DAT inhibition as an off-target effect of potent adenosine receptor agonists. Thus, we have identified a new class of allosteric DAT ligands, rigidified adenosine derivatives, and explored their initial structural requirements. They display a very atypical pharmacological profile, i.e., either enhancement by increasing affinity or inhibition of radioligand binding at the DAT, and in some cases the NET and SERT, and inhibition of neurotransmitter uptake

  5. Determination of nucleosides in Cordyceps sinensis and Ganoderma lucidum by high performance liquid chromatography method.

    Science.gov (United States)

    Khan, Masood Shah; Parveen, Rabea; Mishra, Kshipra; Tulsawani, Rajkumar; Ahmad, Sayeed

    2015-01-01

    Nucleosides are supportive in the regulation and modulation of various physiological processes in body, they acts as precursors in nucleic acid synthesis, enhance immune response, help in absorption of iron and influence the metabolism of fatty acids. Cordyceps sinensis and Ganoderma lucidum are well-known for its use in traditional medicine of China, Nepal and India. They are rich in nucleosides such as adenine, adenosine, cordycepin, etc. Hence, a simple, economic and accurate high-performance liquid chromatography (HPLC) analytical method was proposed for determination of adenine and adenosine for the quality control of plants. Chromatographic experiments were conducted on YL9100 HPLC system (South Korea). Reversed-phase chromatography was performed on a C18 column with methanol and dihydrogen phosphate as the mobile phase in isocratic elution method at a flow rate of 1.0 mL/min. Detection was carried out at 254 nm, which gives a sharp peak of adenine and adenosine at a retention time of 6.53 ± 0.02 min and 12.41 ± 0.02, respectively. Linear regression analysis data for the calibration plot showed a good linear relationship between response and concentration in the range of 25-200 µg/mL for adenosine and 100-800 µg/mL for adenine with regression coefficient of 0.999 and 0.996, respectively. The adenine was found 0.16% and 0.71% w/w in G. lucidum and in C. sinensis, respectively, and adenosine was found to be 0.14% w/w in G. lucidum whereas absent in C. sinensis. The developed HPLC method for the quantification of adenosine and adenine can be used for the quality control and standardization of crude drug and for the different herbal formulations, in which adenine and adenosine are present as major constituents. The wide linearity range, sensitivity, accuracy, and simple mobile phase imply the method is suitable for routine quantification of adenosine and adenine with high precision and accuracy.

  6. Carbocyclic Carbohydrate Mimics as Potential Glycosidase Inhibitors

    DEFF Research Database (Denmark)

    Fanefjord, Mette; Lundt, Inge

    It has been proven that aminocyclopentanols having the aminogroup adjacent to a carbon sidechain could be potential anomer-selective glycosidase inhibitors [1]. A successful pathway for synthesising mimics to L-carbohydrates 2, by introducing nitrogen to the C6 position in compound 1, has been...... developed in our group. A similar strategy has been used for synthesising mimics of D-carbohydrates. The α,β-unsaturated lactone 3 was cyclised to compound 4 which was further transformed into 5. The nitrogen functionality in compound 7 is introduced by an Overman rearrangement of 6 and the hydroxyl...

  7. Tunnel conductance of Watson-Crick nucleoside-base pairs from telegraph noise

    Energy Technology Data Exchange (ETDEWEB)

    Chang Shuai; He Jin; Lin Lisha; Zhang Peiming; Liang Feng; Huang Shuo; Lindsay, Stuart [Biodesign Institute, Arizona State University, Tempe, AZ 85287 (United States); Young, Michael [Department of Physics, Arizona State University, Tempe, AZ 85287 (United States)], E-mail: Stuart.Lindsay@asu.edu

    2009-05-06

    The use of tunneling signals to sequence DNA is presently hampered by the small tunnel conductance of a junction spanning an entire DNA molecule. The design of a readout system that uses a shorter tunneling path requires knowledge of the absolute conductance across base pairs. We have exploited the stochastic switching of hydrogen-bonded DNA base-nucleoside pairs trapped in a tunnel junction to determine the conductance of individual molecular pairs. This conductance is found to be sensitive to the geometry of the junction, but a subset of the data appears to come from unstrained molecular pairs. The conductances determined from these pairs are within a factor of two of the predictions of density functional calculations. The experimental data reproduces the counterintuitive theoretical prediction that guanine-deoxycytidine pairs (3 H-bonds) have a smaller conductance than adenine-thymine pairs (2 H-bonds). A bimodal distribution of switching lifetimes shows that both H-bonds and molecule-metal contacts break.

  8. Tunnel conductance of Watson-Crick nucleoside-base pairs from telegraph noise

    International Nuclear Information System (INIS)

    Chang Shuai; He Jin; Lin Lisha; Zhang Peiming; Liang Feng; Huang Shuo; Lindsay, Stuart; Young, Michael

    2009-01-01

    The use of tunneling signals to sequence DNA is presently hampered by the small tunnel conductance of a junction spanning an entire DNA molecule. The design of a readout system that uses a shorter tunneling path requires knowledge of the absolute conductance across base pairs. We have exploited the stochastic switching of hydrogen-bonded DNA base-nucleoside pairs trapped in a tunnel junction to determine the conductance of individual molecular pairs. This conductance is found to be sensitive to the geometry of the junction, but a subset of the data appears to come from unstrained molecular pairs. The conductances determined from these pairs are within a factor of two of the predictions of density functional calculations. The experimental data reproduces the counterintuitive theoretical prediction that guanine-deoxycytidine pairs (3 H-bonds) have a smaller conductance than adenine-thymine pairs (2 H-bonds). A bimodal distribution of switching lifetimes shows that both H-bonds and molecule-metal contacts break.

  9. A structural study of lamellar phases formed by nucleoside-functionalized lipids

    Energy Technology Data Exchange (ETDEWEB)

    Berti, D.; Fratini, E.; Baglioni, P. [Department of Chemistry and CSGI, University of Florence, Via G. Capponi 9, 50121 Florence (Italy); Dante, S.; Hauss, T. [Berlin Neutron Scattering Center, Hahn Meitner Institut, Glienicker Strasse 100, Wannsee, 14109 Berlin (Germany)

    2002-07-01

    We report a neutron-scattering investigation of lamellar phases formed by novel phospholipids bearing nucleosides at the polar-head-group region. These nucleolipids can interact through stacking and H-bond interactions, following a pattern that resembles base-base coupling in natural nucleic acids (DNA, RNA), i.e. they have similar recognition properties. Bilayer stacks formed of DPP-adenosine, DPP-uridine and their 1:1 mixture were investigated after equilibration in a 98% relative humidity atmosphere. The DPP-adenosine spectrum can be accounted for (in analogy to DPPC) by a lamellar phase with a smectic period of about 60 A. DPP-uridine displays a not so straightforward behavior that we have tentatively ascribed to the coexistence of lamellae with different smectic periods. In the 1:1 mixture the lamellar mesophase of DPP-uridine is retained, suggesting a specific interaction of the uridine polar-head group with the adenosine moiety of DPP-adenosine. It should be stressed that this behavior can be considered as an indication of the recognition process occurring at the polar-head-group region of the mixed phospholiponucleoside membrane. (orig.)

  10. A structural study of lamellar phases formed by nucleoside-functionalized lipids

    CERN Document Server

    Berti, D; Baglioni, P; Dante, S; Hauss, T

    2002-01-01

    We report a neutron-scattering investigation of lamellar phases formed by novel phospholipids bearing nucleosides at the polar-head-group region. These nucleolipids can interact through stacking and H-bond interactions, following a pattern that resembles base-base coupling in natural nucleic acids (DNA, RNA), i.e. they have similar recognition properties. Bilayer stacks formed of DPP-adenosine, DPP-uridine and their 1:1 mixture were investigated after equilibration in a 98% relative humidity atmosphere. The DPP-adenosine spectrum can be accounted for (in analogy to DPPC) by a lamellar phase with a smectic period of about 60 A. DPP-uridine displays a not so straightforward behavior that we have tentatively ascribed to the coexistence of lamellae with different smectic periods. In the 1:1 mixture the lamellar mesophase of DPP-uridine is retained, suggesting a specific interaction of the uridine polar-head group with the adenosine moiety of DPP-adenosine. It should be stressed that this behavior can be considere...

  11. Effect of purine nucleoside analogue-acyclovir on the sperm parameters and testosterone production in rats.

    Science.gov (United States)

    Movahed, Elham; Sadrkhanlou, Rajabali; Ahmadi, Abbas; Nejati, Vahid; Zamani, Zahra

    2013-04-01

    Acyclovir (ACV), a synthetic purine nucleoside analogue derived from guanosine, is known to be toxic to gonads and the aim of this study was to evaluate the effect of ACV on the sperm parameters and testosterone production in rat. In this experimental study, forty adult male Wistar rats (220 ± 20 g) were randomly divided into five groups (n=8 for each group). One group served as control and one group served as sham control [distilled water was intraperitoneally (i.p.) injected]. ACV was administered intraperitoneally in the drug treatment groups (4, 16 and 48 mg/kg/day) for 15 days. Eighteen days after the last injection, rats were sacrificed by CO2 inhalation. After that, cauda epididymides were removed surgically. At the end, sperm concentrations in the cauda epididymis, sperm motility, morphology, viability, chromatin quality and DNA integrity were analyzed. Serum testosterone concentrations were determined. The results showed that ACV did not affect sperm count, but decreased sperm motility and sperm viability at 16 and 48 mg/kg dose-levels. Sperm abnormalities increased at 48 mg/kg dose-level of ACV. Further, ACV significantly increases DNA damage at 16 and 48 mg/kg dose-levels and chromatin abnormality at all doses. Besides, a significant decrease in serum testosterone concentrations was observed at 16 and 48 mg/ kg doses. The present results highly support the idea that ACV induces testicular toxicity by adverse effects on the sperm parameters and serum level of testosterone in male rats.

  12. Cloning, molecular characterization and expression of ecto-nucleoside triphosphate diphosphohydrolase-1 from Torpedo electric organ.

    Science.gov (United States)

    Martín-Satué, Mireia; Torrejón-Escribano, Benjamín; Felipe, Antonio; de Aranda, Inmaculada Gómez; Elías, Marc; Marsal, Jordi; Blasi, Juan; Solsona, Carles

    2007-01-01

    During synaptic transmission large amounts of ATP are released from pre- and post-synaptic sources of Torpedo electric organ. A chain reaction sequentially hydrolyses ATP to adenosine, which inhibits acetylcholine secretion. The first enzyme implicated in this extracellular ATP hydrolysis is an ecto-nucleoside triphosphate diphosphohydrolase (E-NTPDase) that dephosphorylates both ATP and ADP to AMP. This enzyme has been biochemically characterized in the synaptosomal fraction of Torpedo electric organ, having almost equal affinity for ATP as for ADP, a fact that pointed to the type-1 NTPDase enzyme. In the present work we describe the cloning and molecular characterization of the cDNA for an NTPDase from Torpedo marmorata electric organ. The clone, obtained using the RACE-PCR technique, contains and open-reading frame of 1506bp and encodes a 502 amino acids protein that exhibits high homology with other NTPDases1 from vertebrates previously identified, including those of zebrafish and Xenopus, as well as human, rat and mouse. Topology analyses revealed the existence of two transmembrane regions, two short cytoplasmic tails and a long extracellular domain containing five apyrase-conserved regions. Gene expression studies revealed that this gene is expressed in all the Torpedo tissues analyzed. Finally, activity and cellular localization of the protein encoded by this newly cloned cDNA was assessed by heterologous expression experiments involving COS-7 and HeLa cells.

  13. 3D QSAR Studies of DAMNI Analogs as Possible Non-nucleoside Reverse Transcriptase Inhibitors

    Directory of Open Access Journals (Sweden)

    S. Ganguly

    2008-01-01

    Full Text Available The non-nucleoside inhibitors of HIV-1-reverse transcriptase (NNRTIs are an important class of drugs employed in antiviral therapy. Recently, a novel family of NNRTIs commonly referred to as 1-[2-diarylmethoxy] ethyl 2-methyl-5-nitroimidazoles (DAMNI derivatives have been discovered. The 3D-QSAR studies on DAMNI derivatives as NNRTIs was performed by comparative molecular field analysis (CoMFA and comparative molecular similarity indices analysis (CoMSIA methods to determine the factors required for the activity of these compounds. The global minimum energy conformer of the template molecule 15, the most active molecule of the series, was obtained by simulated annealing method and used to build the structures of the molecules in the dataset. The combination of steric and electrostatic fields in CoMSIA gave the best results with cross-validated and conventional correlation coefficients of 0.654 and 0.928 respectively. The predictive ability of CoMFA and CoMSIA were determined using a test set of ten DAMNI derivatives giving predictive correlation coefficients of 0.92 and 0.98 respectively indicating good predictive power. Further, the robustness of the models was verified by bootstrapping analysis. The information obtained from CoMFA and CoMSIA 3D contour maps may be of utility in the design of more potent DAMNI analogs as NNRTIs in future.

  14. The pharmacology of antiretroviral nucleoside and nucleotide reverse transcriptase inhibitors: implications for once-daily dosing.

    Science.gov (United States)

    Back, David J; Burger, David M; Flexner, Charles W; Gerber, John G

    2005-08-01

    The trend toward once-daily dosing in HIV antiretroviral therapy is based on the association between adherence, treatment outcome, and patient preferences. Patients prefer simpler treatments, fewer pills, less frequent dosing, and no food restrictions. When a regimen meets a patient's preferences, the patient is more likely to be adherent, and with good adherence, the regimen is more likely to be effective. Nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs) have been a prime focus for developing once-daily therapies primarily because they form the backbone of most current regimens. Within the NRTI class, however, drugs differ in their pharmacokinetic properties, such as plasma and intracellular half-lives, and thus in their suitability for once-daily dosing. For example, newer NRTIs, such as tenofovir and emtricitabine, combine longer plasma half-lives with longer intracellular half-lives, prolonging exposure and the period of pharmacologic activity. Of equal importance, the clinical impact of systemic and intracellular interactions between concomitant drugs defines which once-daily drugs may be combined in once-daily regimens. To construct simplified and effective therapies for individual patients, clinicians require an understanding of the plasma and intracellular pharmacokinetic properties of NRTIs and how these properties determine a drug's appropriateness for once-daily dosing and placement within a once-daily regimen.

  15. Structures and kinetics for plant nucleoside triphosphate diphosphohydrolases support a domain motion catalytic mechanism.

    Science.gov (United States)

    Summers, Emma L; Cumming, Mathew H; Oulavallickal, Tifany; Roberts, Nicholas J; Arcus, Vickery L

    2017-08-01

    Extracellular nucleoside triphosphate diphosphohydrolases (NTPDases) are enzymes that hydrolyze extracellular nucleotides to the respective monophosphate nucleotides. In the past 20 years, NTPDases belonging to mammalian, parasitic and prokaryotic domains of life have been discovered, cloned and characterized. We reveal the first structures of NTPDases from the legume plant species Trifolium repens (7WC) and Vigna unguiculata subsp. cylindrica (DbLNP). Four crystal structures of 7WC and DbLNP were determined at resolutions between 1.9 and 2.6 Å. For 7WC, structures were determined for an -apo form (1.89 Å) and with the product AMP (2.15 Å) and adenine and phosphate (1.76 Å) bound. For DbLNP, a structure was solved with phosphate and manganese bound (2.60 Å). Thorough kinetic data and analysis is presented. The structure of 7WC and DbLNP reveals that these NTPDases can adopt two conformations depending on the molecule and co-factor bound in the active site. A central hinge region creates a "butterfly-like" motion of the domains that reduces the width of the inter-domain active site cleft upon molecule binding. This phenomenon has been previously described in Rattus norvegicus and Legionella pneumophila NTPDaseI and Toxoplasma gondii NTPDaseIII suggesting a common catalytic mechanism across the domains of life. © 2017 The Protein Society.

  16. Structure-function relationship of substituted bromomethylcoumarins in nucleoside specificity of RNA alkylation.

    Science.gov (United States)

    Kellner, Stefanie; Kollar, Laura Bettina; Ochel, Antonia; Ghate, Manjunath; Helm, Mark

    2013-01-01

    Selective alkylation of RNA nucleotides is an important field of RNA biochemistry, e.g. in applications of fluorescent labeling or in structural probing experiments, yet detailed structure-function studies of labeling agents are rare. Here, bromomethylcoumarins as reactive compounds for fluorescent labeling of RNA are developed as an attractive scaffold on which electronic properties can be modulated by varying the substituents. Six different 4-bromomethyl-coumarins of various substitution patterns were tested for nucleotide specificity of RNA alkylation using tRNA from Escherichia coli as substrate. Using semi-quantitative LC-MS/MS analysis, reactions at mildly acidic and slightly alkaline pH were compared. For all tested compounds, coumarin conjugates with 4-thiouridine, pseudouridine, guanosine, and uridine were identified, with the latter largely dominating. This data set shows that selectivity of ribonucleotide alkylation depends on the substitution pattern of the reactive dye, and even more strongly on the modulation of the reaction conditions. The latter should be therefore carefully optimized when striving to achieve selectivity. Interestingly, the highest selectivity for labeling of a modified nucleoside, namely of 4-thiouridine, was achieved with a compound whose selectivity was somewhat less dependent on reaction conditions than the other compounds. In summary, bromomethylcoumarin derivatives are a highly interesting class of compounds, since their selectivity for 4-thiouridine can be efficiently tuned by variation of substitution pattern and reaction conditions.

  17. Radiochromatographic determination of activity of adenosine deaminase and purine nucleoside phosphorylase in blood cells

    International Nuclear Information System (INIS)

    Pechan, I.; Rendekova, V.; Pechanova, E.; Krizko, J.

    1982-01-01

    Expeditious and sensitive methods are described for determining the activities of adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP) in human lymphocytes and erythrocytes. ADA and PNP activity is determined on the basis of the reaction of (U- 14 C)adenosine or (8- 14 C)inosine with the lysate of human blood cells. Reaction products are separated using paper chromatography. Following the measurement of the radioactivity of spots of adenosine, inosine and hypoxanthine, a calculation is made of ADA and PNP activity from the results of the said measurements. On a sample of 52 clinically healthy people average ADA and PNP activity in isolated lymphocytes was found to be (51.6+-18.8) and (185.6+-94.7) pcat/10 6 cells and in erythrocytes (9.8+-2.98) and (17.1+-3.19) pcat/mg of proteins, respectively. The advantage of the method is the small amount of sample needed (1 to 2 ml) which allows its application in pediatrics. (Ha)

  18. Structural insight into activation mechanism of Toxoplasma gondii nucleoside triphosphate diphosphohydrolases by disulfide reduction.

    Science.gov (United States)

    Krug, Ulrike; Zebisch, Matthias; Krauss, Michel; Sträter, Norbert

    2012-01-27

    The intracellular parasite Toxoplasma gondii produces two nucleoside triphosphate diphosphohydrolases (NTPDase1 and -3). These tetrameric, cysteine-rich enzymes require activation by reductive cleavage of a hitherto unknown disulfide bond. Despite a 97% sequence identity, both isozymes differ largely in their ability to hydrolyze ATP and ADP. Here, we present crystal structures of inactive NTPDase3 as an apo form and in complex with the product AMP to resolutions of 2.0 and 2.2 Å, respectively. We find that the enzyme is present in an open conformation that precludes productive substrate binding and catalysis. The cysteine bridge 258-268 is identified to be responsible for locking of activity. Crystal structures of constitutively active variants of NTPDase1 and -3 generated by mutation of Cys(258)-Cys(268) show that opening of the regulatory cysteine bridge induces a pronounced contraction of the whole tetramer. This is accompanied by a 12° domain closure motion resulting in the correct arrangement of all active site residues. A complex structure of activated NTPDase3 with a non-hydrolyzable ATP analog and the cofactor Mg(2+) to a resolution of 2.85 Å indicates that catalytic differences between the NTPDases are primarily dictated by differences in positioning of the adenine base caused by substitution of Arg(492) and Glu(493) in NTPDase1 by glycines in NTPDase3.

  19. Molecular mechanism of distinct salt-dependent enzyme activity of two halophilic nucleoside diphosphate kinases.

    Science.gov (United States)

    Yamamura, Akihiro; Ichimura, Takefumi; Kamekura, Masahiro; Mizuki, Toru; Usami, Ron; Makino, Tsukasa; Ohtsuka, Jun; Miyazono, Ken-ichi; Okai, Masahiko; Nagata, Koji; Tanokura, Masaru

    2009-06-03

    Nucleoside diphosphate kinases from haloarchaea Haloarcula quadrata (NDK-q) and H. sinaiiensis (NDK-s) are identical except for one out of 154 residues, i.e., Arg(31) in NDK-q and Cys(31) in NDK-s. However, the salt-dependent activity profiles of NDK-q and NDK-s are quite different: the optimal NaCl concentrations of NDK-q and NDK-s are 1 M and 2 M, respectively. We analyzed the relationships of the secondary, tertiary, and quaternary structures and NDK activity of these NDKs at various salt concentrations, and revealed that 1), NDK-q is present as a hexamer under a wide range of salt concentrations (0.2-4 M NaCl), whereas NDK-s is present as a hexamer at an NaCl concentration above 2 M and as a dimer at NaCl concentrations below 1 M; 2), dimeric NDK-s has lower activity than hexameric NDK-s; and 3), dimeric NDK-s has higher helicity than hexameric NDK-s. We also determined the crystal structure of hexameric NDK-q, and revealed that Arg(31) plays an important role in stabilizing the hexamer. Thus the substitution of Arg (as in NDK-q) to Cys (as in NDK-s) at position 31 destabilizes the hexameric assembly, and causes dissociation to less active dimers at low salt concentrations.

  20. An Unusual Protector-Protégé Strategy for the Biosynthesis of Purine Nucleoside Antibiotics.

    Science.gov (United States)

    Wu, Pan; Wan, Dan; Xu, Gudan; Wang, Gui; Ma, Hongmin; Wang, Tingting; Gao, Yaojie; Qi, Jianzhao; Chen, Xiaoxia; Zhu, Jian; Li, Yong-Quan; Deng, Zixin; Chen, Wenqing

    2017-02-16

    Pentostatin (PTN, deoxycoformycin) and arabinofuranosyladenine (Ara-A, vidarabine) are purine nucleoside antibiotics used clinically to treat hematological cancers and human DNA virus infections, respectively. PTN has a 1,3-diazepine ring, and Ara-A is an adenosine analog with an intriguing epimerization at the C-2' hydroxyl group. However, the logic underlying the biosynthesis of these interesting molecules has long remained elusive. Here, we report that the biosynthesis of PTN and Ara-A employs an unusual protector-protégé strategy. To our surprise, we determined that a single gene cluster governs PTN and Ara-A biosynthesis via two independent pathways. Moreover, we verified that PenB functions as a reversible oxidoreductase for the final step of PTN. Remarkably, we provided the first direct biochemical evidence that PTN can protect Ara-A from deamination by selective inhibition of the host adenosine deaminase. These findings expand our knowledge of natural product biosynthesis and open the way for target-directed genome mining of Ara-A/PTN-related antibiotics. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Analogues of uracil nucleosides with intrinsic fluorescence (NIF-analogues): synthesis and photophysical properties.

    Science.gov (United States)

    Segal, Meirav; Fischer, Bilha

    2012-02-28

    Uridine cannot be utilized as fluorescent probe due to its extremely low quantum yield. For improving the uracil fluorescence characteristics we extended the natural chromophore at the C5 position by coupling substituted aromatic rings directly or via an alkenyl or alkynyl linker to create fluorophores. Extension of the uracil base was achieved by treating 5-I-uridine with the appropriate boronic acid under the Suzuki coupling conditions. Analogues containing an alkynyl linker were obtained from 5-I-uridine and the suitable boronic acid in a Sonogashira coupling reaction. The uracil fluorescent analogues proposed here were designed to satisfy the following requirements: a minimal chemical modification at a position not involved in base-pairing, resulting in relatively long absorption and emission wavelengths and high quantum yield. 5-((4-Methoxy-phenyl)-trans-vinyl)-2'-deoxy-uridine, 6b, was found to be a promising fluorescent probe. Probe 6b exhibits a quantum yield that is 3000-fold larger than that of the natural chromophore (Φ 0.12), maximum emission (478 nm) which is 170 nm red shifted as compared to uridine, and a Stokes shift of 143 nm. In addition, since probe 6b adopts the anti conformation and S sugar puckering favored by B-DNA, it makes a promising nucleoside analogue to be incorporated in an oligonucleotide probe for detection of genetic material.

  2. Novel anomeric sugar phosphodiesters synthesis, hydrolytic mechanism, structure and interaction with purine nucleoside phosphorylase

    International Nuclear Information System (INIS)

    Fathi, R.

    1988-01-01

    Some five-membered ring ribofuranosyl-1,2-cyclic phosphates were synthesized, purified, and characterized for the purpose of employing them as stereoselective electrophilic substrate analogs with a potential to trap enzymic nucleophiles on the purine salvage pathway. The purine salvage enzyme purine nucleoside phosphorylase from mammalian sources was irreversibly inactivated at its catalytic center by α-D-ribofuranosyl-1,2-cyclic monophosphate. The product distribution and kinetics of hydronium and hydroxide catalyzed hydrolysis of cyclic phosphates were monitored by 31 P NMR. Alkaline hydrolysis was demonstrated to proceed exclusively by O-P bond cleavage by employing a specifically 18 O-labelled substrate. Acid hydrolysis proceeded by C-O bond cleavage. The high rates of alkaline hydrolysis were similar to those reported for ethylene phosphate, presumably due to the presence of a strained cyclic phosphate ring. Extensive NMR ( 1 H, 13 C, and 31 P) data on the cyclic phosphates were consistent with a C3-endo ribofuranosyl conformation

  3. The dipeptide monoester prodrugs of floxuridine and gemcitabine-feasibility of orally administrable nucleoside analogs.

    Science.gov (United States)

    Tsume, Yasuhiro; Borras Bermejo, Blanca; Amidon, Gordon L

    2014-01-27

    Dipeptide monoester prodrugs of floxuridine and gemcitabine were synthesized. Their chemical stability in buffers, enzymatic stability in cell homogenates, permeability in mouse intestinal membrane along with drug concentration in mouse plasma, and anti-proliferative activity in cancer cells were determined and compared to their parent drugs. Floxuridine prodrug was more enzymatically stable than floxuridine and the degradation from prodrug to parent drug works as the rate-limiting step. On the other hand, gemcitabine prodrug was less enzymatically stable than gemcitabine. Those dipeptide monoester prodrugs exhibited 2.4- to 48.7-fold higher uptake than their parent drugs in Caco-2, Panc-1, and AsPC-1 cells. Floxuridine and gemcitabine prodrugs showed superior permeability in mouse jejunum to their parent drugs and exhibited the higher drug concentration in plasma after in situ mouse perfusion. Cell proliferation assays in ductal pancreatic cancer cells, AsPC-1 and Panc-1, indicated that dipeptide prodrugs of floxuridine and gemcitabine were more potent than their parent drugs. The enhanced potency of nucleoside analogs was attributed to their improved membrane permeability. The prodrug forms of 5¢-L-phenylalanyl-l-tyrosyl-floxuridine and 5¢-L-phenylalanyl-L-tyrosyl-gemcitabine appeared in mouse plasma after the permeation of intestinal membrane and the first-pass effect, suggesting their potential for the development of oral dosage form for anti-cancer agents.

  4. The Dipeptide Monoester Prodrugs of Floxuridine and Gemcitabine—Feasibility of Orally Administrable Nucleoside Analogs

    Directory of Open Access Journals (Sweden)

    Yasuhiro Tsume

    2014-01-01

    Full Text Available Dipeptide monoester prodrugs of floxuridine and gemcitabine were synthesized. Their chemical stability in buffers, enzymatic stability in cell homogenates, permeability in mouse intestinal membrane along with drug concentration in mouse plasma, and anti-proliferative activity in cancer cells were determined and compared to their parent drugs. Floxuridine prodrug was more enzymatically stable than floxuridine and the degradation from prodrug to parent drug works as the rate-limiting step. On the other hand, gemcitabine prodrug was less enzymatically stable than gemcitabine. Those dipeptide monoester prodrugs exhibited 2.4- to 48.7-fold higher uptake than their parent drugs in Caco-2, Panc-1, and AsPC-1 cells. Floxuridine and gemcitabine prodrugs showed superior permeability in mouse jejunum to their parent drugs and exhibited the higher drug concentration in plasma after in situ mouse perfusion. Cell proliferation assays in ductal pancreatic cancer cells, AsPC-1 and Panc-1, indicated that dipeptide prodrugs of floxuridine and gemcitabine were more potent than their parent drugs. The enhanced potency of nucleoside analogs was attributed to their improved membrane permeability. The prodrug forms of 5¢-L-phenylalanyl-l-tyrosyl-floxuridine and 5¢-L-phenylalanyl-L-tyrosyl-gemcitabine appeared in mouse plasma after the permeation of intestinal membrane and the first-pass effect, suggesting their potential for the development of oral dosage form for anti-cancer agents.

  5. The Dipeptide Monoester Prodrugs of Floxuridine and Gemcitabine—Feasibility of Orally Administrable Nucleoside Analogs

    Science.gov (United States)

    Tsume, Yasuhiro; Bermejo, Blanca Borras; Amidon, Gordon L.

    2014-01-01

    Dipeptide monoester prodrugs of floxuridine and gemcitabine were synthesized. Their chemical stability in buffers, enzymatic stability in cell homogenates, permeability in mouse intestinal membrane along with drug concentration in mouse plasma, and anti-proliferative activity in cancer cells were determined and compared to their parent drugs. Floxuridine prodrug was more enzymatically stable than floxuridine and the degradation from prodrug to parent drug works as the rate-limiting step. On the other hand, gemcitabine prodrug was less enzymatically stable than gemcitabine. Those dipeptide monoester prodrugs exhibited 2.4- to 48.7-fold higher uptake than their parent drugs in Caco-2, Panc-1, and AsPC-1 cells. Floxuridine and gemcitabine prodrugs showed superior permeability in mouse jejunum to their parent drugs and exhibited the higher drug concentration in plasma after in situ mouse perfusion. Cell proliferation assays in ductal pancreatic cancer cells, AsPC-1 and Panc-1, indicated that dipeptide prodrugs of floxuridine and gemcitabine were more potent than their parent drugs. The enhanced potency of nucleoside analogs was attributed to their improved membrane permeability. The prodrug forms of 5′-l-phenylalanyl-l-tyrosyl-floxuridine and 5′-l-phenylalanyl-l-tyrosyl-gemcitabine appeared in mouse plasma after the permeation of intestinal membrane and the first-pass effect, suggesting their potential for the development of oral dosage form for anti-cancer agents. PMID:24473270

  6. Supramolecular assemblies of nucleoside functionalized carbon nanotubes: synthesis, film preparation, and properties.

    Science.gov (United States)

    Micoli, Alessandra; Turco, Antonio; Araujo-Palomo, Elsie; Encinas, Armando; Quintana, Mildred; Prato, Maurizio

    2014-04-25

    Nucleoside-functionalized multi-walled carbon nanotubes (N-MWCNTs) were synthesized and characterized. A self-organization process using hydrogen bonding interactions was then used for the fabrication of self-assembled N-MWCNTs films free of stabilizing agents, polymers, or surfactants. Membranes were produced by using a simple water-dispersion-based vacuum-filtration method. Hydrogen-bond recognition was confirmed by analysis with IR spectroscopy and TEM images. Restoration of the electronic conduction properties in the N-MWCNTs membranes was performed by removing the organic portion by thermal treatment under an argon atmosphere to give d-N-MWCNTs. Electrical conductivity and thermal gravimetric analysis (TGA) measurements confirmed the efficiency of the annealing process. Finally, oxidative biodegradation of the films N-MWCNTs and d-N-MWCNTs was performed by using horseradish peroxidase (HRP) and low concentrations of H2 O2 . Our results confirm that functional groups play an important role in the biodegradation of CNT by HRP: N-MWCNTs films were completely biodegraded, whereas for d-N-MWCNTs films no degradation was observed, showing that the pristine CNT undergoes minimal enzyme-catalyzed oxidation This novel methodology offers a straightforward supramolecular strategy for the construction of conductive and biodegradable carbon nanotube films. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Application of nucleoside analogues to liver transplant recipients with hepatitis B

    Science.gov (United States)

    Song, Zhuo-Lun; Cui, Yu-Jun; Zheng, Wei-Ping; Teng, Da-Hong; Zheng, Hong

    2015-01-01

    Hepatitis B is a common yet serious infectious disease of the liver, affecting millions of people worldwide. Liver transplantation is the only possible treatment for those who advance to end-stage liver disease. Donors positive for hepatitis B virus (HBV) core antibody (HBcAb) have previously been considered unsuitable for transplants. However, those who test negative for the more serious hepatitis B surface antigen can now be used as liver donors, thereby reducing organ shortages. Remarkable improvements have been made in the treatment against HBV, most notably with the development of nucleoside analogues (NAs), which markedly lessen cirrhosis and reduce post-transplantation HBV recurrence. However, HBV recurrence still occurs in many patients following liver transplantation due to the development of drug resistance and poor compliance with therapy. Optimized prophylactic treatment with appropriate NA usage is crucial prior to liver transplantation, and undetectable HBV DNA at the time of transplantation should be achieved. NA-based and hepatitis B immune globulin-based treatment regimens can differ between patients depending on the patients’ condition, virus status, and presence of drug resistance. This review focuses on the current progress in applying NAs during the perioperative period of liver transplantation and the prophylactic strategies using NAs to prevent de novo HBV infection in recipients of HBcAb-positive liver grafts. PMID:26576094

  8. 6-methylmercaptopurine riboside, a thiopurine nucleoside with antiviral activity against canine distemper virus in vitro.

    Science.gov (United States)

    de Carvalho, Otávio Valério; Félix, Daniele Mendes; de Camargo Tozato, Claudia; Fietto, Juliana Lopes Rangel; de Almeida, Márcia Rogéria; Bressan, Gustavo Costa; Pena, Lindomar José; Silva-Júnior, Abelardo

    2017-06-26

    Canine distemper (CD) is a widespread infectious disease that can severely impact a variety of species in the order Carnivora, as well as non-carnivore species such as non-human primates. Despite large-scale vaccination campaigns, several fatal outbreaks have been reported in wild and domestic carnivore populations. This, in association with expansion of the disease host range and the development of vaccine-escape strains, has contributed to an increased demand for therapeutic strategies synergizing with vaccine programs for effectively controlling canine distemper. 6-methylmercaptopurine riboside (6MMPr) is a modified thiopurine nucleoside with known antiviral properties against certain RNA viruses. We tested the inhibitory effects of 6MMPr against a wild-type CDV strain infection in cell culture. We measured infectious particle production and viral RNA levels in treated and untreated CDV-infected cells. Ribavirin (RIB) was used as a positive control. Here, we report for the first time the antiviral effects of 6MMPr against canine distemper virus (CDV) in vitro. 6MMPr was able to reduce viral RNA levels and to inhibit the production of infectious CDV particles. The therapeutic selectivity of 6MMPr was approximately six times higher than that of ribavirin. Our results indicate that 6MMPr has high anti-CDV potential and warrants further testing against other paramyxoviruses, as well as clinical testing of the compound against CDV.

  9. Structure-function relationship of substituted bromomethylcoumarins in nucleoside specificity of RNA alkylation.

    Directory of Open Access Journals (Sweden)

    Stefanie Kellner

    Full Text Available Selective alkylation of RNA nucleotides is an important field of RNA biochemistry, e.g. in applications of fluorescent labeling or in structural probing experiments, yet detailed structure-function studies of labeling agents are rare. Here, bromomethylcoumarins as reactive compounds for fluorescent labeling of RNA are developed as an attractive scaffold on which electronic properties can be modulated by varying the substituents. Six different 4-bromomethyl-coumarins of various substitution patterns were tested for nucleotide specificity of RNA alkylation using tRNA from Escherichia coli as substrate. Using semi-quantitative LC-MS/MS analysis, reactions at mildly acidic and slightly alkaline pH were compared. For all tested compounds, coumarin conjugates with 4-thiouridine, pseudouridine, guanosine, and uridine were identified, with the latter largely dominating. This data set shows that selectivity of ribonucleotide alkylation depends on the substitution pattern of the reactive dye, and even more strongly on the modulation of the reaction conditions. The latter should be therefore carefully optimized when striving to achieve selectivity. Interestingly, the highest selectivity for labeling of a modified nucleoside, namely of 4-thiouridine, was achieved with a compound whose selectivity was somewhat less dependent on reaction conditions than the other compounds. In summary, bromomethylcoumarin derivatives are a highly interesting class of compounds, since their selectivity for 4-thiouridine can be efficiently tuned by variation of substitution pattern and reaction conditions.

  10. Complexes of cadmium(II) and mercury(II) with polyamines, nucleosides and nucleotides

    International Nuclear Information System (INIS)

    Lomozik, L.; Bregier-Jarzebowska, R.

    1999-01-01

    Computer analysis of potentiometric titration data was applied for determination of stability constants of Cd(II) and Hg(II) complexes in binary systems with polyamines (PA), nucleosides (Nuc) and nucleotides (NMP). For the systems of Hg(II) and PA an untypical increase in the complex stability with increasing ring size was observed and interpreted as the mercury preference to formation of linear complexes. Results of potentiometric and 13 C NMR studies for complexes of both metals indicate the involvement of all donor nitrogen atoms of di- and triamines in the coordination, leading to formation of N2 and N3 type chromophores, respectively. Monodentate complexes of Hg(II) with Cyd are formed already at very low pH (complexes with Cd from pH about 4). In the systems with AMP apart from nitrogen donor atoms, also the phosphate groups are involved in coordination. In the solid complexes of Cd(II) and Hg(II) with PA all donor atoms from the polyamines were found to be involved in the coordination and the presence of nitrate ions was established both in the inner and in the outer coordination spheres. (author)

  11. Nicotinamide riboside, an unusual, non-typical, substrate of purified purine-nucleoside phosphorylases.

    Science.gov (United States)

    Wielgus-Kutrowska, B; Kulikowska, E; Wierzchowski, J; Bzowska, A; Shugar, D

    1997-01-15

    Nicotinamide 1-beta-D-riboside (Nir), the cationic, reducible moiety of the coenzyme NAD+, has been confirmed as an unusual substrate for purified purine-nucleoside phosphorylase (PNP) from a mammalian source (calf spleen). It is also a substrate of the enzyme from Escherichia coli. The Km values at pH 7, 1.48 mM and 0.62 mM, respectively, were 1-2 orders of magnitude higher than for the natural substrate inosine, but the Vmax values were comparable, 96% and 35% that for Ino. The pseudo first-order rate constants, Vmax/Km, were 1.1% and 2.5% for the calf spleen and E. coli enzymes. The aglycon, nicotinamide, was neither a substrate nor an inhibitor of PNP. Nir was a weak inhibitor of inosine phosphorolysis catalyzed by both enzymes, with Ki values close to the Km for its phosphorolysis, consistent with simple competitive inhibition; this was further confirmed by Dixon plots. Phosphorolysis of the fluorescent positively charged substrate 7-methylguanosine was also inhibited in a competitive manner by both Ino and Nir. Phosphorolysis of Nir by both enzymes was inhibited competitively by several specific inhibitors of calf spleen and E. coli PNP, with Ki values similar to those for inhibition of other natural substrates. The pH dependence of the kinetic constants for the phosphorolysis of Nir and of a variety of other substrates, was extensively investigated, particularly in the alkaline pH range, where Nir exhibited abnormally high substrate activity relative to the reduced reaction rates of both enzymes towards other anionic or neutral substrates. The overall results are discussed in relation to present concepts regarding binding and phosphorolysis of substrates by PNP based on crystallographic data of enzyme-inhibitor complexes, and current studies on enzymatic and nonenzymatic mechanisms of the cleavage of the Nir glycosidic bond.

  12. Recognition of Nucleoside Monophosphate Substrates by Haemophilus influenzae Class C Acid Phosphatase

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Harkewal; Schuermann, Jonathan P.; Reilly, Thomas J.; Calcutt, Michael J.; Tanner, John J. (Cornell); (UMC)

    2010-12-08

    The e (P4) phosphatase from Haemophilus influenzae functions in a vestigial NAD{sup +} utilization pathway by dephosphorylating nicotinamide mononucleotide to nicotinamide riboside. P4 is also the prototype of class C acid phosphatases (CCAPs), which are nonspecific 5{prime},3{prime}-nucleotidases localized to the bacterial outer membrane. To understand substrate recognition by P4 and other class C phosphatases, we have determined the crystal structures of a substrate-trapping mutant P4 enzyme complexed with nicotinamide mononucleotide, 5{prime}-AMP, 3{prime}-AMP, and 2{prime}-AMP. The structures reveal an anchor-shaped substrate-binding cavity comprising a conserved hydrophobic box that clamps the nucleotide base, a buried phosphoryl binding site, and three solvent-filled pockets that contact the ribose and the hydrogen-bonding edge of the base. The span between the hydrophobic box and the phosphoryl site is optimal for recognizing nucleoside monophosphates, explaining the general preference for this class of substrate. The base makes no hydrogen bonds with the enzyme, consistent with an observed lack of base specificity. Two solvent-filled pockets flanking the ribose are key to the dual recognition of 5{prime}-nucleotides and 3{prime}-nucleotides. These pockets minimize the enzyme's direct interactions with the ribose and provide sufficient space to accommodate 5{prime} substrates in an anti conformation and 3{prime} substrates in a syn conformation. Finally, the structures suggest that class B acid phosphatases and CCAPs share a common strategy for nucleotide recognition.

  13. Comparison of agrobacterium mediated wheat and barley transformation with nucleoside diphosphate kinase 2 (NDPK2) gene

    International Nuclear Information System (INIS)

    Waheed, U.; Shah, M.M.; Smedley, M.; Harwood, W.

    2016-01-01

    An efficient and reliable transformation system is imperative for improvement of important crop species like barley and wheat. Wheat transformation is complex due to larger genome size and polyploidy while barley has a limitation of genotypic dependency. The objective of current study was to compare the relative transformation efficiency of wheat and barley using specific expression vector pBRACT 214-NDPK2 constructed through gateway cloning carrying Nucleoside Diphosphate Kinase 2 (NDPK2) gene. The vector was used to compare the transformation response in both crops using immature embryos through Agrobacterium mediated transformation. Both wheat and barley showed different responses towards callus induction and regeneration. Immature embryos of 1.5 to 2 mm in diameter was found optimum for wheat callus induction while 1 to 1.5 mm for barley. Both embryogenic and non-embryogenic calli were found in wheat with significantly greater tendency for embryogenecity in barley. The overall regeneration response was found different for all transformed wheat and barley cultivars. Wheat cultivars showed good response initially that drastically slowed down in later stages with the exception of Fielder that reached to the green shoots with good roots. The barley transformed lines showed good regeneration response as compared to wheat. PCR analysis of putative transformants using genomic DNA showed a maximum of 27% transformation efficiency in barely. No true transformation response was obtained in all cultivars of wheat used in this study. The protocol developed for wheat and barley transformation will greatly be helpful in crop improvement programme through genetic engineering especially in diploid relatives of cereals. (author)

  14. Kinetic characterization of ecto-nucleoside triphosphate diphosphohydrolases in brain nerve terminals during rat postnatal development

    Science.gov (United States)

    Stanojević, I.; Drakulić, D.; Petrović, S.; Milošević, M.; Jovanović, N.; Horvat, A.

    2011-12-01

    A family of enzymes named ecto-nucleoside triphosphate diphosphohydrolase (NTPDases) catalyzes the termination of ATP and ADP actions. Three different NTPDases (NTPDase 1-3), differing in their preference for a substrate, have been localized in the brain of adult mammals. The goal of our study was to clarify ATP and ADP hydrolyzing activities and kinetic parameters of NTPDases in synaptic plasma membranes (SPM) isolated from 15-, 30-, 60- and 90-days-old female rat brains. ATP and ADP hydrolysis were maximal in the presence of Mg2+ and showed insensitivity to ion-transporting ATPase inhibitors. The pronounced increase in both, ATP and ADP hydrolysis, were found in the SPM isolated from rats in the first month of life, stayed at the same level in the second month, and then decreased in adulthood. Kinetic analysis are also developmental-dependent, and together with the rate of ATP:ADP hydrolysis, point that all three NTPDases are present in SPM isolated from different developmental stages, with different, developmental-dependent proportion of activities. The lowest velocity and the highest affinity were observed for ATP hydrolyses, while the highest velocity and lowest affinity were detected for ADP hydrolyses in SPM isolated from 15-day old rats. Since specific ATP and ADP hydrolysis were lowest in this stage, we concluded that velocity is crucial for ATPase-, while affinity is for ADPase-part of NTPDases. Increased NTPDases activities, changes in their hydrolysis velocity and substrates affinities during rat postnatal development indicate involvement of adenine nucleotides in processes implicated to neuronal maturation and augmented neuroprotection.

  15. Preformulation studies of EFdA, a novel nucleoside reverse transcriptase inhibitor for HIV prevention.

    Science.gov (United States)

    Zhang, Wei; Parniak, Michael A; Mitsuya, Hiroaki; Sarafianos, Stefan G; Graebing, Phillip W; Rohan, Lisa C

    2014-08-01

    4'-Ethynyl-2-fluoro-2'-deoxyadenosine (EFdA) is a novel nucleoside analog of great interest because of its superior activity against wild-type and multidrug-resistant HIV-1 strains, and favorable safety profiles in vitro and in vivo. The aim of this work was to provide preformulation information of EFdA important for delivery system development. A simple, accurate and specific reverse-phase high performance liquid chromatographic (RP-HPLC) method with UV detection was developed for quantification of EFdA. In addition, physicochemical characterizations including pH solubility profile, octanol/water partition coefficient (Log Po/w), DSC analysis, field emission scanning electron microscopy, and stability studies under various conditions were conducted. EFdA existed in planar or flake shape, with a melting point of ∼130 °C, and had a pH dependent solubility. The log Po/w value of EFdA was -1.19. The compound was stable upon exposure to pH levels from 3 to 9 and showed good stability at elevated temperature (65 °C). In vitro cytotoxicity assessments were performed in two different epithelial cell lines. In cell-based studies, the EFdA selectivity index (50% cytotoxic concentration [CC50] values/50% effective concentration [EC50]) was found to be greater than 1 × 10(3). Permeability studies using cell- and tissue-based models showed that EFdA had an apparent permeability coefficient (Papp) <1 × 10(-6)cm/s and that the paracelluar pathway was the dominant transport route for EFdA. Overall, EFdA possesses favorable characteristics for further formulation development.

  16. Influence of Ecto-nucleoside triphosphate diphosphohydrolase activity on Trypanosoma cruzi infectivity and virulence.

    Directory of Open Access Journals (Sweden)

    Ramon F Santos

    Full Text Available BACKGROUND: The protozoan Trypanosoma cruzi is the causative agent of Chagas disease. There are no vaccines or effective treatment, especially in the chronic phase when most patients are diagnosed. There is a clear necessity to develop new drugs and strategies for the control and treatment of Chagas disease. Recent papers have suggested the ecto-nucleotidases (from CD39 family from pathogenic agents as important virulence factors. In this study we evaluated the influence of Ecto-Nucleoside-Triphosphate-Diphosphohydrolase (Ecto-NTPDase activity on infectivity and virulence of T. cruzi using both in vivo and in vitro models. METHODOLOGY/PRINCIPAL FINDINGS: We followed Ecto-NTPDase activities of Y strain infective forms (trypomastigotes obtained during sequential sub-cultivation in mammalian cells. ATPase/ADPase activity ratios of cell-derived trypomastigotes decreased 3- to 6-fold and infectivity was substantially reduced during sequential sub-cultivation. Surprisingly, at third to fourth passages most of the cell-derived trypomastigotes could not penetrate mammalian cells and had differentiated into amastigote-like parasites that exhibited 3- to 4-fold lower levels of Ecto-NTPDase activities. To evidence the participation of T. cruzi Ecto-NTPDase1 in the infective process, we evaluated the effect of known Ecto-ATPDase inhibitors (ARL 67156, Gadolinium and Suramin, or anti-NTPDase-1 polyclonal antiserum on ATPase and ADPase hydrolytic activities in recombinant T. cruzi NTPDase-1 and in live trypomastigotes. All tests showed a partial inhibition of Ecto-ATPDase activities and a marked inhibition of trypomastigotes infectivity. Mice infections with Ecto-NTPDase-inhibited trypomastigotes produced lower levels of parasitemia and higher host survival than with non-inhibited control parasites. CONCLUSIONS/SIGNIFICANCE: Our results suggest that Ecto-ATPDases act as facilitators of infection and virulence in vitro and in vivo and emerge as target

  17. The structure of FIV reverse transcriptase and its implications for non-nucleoside inhibitor resistance.

    Directory of Open Access Journals (Sweden)

    Meytal Galilee

    2018-01-01

    Full Text Available Reverse transcriptase (RT is the target for the majority of anti-HIV-1 drugs. As with all anti-AIDS treatments, continued success of RT inhibitors is persistently disrupted by the occurrence of resistance mutations. To explore latent resistance mechanisms potentially accessible to therapeutically challenged HIV-1 viruses, we examined RT from the related feline immunodeficiency virus (FIV. FIV closely parallels HIV-1 in its replication and pathogenicity, however, is resistant to all non-nucleoside inhibitors (NNRTI. The intrinsic resistance of FIV RT is particularly interesting since FIV harbors the Y181 and Y188 sensitivity residues absent in both HIV-2 and SIV. Unlike RT from HIV-2 or SIV, previous efforts have failed to make FIV RT susceptible to NNRTIs concluding that the structure or flexibility of the feline enzyme must be profoundly different. We report the first crystal structure of FIV RT and, being the first structure of an RT from a non-primate lentivirus, enrich the structural and species repertoires available for RT. The structure demonstrates that while the NNRTI binding pocket is conserved, minor subtleties at the entryway can render the FIV RT pocket more restricted and unfavorable for effective NNRTI binding. Measuring NNRTI binding affinity to FIV RT shows that the "closed" pocket configuration inhibits NNRTI binding. Mutating the loop residues rimming the entryway of FIV RT pocket allows for NNRTI binding, however, it does not confer sensitivity to these inhibitors. This reveals a further layer of resistance caused by inherent FIV RT variances that could have enhanced the dissociation of bound inhibitors, or, perhaps, modulated protein plasticity to overcome inhibitory effects of bound NNRTIs. The more "closed" conformation of FIV RT pocket can provide a template for the development of innovative drugs that could unlock the constrained pocket, and the resilient mutant version of the enzyme can offer a fresh model for the study

  18. Metabolites of purine nucleoside phosphorylase (NP in serum have the potential to delineate pancreatic adenocarcinoma.

    Directory of Open Access Journals (Sweden)

    Shaiju K Vareed

    2011-03-01

    Full Text Available Pancreatic Adenocarcinoma (PDAC, the fourth highest cause of cancer related deaths in the United States, has the most aggressive presentation resulting in a very short median survival time for the affected patients. Early detection of PDAC is confounded by lack of specific markers that has motivated the use of high throughput molecular approaches to delineate potential biomarkers. To pursue identification of a distinct marker, this study profiled the secretory proteome in 16 PDAC, 2 carcinoma in situ (CIS and 7 benign patients using label-free mass spectrometry coupled to 1D-SDS-PAGE and Strong Cation-Exchange Chromatography (SCX. A total of 431 proteins were detected of which 56 were found to be significantly elevated in PDAC. Included in this differential set were Parkinson disease autosomal recessive, early onset 7 (PARK 7 and Alpha Synuclein (aSyn, both of which are known to be pathognomonic to Parkinson's disease as well as metabolic enzymes like Purine Nucleoside Phosphorylase (NP which has been exploited as therapeutic target in cancers. Tissue Microarray analysis confirmed higher expression of aSyn and NP in ductal epithelia of pancreatic tumors compared to benign ducts. Furthermore, extent of both aSyn and NP staining positively correlated with tumor stage and perineural invasion while their intensity of staining correlated with the existence of metastatic lesions in the PDAC tissues. From the biomarker perspective, NP protein levels were higher in PDAC sera and furthermore serum levels of its downstream metabolites guanosine and adenosine were able to distinguish PDAC from benign in an unsupervised hierarchical classification model. Overall, this study for the first time describes elevated levels of aSyn in PDAC as well as highlights the potential of evaluating NP protein expression and levels of its downstream metabolites to develop a multiplex panel for non-invasive detection of PDAC.

  19. Virological and immunological outcomes at 3 years after starting antiretroviral therapy with regimens containing non-nucleoside reverse transcriptase inhibitor, protease inhibitor, or both in INITIO: open-label randomised trial

    DEFF Research Database (Denmark)

    Yeni, P; Cooper, DA; Aboulker, J-P

    2006-01-01

    antiretroviral therapy with two nucleoside analogue reverse transcriptase inhibitors (didanosine+stavudine) plus either a non-nucleoside reverse transcriptase inhibitor (efavirenz, EFV) or a protease inhibitor (nelfinavir, NFV), or both (EFV/NFV), in patients with HIV-1 infection who had not previously received...

  20. Simultaneous determination of nucleosides, myriocin, and carbohydrates in Cordyceps by HPLC coupled with diode array detection and evaporative light scattering detection.

    Science.gov (United States)

    Wang, Shuang; Yang, Feng-Qing; Feng, Kun; Li, De-Qiang; Zhao, Jing; Li, Shao-Ping

    2009-12-01

    A HPLC coupled with diode array detection (DAD) and evaporative light scattering detection (ELSD) method for qualitative and quantitative analysis of eight nucleosides and nucleobases, three carbohydrates and myriocin in Cordyceps was developed. A Prevail Carbohydrate ES column was employed for the separation within 50 min. Nucleosides and their bases were tested at UV 254 nm. ELSD was connected with DAD to determine myriocin and carbohydrates. The optimum drift tube temperature of ELSD was at 94 degrees C with the nitrogen flow rate of 2.0 L/min. All calibration curves showed good linearity (R(2)>0.9933) during the test ranges. The precision, repeatability, accuracy, LOD and LOQ were also fully investigated. This developed method was successfully applied to quantify 12 components, eight nucleosides and nucleobases, three carbohydrates and myriocin, in natural and cultured Cordyceps, which provides another view for quality control of Cordyceps sinensis.

  1. A randomized trial comparing initial HAART regimens of nelfinavir/nevirapine and ritonavir/saquinavir in combination with two nucleoside reverse transcriptase inhibitors

    DEFF Research Database (Denmark)

    Kirk, Ole; Lundgren, Jens D; Pedersen, Court

    2003-01-01

    BACKGROUND: A triple-class HAART regimen may be associated with a better virological effect than conventional regimens, but may also lead to toxicity and more profound resistance. METHODS: Randomized, controlled, open-label trial of 233 protease inhibitor- and non-nucleoside reverse transcriptase...... inhibitor-naive HIV-infected patients allocated to a regimen of nelfinavir and nevirapine (1250/200 mg twice daily; n = 118) or ritonavir and saquinavir (400/400 mg twice daily; n = 115), both in combination with two nucleoside reverse transcriptase inhibitors. The primary end-point was HIV RNA ....037. CONCLUSION: A regimen of nelfinavir/nevirapine had a favourable virological effect and tolerability over a 48-week period compared with ritonavir/saquinavir, when administered in combination with two nucleoside reverse transcriptase inhibitors. However, more extensive follow-up is required to determine...

  2. Synthesis and anticancer activity of new [(Indolyl)pyrazolyl]-1,3,4-oxadiazole thioglycosides and acyclic nucleoside analogs.

    Science.gov (United States)

    El-Sayed, Wael A; El-Sofany, Walaa I; Hussein, Hoda A R; Fathy, Nahed M

    2017-07-03

    New [(Indolyl)pyrazolyl]-1,3,4-oxadiazole compounds and their derived thioglycosides as well as the corresponding sugar hydrazones were synthesized. The acyclo C-nucleoside analogs of the oxadiazoline base system were also prepared by reaction of acid hydrazides with aldehydo sugars followed by one pot process encompassing acetylation and cyclization of the synthesized hydrazones. The anticancer activity of the newly synthesized compounds was studied against colorectal carcinoma (HCT116), breast adenocarcinoma (MCF7) and prostate cancer (PC3) human tumor cell lines and a number of compounds showed moderate to high activities.

  3. 1,3-Dipolar Cycloaddition (Part I: Synthesis of 1,2,3-Triazole Derivatives in Nucleoside Chemistry

    Directory of Open Access Journals (Sweden)

    Saftić D.

    2015-09-01

    Full Text Available Copper catalysed Huisgen 1,3-dipolar cycloaddition of azides and alkynes which leads to 1,4-disubstituted-1,2,3-triazoles is a frequently used method in synthetic organic chemistry. Due to the simple reaction conditions, it has occupied an important place in the field of nucleoside chemistry since it enables preparation of a large number of potentially biologically active compounds with a number of interesting additional properties induced by the presence of 1,2,3-triazole structural motif in the molecule.

  4. Pyrrolo[2,3-d]pyrimidine (7-deazapurine) as a privileged scaffold in design of antitumor and antiviral nucleosides

    Czech Academy of Sciences Publication Activity Database

    Perlíková, Pavla; Hocek, Michal

    2017-01-01

    Roč. 37, č. 6 (2017), s. 1429-1460 ISSN 0198-6325 R&D Projects: GA ČR(CZ) GA16-00178S Grant - others:AV ČR(CZ) AP1501 Program:Akademická prémie - Praemium Academiae Institutional support: RVO:61388963 Keywords : antivirals * cytostatics * deazapurines * nucleosides * nucleotides Subject RIV: CC - Organic Chemistry OBOR OECD: Organic chemistry Impact factor: 8.763, year: 2016 http://onlinelibrary.wiley.com/doi/10.1002/med.21465/full

  5. Muraymycin nucleoside-peptide antibiotics: uridine-derived natural products as lead structures for the development of novel antibacterial agents

    Directory of Open Access Journals (Sweden)

    Daniel Wiegmann

    2016-04-01

    Full Text Available Muraymycins are a promising class of antimicrobial natural products. These uridine-derived nucleoside-peptide antibiotics inhibit the bacterial membrane protein translocase I (MraY, a key enzyme in the intracellular part of peptidoglycan biosynthesis. This review describes the structures of naturally occurring muraymycins, their mode of action, synthetic access to muraymycins and their analogues, some structure–activity relationship (SAR studies and first insights into muraymycin biosynthesis. It therefore provides an overview on the current state of research, as well as an outlook on possible future developments in this field.

  6. Anthraquinone as a redox label for DNA. Synthesis, enzymatic incorporation and electrochemistry of anthraquinone modified nucleosides, nucleotides and DNA

    Czech Academy of Sciences Publication Activity Database

    Balintová, J.; Havran, Luděk; Fojta, Miroslav; Hocek, Michal

    2012-01-01

    Roč. 106, - (2012), s1033-s1033 ISSN 0009-2770. [EuCheMS Chemistry Congress /4./. 26.08.2012-30.08.2012, Prague] R&D Projects: GA ČR GA203/09/0317; GA MŠk LC512; GA MŠk 1M0508; GA AV ČR(CZ) IAA400040901 Institutional research plan: CEZ:AV0Z40550506; CEZ:AV0Z50040702 Keywords : nucleosides * oligonucleotides * electrochemistry Subject RIV: CC - Organic Chemistry

  7. Structure-enhanced methods in the development of non-nucleoside inhibitors targeting HIV reverse transcriptase variants.

    Science.gov (United States)

    Frey, Kathleen M

    2015-01-01

    Resistance continues to emerge as a leading cause for antiretroviral treatment failure. Several mutations in HIV reverse transcriptase (RT) confer resistance to non-nucleoside inhibitors (NNRTIs), vital components of antiretroviral combination therapies. Since the majority of mutations are located in the NNRTI binding pocket, crystal structures of RT variants in complex with NNRTIs have provided ideas for new drug design strategies. This article reviews the impact of RT crystal structures on the multidisciplinary design and development of new inhibitors with improved resistance profiles.

  8. Inhibitory activities of three classes of acyclic nucleoside phosphonates against murine polyomavirus and primate simian virus 40 strains

    Czech Academy of Sciences Publication Activity Database

    Lebeau, I.; Andrei, G.; Krečmerová, Marcela; De Clercq, E.; Holý, Antonín; Snoeck, R.

    2007-01-01

    Roč. 51, č. 6 (2007), s. 2268-2273 ISSN 0066-4804 R&D Projects: GA AV ČR 1QS400550501; GA MŠk 1M0508 Grant - others:FWO(BE) G.0267.04; NIH(US) AI 062540-01; René Descartes Prize 2001(XE) HPAV-2002-100096 Institutional research plan: CEZ:AV0Z40550506 Keywords : acyclic nucleoside phosphonates * polyomavirus * 5-azacytosine Subject RIV: CC - Organic Chemistry Impact factor: 4.390, year: 2007

  9. Synthesis of Conformationally North-Locked Pyrimidine Nucleosides Built on an Oxabicyclo[3.1.0]hexane Scaffold | Center for Cancer Research

    Science.gov (United States)

    Beginning with a known 3-oxabicyclo[3.1.0]-hexane scaffold, the relocation of the fused cyclopropane ring bond and the shifting of the oxygen atom to an alternative location engendered a new 2-oxabicyclo[3.1.0]hexane template that mimics more closely the tetrahydrofuran ring of conventional nucleosides. The synthesis of this new class of locked nucleosides involved a novel approach that required the isocyanate with a hydroxyl-protected scaffold as a pivotal intermediate that was obtained in 11 steps from a known dihydrofuran precursor.

  10. Long-lasting protection of activity of nucleoside reverse transcriptase inhibitors and protease inhibitors (PIs by boosted PI containing regimens.

    Directory of Open Access Journals (Sweden)

    Alexandra U Scherrer

    Full Text Available BACKGROUND: The accumulation of mutations after long-lasting exposure to a failing combination antiretroviral therapy (cART is problematic and severely reduces the options for further successful treatments. METHODS: We studied patients from the Swiss HIV Cohort Study who failed cART with nucleoside reverse transcriptase inhibitors (NRTIs and either a ritonavir-boosted PI (PI/r or a non-nucleoside reverse transcriptase inhibitor (NNRTI. The loss of genotypic activity 6 months after virological failure was analyzed with Stanford algorithm. Risk factors associated with early emergence of drug resistance mutations (6 months after failure: 8.8% vs. 38.2% (p = 0.009, 7.1% vs. 46.9% (p6 months after failure compared to 41.2%, 49.0% and 63.0% of those who have lost NNRTI activity (all p<0.001. The risk to accumulate an early NRTI mutation was strongly associated with NNRTI-containing cART (adjusted odds ratio: 13.3 (95% CI: 4.1-42.8, p<0.001. CONCLUSIONS: The loss of activity of PIs and NRTIs was low among patients treated with PI/r, even after long-lasting exposure to a failing cART. Thus, more options remain for second-line therapy. This finding is potentially of high relevance, in particular for settings with poor or lacking virological monitoring.

  11. Nucleoside uptake in macrophages from various murine strains: a short-time and a two-step stimulation model

    International Nuclear Information System (INIS)

    Busolo, F.; Conventi, L.; Grigolon, M.; Palu, G.

    1991-01-01

    Kinetics of [3H]-uridine uptake by murine peritoneal macrophages (pM phi) is early altered after exposure to a variety of stimuli. Alterations caused by Candida albicans, lipopolysaccharide (LPS) and recombinant interferon-gamma (rIFN-gamma) were similar in SAVO, C57BL/6, C3H/HeN and C3H/HeJ mice, and were not correlated with an activation process as shown by the amount of tumor necrosis factor-alpha (TNF-alpha) being released. Short-time exposure to all stimuli resulted in an increased nucleoside uptake by SAVO pM phi, suggesting that the tumoricidal function of this cell either depends from the type of stimulus or the time when the specific interaction with the cell receptor is taking place. Experiments with priming and triggering signals confirmed the above findings, indicating that the increase or the decrease of nucleoside uptake into the cell depends essentially on the chemical nature of the priming stimulus. The triggering stimulus, on the other hand, is only able to amplify the primary response

  12. Trifunctional fluorescent unnatural nucleoside: Label free detection of T-T/C-C base mismatches, abasic site and bulge DNA.

    Science.gov (United States)

    Bag, Subhendu Sekhar; Pradhan, Manoj Kumar; Talukdar, Sangita

    2017-08-01

    The detection and targeting of both the mismatched and abasic DNA is highly important which would ultimately help in designing new diagnostics and chemotherapeutics. Furthermore, sensing and targeting the bulge sequence with a fluorescent probe would be useful to study the role of bulges in nucleic acid function or could have significant therapeutic potential. Thus, detection of specific bulges by small fluorescent molecules is an attractive research area since the past several years. Many attempts have been made to prepare such compounds. We report herein a label free strategy for the detection of pyrimidine base mismatches (T/T and C/C), sensing of abasic site, and pyrimidine base bulge DNA using an unnatural tetrazolylpyrene nucleoside ( TPy B Do ) as a bare fluorescent probe. The H-bonding/hydrophobic force mediated interactions allow the sensing of all three deformed DNA via an enhancement of fluorescence signal using our simple "Just-Mix and Read" strategy. The binding of the probe to all the three deformed DNA duplexes is accompanied by an increase in the thermal melting stability of the deformed DNAs. That the probe binds efficiently to the minor groove near the deformed site was evident from spectroscopic studies. All the spectral evidences open up a multitude of possibilities for using our probe, tetrazolylpyrene nucleoside, as an efficient fluorescent light-up bio-probe for label free DNA detection. Copyright © 2017. Published by Elsevier B.V.

  13. Fast Simultaneous Determination of 13 Nucleosides and Nucleobases in Cordyceps sinensis by UHPLC–ESI–MS/MS

    Directory of Open Access Journals (Sweden)

    Shi-Yu Zong

    2015-12-01

    Full Text Available A reliable ultra-high-performance liquid chromatography–electrospray ionization–tandem mass spectrometry (UHPLC–ESI–MS/MS method for the fast simultaneous determination of 13 nucleosides and nucleobases in Cordyceps sinensis (C. sinensis with 2-chloroadenosine as internal standard was developed and validated. Samples were ultrasonically extracted in an ice bath thrice, and the optimum analyte separation was performed on an ACQUITY UPLCTM HSS C18 column (100 mm × 2.1 mm, 1.8 μm with gradient elution. All targeted analytes were separated in 5.5 min. Furthermore, all calibration curves showed good linear regression (r > 0.9970 within the test ranges, and the limits of quantitation and detection of the 13 analytes were less than 150 and 75 ng/mL, respectively. The relative standard deviations (RSDs of intra- and inter-day precisions were <6.23%. Recoveries of the quantified analytes ranged within 85.3%–117.3%, with RSD < 6.18%. The developed UHPLC–ESI–MS/MS method was successfully applied to determine nucleosides and nucleobases in 11 batches of C. sinensis samples from different regions in China. The range for the total content in the analyzed samples was 1329–2057 µg/g.

  14. Characterization of nucleoside triphosphate diphosphohydrolase activity in Trichomonas gallinae and the influence of penicillin and streptomycin in extracellular nucleotide hydrolysis.

    Science.gov (United States)

    Borges, Fernanda Pires; de Brum Vieira, Patrícia; Wiltuschnig, Renata C M; Tasca, Tiana; De Carli, Geraldo Attilio; Bonan, Carla Denise

    2008-06-01

    Here we described an nucleoside triphosphate diphosphohydrolase (NTPDase) activity in living trophozoites of Trichomonas gallinae. The enzyme hydrolyzes a variety of purine and pyrimidine nucleoside di- and triphosphates in an optimum pH range of 6.0-8.0. This enzyme activity was activated by high concentrations of divalent cations, such as calcium and magnesium. Contaminant activities were ruled out because the enzyme was not inhibited by classical inhibitors of ATPases (ouabain, 5.0 mM sodium azide, oligomycin) and alkaline phosphatases (levamisole). A significant inhibition of ATP hydrolysis (38%) was observed in the presence of 20 mM sodium azide. Sodium orthovanadate inhibited ATP and ADP hydrolysis (24% and 78%), respectively. The apparent K(M) (Michaelis constant) values were 667.62+/-13 microM for ATP and 125+/-5.3 microM for ADP. V(max) (maximum velocity) values were 0.44+/-0.007 nmol Pi min(-1) per 10(6) trichomonads and 0.91+/-0.12 nmol Pi min(-1) per 10(6) trichomonads for ATP and ADP, respectively. Moreover, we showed a marked decrease in ATP, ADP and AMP hydrolysis when the parasites were grown in the presence of penicillin and streptomycin. The existence of an NTPDase activity in T. gallinae may be involved in pathogenicity, protecting the parasite from the cytolytic effects of the extracellular nucleotides.

  15. Production, purification, crystallization and preliminary X-ray diffraction studies of the nucleoside diphosphate kinase b from Leishmania major

    International Nuclear Information System (INIS)

    Tonoli, Celisa Caldana Costa; Vieira, Plinio Salmazo; Ward, Richard John; Arni, Raghuvir Krishnaswamy; Oliveira, Arthur Henrique Cavalcante de; Murakami, Mario Tyago

    2009-01-01

    Overexpression, purification, crystallization and preliminary X-ray diffraction analysis of the nucleoside diphosphate kinase b from Leishmania major are reported. The crystals belonged to the trigonal space group P3 2 21 and diffracted to 2.18 Å resolution. Nucleoside diphosphate kinases (NDKs; EC 2.7.4.6) play an essential role in the synthesis of nucleotides from intermediates in the salvage pathway in all parasitic trypanosomatids and their structural studies will be instrumental in shedding light on the biochemical machinery involved in the parasite life cycle and host–parasite interactions. In this work, NDKb from Leishmania major was overexpressed in Escherichia coli, purified to homogeneity and crystallized using the sitting-drop vapour-diffusion method. The NDK crystal diffracted to 2.2 Å resolution and belonged to the trigonal crystal system, with unit-cell parameters a = 114.2, c = 93.9 Å. Translation-function calculations yielded an unambiguous solution in the enantiomorphic space group P3 2 21

  16. OH-induced free radicals in purine nucleoside monophosphates: e.s.r. and spin-trapping

    International Nuclear Information System (INIS)

    Hiraoka, W.; Kuwabara, M.; Sato, F.

    1989-01-01

    Free radicals produced by the reactions of OH radicals with six purine nucleoside monophosphates (3'-AMP, 5'-AMP, 5'-dAMP, 3'-GMP, 5'GMP and 5'-dGMP) were investigated by a method combining e.s.r. spin-trapping and high-performance liquid chromatography (HPLC). The N 2 O-saturated aqueous solutions of purine nucleoside monophosphates, containing 2-methyl-2-nitrosopropane as a spin-trap, were X-irradiated and the resulting spin-adducts were separated by reverse-phase HPLC in the ion suppression mode. The separated spin-adducts were characterized by e.s.r. spectrometry and UV spectrophotometry. Consequently, the radicals due to H-abstraction at the C4' position of the sugar moiety were identified arising from 5'-dAMP and 5'-dGMP. In all cases, e.s.r. spectra consisting of a secondary doublet were observed and assigned to the radical due to H-abstraction at the C5' position of the sugar moiety. (author)

  17. Purine (N)-Methanocarba Nucleoside Derivatives Lacking an Exocyclic Amine as Selective A3 Adenosine Receptor Agonists

    Science.gov (United States)

    2016-01-01

    Purine (N)-methanocarba-5′-N-alkyluronamidoriboside A3 adenosine receptor (A3AR) agonists lacking an exocyclic amine resulted from an unexpected reaction during a Sonogashira coupling and subsequent aminolysis. Because the initial C6-Me and C6-styryl derivatives had unexpectedly high A3AR affinity, other rigid nucleoside analogues lacking an exocyclic amine were prepared. Of these, the C6-Me-(2-phenylethynyl) and C2-(5-chlorothienylethynyl) analogues were particularly potent, with human A3AR Ki values of 6 and 42 nM, respectively. Additionally, the C2-(5-chlorothienyl)-6-H analogue was potent and selective at A3AR (MRS7220, Ki 60 nM) and also completely reversed mouse sciatic nerve mechanoallodynia (in vivo, 3 μmol/kg, po). The lack of a C6 H-bond donor while maintaining A3AR affinity and efficacy could be rationalized by homology modeling and docking of these hypermodified nucleosides. The modeling suggests that a suitable combination of stabilizing features can partially compensate for the lack of an exocyclic amine, an otherwise important contributor to recognition in the A3AR binding site. PMID:26890707

  18. Characterization of Nucleosides and Nucleobases in Natural Cordyceps by HILIC–ESI/TOF/MS and HILIC–ESI/MS

    Directory of Open Access Journals (Sweden)

    Bin Yang

    2013-08-01

    Full Text Available A method combining hydrophilic interaction chromatography (HILIC and electrospray ionization mass spectrometry (ESI-MS was developed for the characterization and determination of natural Cordyceps. Separation was achieved on a Waters Xbridge Amide column with gradient elution. Identification of 15 target nucleosides and nucleobases was based on retention time, UV spectra and mass measurements of the protonated molecules ([M+H]+ and main fragment ions (ESI-TOF/MS. Eight non-target compounds were tentatively identified by ESI-TOF/MS. The 15 target compounds were quantified by HILIC-ESI-MS/MS using time-programmed selective ion monitoring or multiple reaction monitoring in positive-ion mode under optimized mass conditions. This technique showed good linearity, repeatability and recovery. This approach was also successfully implemented in the analysis of nucleosides and nucleobases in 12 batches of natural Cordyceps samples that were collected from different regions in China. The developed HILIC-ESI-MS method exhibited clear advantages in identifying and determining highly polar bioactive components in Cordyceps, as well as their quality control.

  19. Purine nucleoside phosphorylase and xanthine oxidase activities in erythrocytes and plasma from marine, semiaquatic and terrestrial mammals.

    Science.gov (United States)

    López-Cruz, Roberto I; Pérez-Milicua, Myrna Barjau; Crocker, Daniel E; Gaxiola-Robles, Ramón; Bernal-Vertiz, Jaime A; de la Rosa, Alejandro; Vázquez-Medina, José P; Zenteno-Savín, Tania

    2014-05-01

    Purine nucleoside phosphorylase (PNP) and xanthine oxidase (XO) are key enzymes involved in the purine salvage pathway. PNP metabolizes purine bases to synthetize purine nucleotides whereas XO catalyzes the oxidation of purines to uric acid. In humans, PNP activity is reported to be high in erythrocytes and XO activity to be low in plasma; however, XO activity increases after ischemic events. XO activity in plasma of northern elephant seals has been reported during prolonged fasting and rest and voluntary associated apneas. The objective of this study was to analyze circulating PNP and XO activities in marine mammals adapted to tolerate repeated cycles of ischemia/reperfusion associated with diving (bottlenose dolphin, northern elephant seal) in comparison with semiaquatic (river otter) and terrestrial mammals (human, pig). PNP activities in plasma and erythrocytes, as well as XO activity in plasma, from all species were quantified by spectrophotometry. No clear relationship in circulating PNP or XO activity could be established between marine, semiaquatic and terrestrial mammals. Erythrocytes from bottlenose dolphins and humans are highly permeable to nucleosides and glucose, intraerythrocyte PNP activity may be related to a release of purine nucleotides from the liver. High-energy costs will probably mean a higher ATP degradation rate in river otters, as compared to northern elephant seals or dolphins. Lower erythrocyte PNP activity and elevated plasma XO activity in northern elephant seal could be associated with fasting and/or sleep- and dive-associated apneas. Copyright © 2014 Elsevier Inc. All rights reserved.

  20. Insights into the Molecular Mechanism of Polymerization and Nucleoside Reverse Transcriptase Inhibitor Incorporation by Human PrimPol.

    Science.gov (United States)

    Mislak, Andrea C; Anderson, Karen S

    2016-01-01

    Human PrimPol is a newly identified DNA and RNA primase-polymerase of the archaeo-eukaryotic primase (AEP) superfamily and only the second known polymerase in the mitochondria. Mechanistic studies have shown that interactions of the primary mitochondrial DNA polymerase γ (mtDNA Pol γ) with nucleoside reverse transcriptase inhibitors (NRTIs), key components in treating HIV infection, are a major source of NRTI-associated toxicity. Understanding the interactions of host polymerases with antiviral and anticancer nucleoside analog therapies is critical for preventing life-threatening adverse events, particularly in AIDS patients who undergo lifelong treatment. Since PrimPol has only recently been discovered, the molecular mechanism of polymerization and incorporation of natural nucleotide and NRTI substrates, crucial for assessing the potential for PrimPol-mediated NRTI-associated toxicity, has not been explored. We report for the first time a transient-kinetic analysis of polymerization for each nucleotide and NRTI substrate as catalyzed by PrimPol. These studies reveal that nucleotide selectivity limits chemical catalysis while the release of the elongated DNA product is the overall rate-limiting step. Remarkably, PrimPol incorporates four of the eight FDA-approved antiviral NRTIs with a kinetic profile distinct from that of mtDNA Pol γ that may manifest in toxicity. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  1. One-step enzymatic synthesis of nucleosides from low water-soluble purine bases in non-conventional media.

    Science.gov (United States)

    Fernández-Lucas, Jesús; Fresco-Taboada, Alba; de la Mata, Isabel; Arroyo, Miguel

    2012-07-01

    The effect of several water-miscible cosolvents on activity and stability of soluble and immobilized 2'-deoxyribosyltransferase from Lactobacillus reuteri on Sepabeads® has been studied in order to establish optimal conditions for enzymatic synthesis of nucleosides using purine bases with low solubility in aqueous buffer. As a rule of thumb, there was a general reduction of soluble enzyme activity when cosolvent content was gradually increased in reaction medium. In contrast, immobilized enzyme activity was enhanced 1.2-1.4-fold at 20% of methanol, ethanol, 2-propanol, diethylene glycol, and acetone; and at 10% and 30% acetonitrile. Likewise, highest increased activity (1.8-fold) was also obtained in presence of 20% acetonitrile. Immobilized enzyme was successfully used in the synthesis of 2'-deoxyxanthosine and 2'-deoxyguanosine using 2'-deoxyuridine as sugar donor and the corresponding poor water-soluble base in the presence of 30% of methanol, ethanol, 2-propanol, ethylene glycol, acetonitrile, and DMSO, giving high nucleoside yields at 4h. Copyright © 2011 Elsevier Ltd. All rights reserved.

  2. First-line therapy and mitochondrial damage: different nucleosides, different findings.

    Science.gov (United States)

    Blanco, Francisco; García-Benayas, Teresa; José de la Cruz, Juan; González-Lahoz, Juan; Soriano, Vincent

    2003-01-01

    Antiretroviral therapy has been associated with the development of morphologic body-shape changes and metabolic abnormalities, including dislipemia, insulin resistance, and hyperlactatemia. Mitochondrial damage secondary to the use of nucleoside analogue reverse transcriptase inhibitors (NRTIs) has been related to some of these complications, although the role of different NRTIs in their development is not well established. To assess the incidence of hyperlactatemia and lipodystrophy body-shape changes in drug-naïve HIV-infected patients who began highly active antiretroviral therapy (HAART) based on a backbone of two different NRTI combinations. Prospective, longitudinal, observational study of all consecutive drug-naïve HIV-infected individuals who started HAART with zidovudine (AZT) plus lamivudine (3TC) or didanosine (ddI) plus stavudine (d4T) between June 2000 and June 2001 at one single institution. Serum lactate levels and lipodystrophy body-shape changes were monitored periodically during 12 months. At 1 year, mean lactate values remained or=2 mmol/L) steadily increased in those on ddI+d4T (from 30% at 3 months to 71% at 12 months), whereas it remained below 10% in patients treated with AZT+3TC. Two patients on ddI+d4T developed lactic acidosis. Mean serum lactate dehydrogenase (LDH), gamma-glutamyltransferase (GGT), and amylase significantly increased in patients treated with ddI+d4T, whereas they remained unaltered in patients under AZT+3TC. Significant correlations were found between lactate and LDH, alkaline phosphatase (AP), and GGT. In the multivariate analysis, treatment with ddI+d4T, LDH, and AP was significantly associated with lactate levels. At 12 months, subcutaneous lipoatrophy was significantly more frequent in patients treated with ddI+d4T than in those on AZT+3TC (35% vs. 8%; p =.01). In drug-naïve HIV-infected patients who start antiretroviral therapy, ddI+d4T-based combinations produce a greater increase in serum lactate and lipoatrophy

  3. Tempol protects cardiomyocytes from nucleoside reverse transcriptase inhibitor-induced mitochondrial toxicity.

    Science.gov (United States)

    Liu, Yongmin; Shim, Eunwoo; Nguyen, Phuonggiang; Gibbons, Alexander T; Mitchell, James B; Poirier, Miriam C

    2014-05-01

    Nucleoside reverse transcriptase inhibitors (NRTIs), essential components of combinational therapies used for treatment of human immunodeficiency virus-1, damage heart mitochondria. Here, we have shown mitochondrial compromise in H9c2 rat cardiomyocytes exposed for 16 passages (P) to the NRTIs zidovudine (AZT, 50μM) and didanosine (ddI, 50μM), and we have demonstrated protection from mitochondrial compromise in cells treated with 200μM 1-oxyl-2,2,6,6-tetramethyl-4-hydroxypiperidine (Tempol) or 200μM 1-hydroxy-4-[2-triphenylphosphonio)-acetamido]-2,2,6,6-tetramethylpiperidine (Tempol-H), along with AZT/ddI, for 16P. Exposure to AZT/ddI caused a moderate growth inhibition at P3, P5, P7, and P13, which was not altered by addition of Tempol or Tempol-H. Mitochondrial oxidative phosphorylation capacity was determined as uncoupled oxygen consumption rate (OCR) by Seahorse XF24 Analyzer. At P5, P7, and P13, AZT/ddI-exposed cells showed an OCR reduction of 8.8-57.2% in AZT/ddI-exposed cells, compared with unexposed cells. Addition of Tempol or Tempol-H, along with AZT/ddI, resulted in OCR levels increased by about 300% above the values seen with AZT/ddI alone. The Seahorse data were further supported by electron microscopy (EM) studies in which P16 cells exposed to AZT/ddI/Tempol had less mitochondrial pathology than P16 cells exposed to AZT/ddI. Western blots of P5 cells showed that Tempol and Tempol-H upregulated expression of mitochondrial uncoupling protein-2 (UCP-2). However, Complex I activity that was reduced by AZT/ddI, was not restored in the presence of AZT/ddI/Tempol. Superoxide levels were increased in the presence of AZT/ddI and significantly decreased in cells exposed to AZT/3TC/Tempol at P3, P7, and P10. In conclusion, Tempol protects against NRTI-induced mitochondrial compromise, and UCP-2 plays a role through mild uncoupling.

  4. Mitochondrial toxicities of nucleoside analogue reverse transcriptase inhibitors in AIDS cases.

    Science.gov (United States)

    Marfatia, Yogesh S; Talwar, Mahima; Agrawal, Meetesh; Sharma, Ajay; Mehta, Kajal

    2014-01-01

    The development of antiretroviral therapy (ART) has been one of the most dramatic progressions in the history of medicine. Concomitant with this momentous therapeutic advance, the mitochondrial toxicities of ART were recognized as an important clinical entity. The aim was to study the mitochondrial toxicities in terms of peripheral neuropathy (PN), lipodystrophy (LD), hepatic steatosis, lactic academia (LA), and pancreatitis developing in AIDS cases on nucleoside analog reverse transcriptase inhibitors (NRTIs) based ART regimens. An observational study, which included 90 AIDS cases, receiving first line ART regimens containing two NRTIs (zidovudine [AZT]/stavudine [d4T] with lamivudine [3TC]) and one nonNRTIs (nevirapine/efavirenz) was conducted at Skin-VD outpatient department of a tertiary care hospital attached to a Medical College. Thorough history was taken, and clinical examination was done. Cases were subjected to measurements of abdominal girth and mid-arm circumference, liver function tests, blood sugar, lipid profile, serum lactate, and amylase levels. Of 90 cases on ART, 66% were males and 34% were females. Mitochondrial toxicities developed in 26 (30%) cases out of 90, which included 3 (7%) out of 42 cases on AZT + 3TC and 23 (48%) out of 48 cases on d4T + 3TC. Most common toxicity was PN seen in 20 (22%) cases; male cases developed PN at a lower CD4 count than female cases. LD was observed in total of 13 (14.5%) cases; deposition of fat in the abdomen in seven cases and at the nape of the neck (buffalo hump) in one case while loss of fat from extremities was seen in seven cases and loss of buccal fat in seven cases. Women presented more with fat accumulation (breast and abdomen), while men with loss of fat (limbs and buttocks). Both PN and LD were more common in d4T based regimen. LA was reported in one case on d4T. Hepatic steatosis was seen in three cases and pancreatitis in one case receiving AZT. Regular monitoring and early diagnosis of

  5. Nicotinamide riboside and nicotinic acid riboside salvage in fungi and mammals. Quantitative basis for Urh1 and purine nucleoside phosphorylase function in NAD+ metabolism.

    Science.gov (United States)

    Belenky, Peter; Christensen, Kathryn C; Gazzaniga, Francesca; Pletnev, Alexandre A; Brenner, Charles

    2009-01-02

    NAD+ is a co-enzyme for hydride transfer enzymes and an essential substrate of ADP-ribose transfer enzymes and sirtuins, the type III protein lysine deacetylases related to yeast Sir2. Supplementation of yeast cells with nicotinamide riboside extends replicative lifespan and increases Sir2-dependent gene silencing by virtue of increasing net NAD+ synthesis. Nicotinamide riboside elevates NAD+ levels via the nicotinamide riboside kinase pathway and by a pathway initiated by splitting the nucleoside into a nicotinamide base followed by nicotinamide salvage. Genetic evidence has established that uridine hydrolase, purine nucleoside phosphorylase, and methylthioadenosine phosphorylase are required for Nrk-independent utilization of nicotinamide riboside in yeast. Here we show that mammalian purine nucleoside phosphorylase but not methylthioadenosine phosphorylase is responsible for mammalian nicotinamide riboside kinase-independent nicotinamide riboside utilization. We demonstrate that so-called uridine hydrolase is 100-fold more active as a nicotinamide riboside hydrolase than as a uridine hydrolase and that uridine hydrolase and mammalian purine nucleoside phosphorylase cleave nicotinic acid riboside, whereas the yeast phosphorylase has little activity on nicotinic acid riboside. Finally, we show that yeast nicotinic acid riboside utilization largely depends on uridine hydrolase and nicotinamide riboside kinase and that nicotinic acid riboside bioavailability is increased by ester modification.

  6. Synthesis of 2'-deoxyadenosine nucleosides bearing bipyridine-type ligands and their Ru-complexes in position 8 through cross-coupling reactions

    Czech Academy of Sciences Publication Activity Database

    Vrábel, Milan; Pohl, Radek; Klepetářová, Blanka; Votruba, Ivan; Hocek, Michal

    2007-01-01

    Roč. 5, č. 17 (2007), s. 2849-2857 ISSN 1477-0520 R&D Projects: GA MŠk LC512; GA ČR GA203/05/0043 Institutional research plan: CEZ:AV0Z40550506 Keywords : nucleosides * purines * cross-coupling * ruthenium Subject RIV: CC - Organic Chemistry Impact factor: 3.167, year: 2007

  7. Novel (2,6-difluorophenyl)(2-(phenylamino)pyrimidin-4-yl) methanones with restricted conformation as potent non-nucleoside reverse transcriptase inhibitors against HIV-1

    Czech Academy of Sciences Publication Activity Database

    Šimon, Petr; Baszczyňski, Ondřej; Šaman, David; Stepan, G.; Hu, E.; Lansdon, E. B.; Jansa, P.; Janeba, Zlatko

    2016-01-01

    Roč. 122, Oct 21 (2016), s. 185-195 ISSN 0223-5234 Institutional support: RVO:61388963 Keywords : diarylpyrimidine (DAPY) * etravirine * human immunodeficiency virus (HIV) * non-nucleoside reverse transcriptase inhibitors * NNRTIs * rilpivirine Subject RIV: CC - Organic Chemistry Impact factor: 4.519, year: 2016

  8. Cytostatic 6-arylpurine nucleosides III. Synthesis and structure-activity relationship study in cytostatic activity of 6-aryl-, 6-hetaryl- and 6-benzylpurine ribonucleosides

    Czech Academy of Sciences Publication Activity Database

    Hocek, Michal; Holý, Antonín; Votruba, Ivan; Dvořáková, H.

    2001-01-01

    Roč. 66, č. 3 (2001), s. 483-499 ISSN 0010-0765 R&D Projects: GA ČR GV203/96/K001; GA ČR GA203/00/0036 Institutional research plan: CEZ:AV0Z4055905 Keywords : purines * nucleosides Subject RIV: CC - Organic Chemistry Impact factor: 0.778, year: 2001

  9. HILIC-UPLC-MS/MS combined with hierarchical clustering analysis to rapidly analyze and evaluate nucleobases and nucleosides in Ginkgo biloba leaves.

    Science.gov (United States)

    Yao, Xin; Zhou, Guisheng; Tang, Yuping; Guo, Sheng; Qian, Dawei; Duan, Jin-Ao

    2015-02-01

    Ginkgo biloba leaf extract has been widely used in dietary supplements and more recently in some foods and beverages. In addition to the well-known flavonol glycosides and terpene lactones, G. biloba leaves are also rich in nucleobases and nucleosides. To determine the content of nucleobases and nucleosides in G. biloba leaves at trace levels, a reliable method has been established by using hydrophilic interaction ultra performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry (HILIC-UPLC-TQ-MS/MS) working in multiple reaction monitoring mode. Eleven nucleobases and nucleosides were simultaneously determined in seven min. The proposed method was fully validated in terms of linearity, sensitivity, and repeatability, as well as recovery. Furthermore, hierarchical clustering analysis (HCA) was performed to evaluate and classify the samples according to the contents of the eleven chemical constituents. The established approach could be helpful for evaluation of the potential values as dietary supplements and the quality control of G. biloba leaves, which might also be utilized for the investigation of other medicinal herbs containing nucleobases and nucleosides. Copyright © 2014 John Wiley & Sons, Ltd.

  10. Antiviral activities of 2,6-diaminopurine-based acyclic nucleoside phosphonates against herpesviruses: In vitro study results with pseudorabies virus (PrV, SuHV-1)

    Czech Academy of Sciences Publication Activity Database

    Zouharová, D.; Lipenská, I.; Fojtiková, M.; Kulich, P.; Neca, J.; Slaný, M.; Kovařčík, K.; Turanek-Knotigová, P.; Hubatka, F.; Celechovská, H.; Mašek, J.; Koudelka, Š.; Procházka, L.; Eyer, L.; Plocková, J.; Bartheldyová, E.; Miller, A. D.; Růžek, Daniel; Raška, M.; Janeba, Zlatko; Turánek, J.

    2016-01-01

    Roč. 184, FEB 29 (2016), s. 84-93 ISSN 0378-1135 Institutional support: RVO:60077344 ; RVO:61388963 Keywords : Pseudorabies * Acyclic nucleoside phosphonates * DNA viruses * Cidofovir * Anti viral drugs * DNA polymerase Subject RIV: EE - Microbiology, Virology; CC - Organic Chemistry (UOCHB-X) Impact factor: 2.628, year: 2016

  11. Familial dysautonomia (FD) patients have reduced levels of the modified wobble nucleoside mcm(5)s(2)U in tRNA.

    Science.gov (United States)

    Karlsborn, Tony; Tükenmez, Hasan; Chen, Changchun; Byström, Anders S

    2014-11-21

    Familial dysautonomia (FD) is a recessive neurodegenerative genetic disease. FD is caused by a mutation in the IKBKAP gene resulting in a splicing defect and reduced levels of full length IKAP protein. IKAP homologues can be found in all eukaryotes and are part of a conserved six subunit protein complex, Elongator complex. Inactivation of any Elongator subunit gene in multicellular organisms cause a wide range of phenotypes, suggesting that Elongator has a pivotal role in several cellular processes. In yeast, there is convincing evidence that the main role of Elongator complex is in formation of modified wobble uridine nucleosides in tRNA and that their absence will influence translational efficiency. To date, no study has explored the possibility that FD patients display defects in formation of modified wobble uridine nucleosides as a consequence of reduced IKAP levels. In this study, we show that brain tissue and fibroblast cell lines from FD patients have reduced levels of the wobble uridine nucleoside 5-methoxycarbonylmethyl-2-thiouridine (mcm(5)s(2)U). Our findings indicate that FD could be caused by inefficient translation due to lower levels of wobble uridine nucleosides. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  12. A novel and efficient one-pot synthesis of symmetrical diamide (bis-amidate) prodrugs of acyclic nucleoside phosphonates and evaluation of their biological activities

    Czech Academy of Sciences Publication Activity Database

    Jansa, Petr; Baszczyňski, Ondřej; Dračínský, Martin; Votruba, Ivan; Zídek, Zdeněk; Bahador, G.; Stepan, G.; Cihlar, T.; Mackman, R.; Holý, Antonín; Janeba, Zlatko

    2011-01-01

    Roč. 46, č. 9 (2011), s. 3748-3754 ISSN 0223-5234 R&D Projects: GA MV VG20102015046 Institutional research plan: CEZ:AV0Z40550506; CEZ:AV0Z50390703 Keywords : prodrugs * phosphonodiamides * bis-amidates * acyclic nucleoside phosphonates * GS-9219 Subject RIV: CC - Organic Chemistry Impact factor: 3.346, year: 2011

  13. Synthesis and antiviral activities of hexadecyloxypropyl prodrugs of acyclic nucleoside phosphonates containing guanine or hypoxanthine and a (S)-HPMP or PEE acyclic moiety

    Czech Academy of Sciences Publication Activity Database

    Tichý, Tomáš; Andrei, G.; Snoeck, R.; Balzarini, J.; Dračínský, Martin; Krečmerová, Marcela

    2012-01-01

    Roč. 55, Sep (2012), s. 307-314 ISSN 0223-5234 R&D Projects: GA ČR GAP207/11/0108; GA MV VG20102015046 Institutional support: RVO:61388963 Keywords : antivirals * prodrugs * acyclic nucleoside phosphonate * phosphonate ester Subject RIV: CC - Organic Chemistry Impact factor: 3.499, year: 2012

  14. Synthesis and biological activity of 5-azacytosine nucleosides derived from 4-thio-2-deoxy-L-threo-pentofuranose and 4-thio-2-deoxy-D-erythro-pentofuranose.

    Science.gov (United States)

    Cappellacci, L; Tiwari, K N; Montgomery, J A; Secrist, J A

    1999-01-01

    1-O-Acetyl-2-deoxy-3,5-di-O-toluoyl-4-thio-D-erythro-pentofuranose and 2-deoxy-1,3,5-tri-O-acetyl-4-thio-L-threo-pentofuranose were coupled with 5-azacytosine to obtain alpha and beta anomers of nucleosides.

  15. Co- and homocyclotrimerization reactions of protected 1-alkynyl-2-deoxyribofuranose. Synthesis of C-nucleosides, C-di- and C-trisaccharide analogues

    Czech Academy of Sciences Publication Activity Database

    Novák, P.; Číhalová, S.; Otmar, Miroslav; Hocek, Michal; Kotora, Martin

    2008-01-01

    Roč. 64, č. 22 (2008), s. 5200-5207 ISSN 0040-4020 R&D Projects: GA MŠk 1M0508 Institutional research plan: CEZ:AV0Z40550506 Keywords : alkynes * cyclotrimerizations * nucleosides Subject RIV: CC - Organic Chemistry Impact factor: 2.897, year: 2008

  16. UHPLC-TQ-MS Coupled with Multivariate Statistical Analysis to Characterize Nucleosides, Nucleobases and Amino Acids in Angelicae Sinensis Radix Obtained by Different Drying Methods.

    Science.gov (United States)

    Zhu, Shaoqing; Guo, Sheng; Duan, Jin-Ao; Qian, Dawei; Yan, Hui; Sha, Xiuxiu; Zhu, Zhenhua

    2017-06-01

    To explore the nutrients in roots of Angelica sinensis (Angelicae Sinensis Radix, ASR), a medicinal and edible plant, and evaluate its nutritional value, a rapid and reliable UHPLC-TQ-MS method was established and used to determine the potential nutritional compounds, including nucleosides, nucleobases and amino acids, in 50 batches of ASR samples obtained using two drying methods. The results showed that ASR is a healthy food rich in nucleosides, nucleobases and amino acids, especially arginine. The total average content of nucleosides and nucleobases in all ASR samples was 3.94 mg/g, while that of amino acids reached as high as 61.79 mg/g. Principle component analysis showed that chemical profile differences exist between the two groups of ASR samples prepared using different drying methods, and the contents of nutritional compounds in samples dried with the tempering-intermittent drying processing method (TIDM) were generally higher than those dried using the traditional solar processing method. The above results suggest that ASR should be considered an ideal healthy food and TIDM could be a suitable drying method for ASR when taking nucleosides, nucleobases and amino acids as the major consideration for their known human health benefits.

  17. A quantitative histochemical procedure for the demonstration of purine nucleoside phosphorylase activity in rat and human liver using Tetranitro BT and xanthine oxidase as auxiliary enzyme

    NARCIS (Netherlands)

    Frederiks, W. M.; Bosch, K. S.; van Gulik, T.

    1993-01-01

    A quantitative histochemical procedure was developed for the demonstration of purine nucleoside phosphorylase in rat liver using unfixed cryostat sections and the auxiliary enzyme xanthine oxidase. The optimum incubation medium contained 18% (w/v) poly(vinyl alcohol), 100 mM phosphate buffer, pH

  18. Synthesis of Nucleosides through Direct Glycosylation of Nucleobases with 5-O-Monoprotected or 5-Modified Ribose: Improved Protocol, Scope, and Mechanism

    Czech Academy of Sciences Publication Activity Database

    Downey, Alan Michael; Pohl, Radek; Roithová, J.; Hocek, Michal

    2017-01-01

    Roč. 23, č. 16 (2017), s. 3910-3917 ISSN 0947-6539 R&D Projects: GA TA ČR(CZ) TE01020028; GA ČR(CZ) GA16-00178S Institutional support: RVO:61388963 Keywords : epoxides * glycosylation * nucleosides * riboses * synthesis design Subject RIV: CC - Organic Chemistry OBOR OECD: Organic chemistry Impact factor: 5.317, year: 2016

  19. Inhibition of nucleoside diphosphate kinase activity by in vitro phosphorylation by protein kinase CK2. Differential phosphorylation of NDP kinases in HeLa cells in culture

    DEFF Research Database (Denmark)

    Biondi, R M; Engel, M; Sauane, M

    1996-01-01

    Although a number of nucleoside diphosphate kinases (NDPKs) have been reported to act as inhibitors of metastasis or as a transcription factor in mammals, it is not known whether these functions are linked to their enzymatic activity or how this protein is regulated. In this report, we show that ...

  20. Pan-pathway based interaction profiling of FDA-approved nucleoside and nucleobase analogs with enzymes of the human nucleotide metabolism.

    Directory of Open Access Journals (Sweden)

    Louise Egeblad

    Full Text Available To identify interactions a nucleoside analog library (NAL consisting of 45 FDA-approved nucleoside analogs was screened against 23 enzymes of the human nucleotide metabolism using a thermal shift assay. The method was validated with deoxycytidine kinase; eight interactions known from the literature were detected and five additional interactions were revealed after the addition of ATP, the second substrate. The NAL screening gave relatively few significant hits, supporting a low rate of "off target effects." However, unexpected ligands were identified for two catabolic enzymes guanine deaminase (GDA and uridine phosphorylase 1 (UPP1. An acyclic guanosine prodrug analog, valaciclovir, was shown to stabilize GDA to the same degree as the natural substrate, guanine, with a ΔT(agg around 7°C. Aciclovir, penciclovir, ganciclovir, thioguanine and mercaptopurine were also identified as ligands for GDA. The crystal structure of GDA with valaciclovir bound in the active site was determined, revealing the binding of the long unbranched chain of valaciclovir in the active site of the enzyme. Several ligands were identified for UPP1: vidarabine, an antiviral nucleoside analog, as well as trifluridine, idoxuridine, floxuridine, zidovudine, telbivudine, fluorouracil and thioguanine caused concentration-dependent stabilization of UPP1. A kinetic study of UPP1 with vidarabine revealed that vidarabine was a mixed-type competitive inhibitor with the natural substrate uridine. The unexpected ligands identified for UPP1 and GDA imply further metabolic consequences for these nucleoside analogs, which could also serve as a starting point for future drug design.

  1. Synthesis and Evaluation of the Biological Profile of Novel Analogues of Nucleosides and of Potential Mimetics of Sugar Phosphates and Nucleotides

    Czech Academy of Sciences Publication Activity Database

    Xavier, N.M.; Lucas, S.D.; Jorda, Radek; Schwarz, S.; Loesche, A.; Csuk, R.; Oliveira, M.C.

    2015-01-01

    Roč. 26, č. 19 (2015), s. 2663-2672 ISSN 0936-5214 R&D Projects: GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : nucleosides * nucleotides * carbohydrates Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.323, year: 2015

  2. Synthesis of enantiomerically pure (purin-6-yl)phenylalanines and their nucleosides, a novel type of purine-amino acid conjugates

    Czech Academy of Sciences Publication Activity Database

    Čapek, Petr; Pohl, Radek; Hocek, Michal

    2005-01-01

    Roč. 70, č. 20 (2005), s. 8001-8008 ISSN 0022-3263 R&D Projects: GA ČR(CZ) GA203/03/0035; GA MŠk(CZ) 1M0508 Institutional research plan: CEZ:AV0Z40550506 Keywords : purin es * nucleosides * amino acids Subject RIV: CC - Organic Chemistry Impact factor: 3.675, year: 2005

  3. Synthesis, cytostatic and anti-HCV activity of 6-(N-substituted aminomethyl)-, 6-(O-substituted hydroxymethyl)- and 6-(S-substituted sulfanylmethyl)purine nucleosides

    Czech Academy of Sciences Publication Activity Database

    Šilhár, Peter; Hocek, Michal; Pohl, Radek; Votruba, Ivan; Shih, I.; Mabery, E.; Mackman, R.

    2008-01-01

    Roč. 16, č. 5 (2008), s. 2329-2366 ISSN 0968-0896 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : purin es * nucleosides * cytostatics * antivirals Subject RIV: CC - Organic Chemistry Impact factor: 3.075, year: 2008

  4. Viral resuppression and detection of drug resistance following interruption of a suppressive non-nucleoside reverse transcriptase inhibitor-based regimen

    DEFF Research Database (Denmark)

    Fox, Zoe; Phillips, Andrew; Cohen, Cal

    2008-01-01

    BACKGROUND: Interruption of a non-nucleoside reverse transcriptase inhibitor (NNRTI)-regimen is often necessary, but must be performed with caution because NNRTIs have a low genetic barrier to resistance. Limited data exist to guide clinical practice on the best interruption strategy to use...

  5. Multi-nucleoside reverse transcriptase inhibitor resistant HIV type-1 in a patient from Sierra Leone failing stavudine, lamivudine and nevirapine

    NARCIS (Netherlands)

    Hamers, Raph L.; Wensing, Annemarie M. J.; Back, Nicole K. T.; Arcilla, Maria S.; Frissen, Jos P. H.

    2011-01-01

    We report a 33-year-old HIV type-1 (HIV-1)-infected male from Sierra Leone who harboured extensive drug resistance mutations to all nucleoside reverse transcriptase inhibitors (NRTIs) and non-NRTIs, including the multi-NRTI-resistance Q151M complex, K65R, M184I and Y181I, after using standard

  6. Bone mineral density changes in protease inhibitor-sparing vs. nucleoside reverse transcriptase inhibitor-sparing highly active antiretroviral therapy: data from a randomized trial

    DEFF Research Database (Denmark)

    Hansen, Ab; Obel, N; Nielsen, H

    2011-01-01

    Objective The aim of the study was to compare changes in bone mineral density (BMD) over 144 weeks in HIV-infected patients initiating nucleoside reverse transcriptase inhibitor (NRTI)-sparing or protease inhibitor-sparing highly active antiretroviral therapy (HAART). Methods Sixty-three HAART...

  7. A systematic review of a single-class maintenance strategy with nucleoside/nucleotide reverse transcriptase inhibitors in HIV/AIDS

    NARCIS (Netherlands)

    Sprenger, Herman G.; Bierman, Wouter F. W.; van der Werf, Tjip S.; Gisolf, Elisabeth H.; Richter, Clemens

    2014-01-01

    Background: Single-drug class regimens with nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs) are generally not recommended as initial therapy because they are inferior compared with therapy with two NRTIs plus efavirenz. However, triple-NRTI combinations can be useful in specific

  8. Interpatient variability in the pharmacokinetics of the HIV non-nucleoside reverse transcriptase inhibitor efavirenz : the effect of gender, race, and CYP2B6 polymorphism

    NARCIS (Netherlands)

    Burger, D; van der Heiden, [No Value; la Porte, C; van der Ende, Marchina E.; Groeneveld, P; Richter, C; Koopmans, P; Sprenger, H; Lindemans, J; Schenk, P; van Schaik, R

    Aims To characterize the demographic and pharmacogenetic factors that influence interpatient variability in the plasma concentrations of the HIV non-nucleoside reverse transcriptase inhibitor efavirenz. Methods Data from all samples analyzed for efavirenz in our TDM service in 2002 and 2003 were

  9. The synthesis of double-headed nucleosides by the CuAAC reaction and their effect in secondary nucleic acid structures

    DEFF Research Database (Denmark)

    Jørgensen, Anna Søndergaard; Shaikh, Khalil Isak; Enderlin, Gerald

    2011-01-01

    Four double-headed nucleosides were prepared by the CuAAC reaction. Hereby, a triazole-containing linker connects an additional thymine or adenine to the 2´-position of 2´-deoxyuridine, a thymine to the 5´-position of thymidine and a thymine to the 6¢-position of an LNA-thymidine monomer. Whereas...

  10. Pan-pathway based interaction profiling of FDA-approved nucleoside and nucleobase analogs with enzymes of the human nucleotide metabolism.

    Science.gov (United States)

    Egeblad, Louise; Welin, Martin; Flodin, Susanne; Gräslund, Susanne; Wang, Liya; Balzarini, Jan; Eriksson, Staffan; Nordlund, Pär

    2012-01-01

    To identify interactions a nucleoside analog library (NAL) consisting of 45 FDA-approved nucleoside analogs was screened against 23 enzymes of the human nucleotide metabolism using a thermal shift assay. The method was validated with deoxycytidine kinase; eight interactions known from the literature were detected and five additional interactions were revealed after the addition of ATP, the second substrate. The NAL screening gave relatively few significant hits, supporting a low rate of "off target effects." However, unexpected ligands were identified for two catabolic enzymes guanine deaminase (GDA) and uridine phosphorylase 1 (UPP1). An acyclic guanosine prodrug analog, valaciclovir, was shown to stabilize GDA to the same degree as the natural substrate, guanine, with a ΔT(agg) around 7°C. Aciclovir, penciclovir, ganciclovir, thioguanine and mercaptopurine were also identified as ligands for GDA. The crystal structure of GDA with valaciclovir bound in the active site was determined, revealing the binding of the long unbranched chain of valaciclovir in the active site of the enzyme. Several ligands were identified for UPP1: vidarabine, an antiviral nucleoside analog, as well as trifluridine, idoxuridine, floxuridine, zidovudine, telbivudine, fluorouracil and thioguanine caused concentration-dependent stabilization of UPP1. A kinetic study of UPP1 with vidarabine revealed that vidarabine was a mixed-type competitive inhibitor with the natural substrate uridine. The unexpected ligands identified for UPP1 and GDA imply further metabolic consequences for these nucleoside analogs, which could also serve as a starting point for future drug design.

  11. An efficient method for the construction of functionalized DNA bearing amino acid groups through cross-coupling reactions of nucleoside triphosphates followed by primer extension or PCR

    Czech Academy of Sciences Publication Activity Database

    Čapek, Petr; Cahová, Hana; Pohl, Radek; Hocek, Michal; Gloeckner, Ch.; Marx, A.

    2007-01-01

    Roč. 13, č. 21 (2007), s. 6196-6203 ISSN 0947-6539 R&D Projects: GA MŠk LC512; GA ČR GA203/05/0043 Institutional research plan: CEZ:AV0Z40550506 Keywords : nucleoside triphosphates * cross-coupling * DNA Subject RIV: CC - Organic Chemistry Impact factor: 5.330, year: 2007

  12. The role of human equilibrative nucleoside transporter 1 on the cellular transport of the DNA methyltransferase inhibitors 5-azacytidine and CP-4200 in human leukemia cells.

    Science.gov (United States)

    Hummel-Eisenbeiss, Johanna; Hascher, Antje; Hals, Petter-Arnt; Sandvold, Marit Liland; Müller-Tidow, Carsten; Lyko, Frank; Rius, Maria

    2013-09-01

    The nucleoside analog 5-azacytidine is an archetypical drug for epigenetic cancer therapy, and its clinical effectiveness has been demonstrated in the treatment of myelodysplastic syndromes (MDS) and acute myelogenous leukemia (AML). However, therapy resistance in patients with MDS/AML remains a challenging issue. Membrane proteins that are involved in drug uptake are potential mediators of drug resistance. The responsible proteins for the transport of 5-azacytidine into MDS/AML cells are unknown. We have now systematically analyzed the expression and activity of various nucleoside transporters. We identified the human equilibrative nucleoside transporter 1 (hENT1) as the most abundant nucleoside transporter in leukemia cell lines and in AML patient samples. Transport assays using [¹⁴C]5-azacytidine demonstrated Na⁺-independent uptake of the drug into the cells, which was inhibited by S-(4-nitrobenzyl)-6-thioinosine (NBTI), a hENT1 inhibitor. The cellular toxicity of 5-azacytidine and its DNA demethylating activity were strongly reduced after hENT1 inhibition. In contrast, the cellular activity of the 5-azacytidine derivative 5-azacytidine-5'-elaidate (CP-4200), a nucleoside transporter-independent drug, persisted after hENT1 inhibition. A strong dependence of 5-azacytidine-induced DNA demethylation on hENT1 activity was also confirmed by array-based DNA methylation profiling, which uncovered hundreds of loci that became demethylated only when hENT1-mediated transport was active. Our data establish hENT1 as a key transporter for the cellular uptake of 5-azacytidine in leukemia cells and raise the possibility that hENT1 expression might be a useful biomarker to predict the efficiency of 5-azacytidine treatments. Furthermore, our data suggest that CP-4200 may represent a valuable compound for the modulation of transporter-related 5-azacytidine resistances.

  13. Carbon-Based Functional Materials Derived from Waste for Water Remediation and Energy Storage.

    Science.gov (United States)

    Ma, Qinglang; Yu, Yifu; Sindoro, Melinda; Fane, Anthony G; Wang, Rong; Zhang, Hua

    2017-04-01

    Carbon-based functional materials hold the key for solving global challenges in the areas of water scarcity and the energy crisis. Although carbon nanotubes (CNTs) and graphene have shown promising results in various fields of application, their high preparation cost and low production yield still dramatically hinder their wide practical applications. Therefore, there is an urgent call for preparing carbon-based functional materials from low-cost, abundant, and sustainable sources. Recent innovative strategies have been developed to convert various waste materials into valuable carbon-based functional materials. These waste-derived carbon-based functional materials have shown great potential in many applications, especially as sorbents for water remediation and electrodes for energy storage. Here, the research progress in the preparation of waste-derived carbon-based functional materials is summarized, along with their applications in water remediation and energy storage; challenges and future research directions in this emerging research field are also discussed. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Imaging expression of adenoviral HSV1-tk suicide gene transfer using the nucleoside analogue FIRU

    International Nuclear Information System (INIS)

    Nanda, Dharmin; Jong, Marion de; Bakker, Willem; Bijster, Magda; Cox, Peter; Vogels, Ronald; Havenga, Menzo; Driesse, Maarten; Avezaat, Cees; Morin, Kevin; Naimi, Ebrahim; Knaus, Edward; Wiebe, Leonard; Smitt, Peter Sillevis

    2002-01-01

    Substrates for monitoring HSV1-tk gene expression include uracil and acycloguanosine derivatives.The most commonly used uracil derivative to monitor HSV1-tk gene transfer is 1-(2-fluoro-2-deoxy-β-D-arabinofuranosyl)-5-[*I]iodouracil (fialuridine; I*-FIAU), where the asterisk denotes any of the radioactive iodine isotopes that can be used. We have previously studied other nucleosides with imaging properties as good as or better than FIAU, including 1-(2-fluoro-2-deoxy-β-D-ribofuranosyl)-5-[*I]iodouracil (FIRU). The first aim of this study was to extend the biodistribution data of 123 I-labelled FIRU. Secondly, we assessed the feasibility of detecting differences in HSV1-tk gene expression levels following adenoviral gene transfer in vivo with 123 I-FIRU. 9L rat gliosarcoma cells were stably transfected with the HSV1-tk gene (9L-tk+). 123 I-FIRU was prepared by radioiodination of 1-(2-fluoro-2-deoxy-β-D-ribofuranosyl)-5-tributylstannyl uracil (FTMRSU; precursor compound) and purified using an activated Sep-Pak column. Incubation of 9L-tk+ cells and the parental 9L cells with 123 I-FIRU resulted in a 100-fold higher accumulation of radioactivity in the 9L-tk+ cells after an optimum incubation time of 4 h. NIH-bg-nu-xid mice were then inoculated subcutaneously with HSV1-tk (-) 9L cells or HSV1-tk (+) 9L-tk+ cells into both flanks. Biodistribution studies and gamma camera imaging were performed at 15 min and 1, 2, 4 and 24 h p.i. At 15 min, the tumour/muscle, tumour/blood and tumour/brain ratios were 5.2, 1.0 and 30.3 respectively. Rapid renal clearance of the tracer from the body resulted in increasing tumour/muscle, tumour/blood and tumour/brain ratios, reaching values of 32.2, 12.5 and 171.6 at 4 h p.i. A maximum specific activity of 22%ID/g tissue was reached in the 9L-tk+ tumours 4 h after 123 I-FIRU injection. Two Ad5-based adenoviral vectors containing the HSV1-tk gene were constructed: a replication-incompetent vector with the transgene in the former E1

  15. Antiviral therapy with nucleotide/nucleoside analogues in chronic hepatitis B: A meta-analysis of prospective randomized trials.

    Science.gov (United States)

    Bedre, Renesh H; Raj, Utkarsh; Misra, Sri Prakash; Varadwaj, Pritish Kumar

    2016-03-01

    Nucleotide/nucleoside analogues (antiviral therapy) are used in the therapy of HBeAg positive and HBeAg negative chronic hepatitis B. We analyzed ten selected randomized controlled with 2557 patients to estimate the effect of antiviral drugs in chronic hepatitis B with compared to placebo. Virological response, biochemical response, histological response, seroconversion of HBeAg, and loss of HBeAg were estimated as primary efficacy measures. The included studies were subjected for heterogeneity and publication bias. The heterogeneity was assessed with χ2 and I(2) statistics. Publication bias was assessed by funnel plot. Greater rates of improvement obtained in antiviral group for virological response [43.96 % vs. 3.15 %, RR = 0.57, 95 % CI = 0.54-0.61, p-value Antiviral therapy provided significant benefit for the treatment of chronic hepatitis B with no measurable adverse effects.

  16. Separation of oligopeptides, nucleobases, nucleosides and nucleotides using capillary electrophoresis/electrochromatography with sol-gel modified inner capillary wall.

    Science.gov (United States)

    Svobodová, Jana; Kofroňová, Olga; Benada, Oldřich; Král, Vladimír; Mikšík, Ivan

    2017-09-29

    The aim of this article is to study the modification of an inner capillary wall with sol-gel coating (pure silica sol-gel or silica sol-gel containing porphyrin-brucine conjugate) and determine its influence on the separation process using capillary electrophoresis/electrochromatography method. After modification of the inner capillary surface the separation of analytes was performed using two different phosphate buffers (pH 2.5 and 9.0) and finally the changes in electrophoretic mobilities of various samples were calculated. To confirm that the modification of the inner capillary surface was successful, the parts of the inner surfaces of capillaries were observed using scanning electron microscopy. The analytes used as testing samples were oligopeptides, nucleosides, nucleobases and finally nucleotides. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. ANN QSAR workflow for predicting the inhibition of HIV-1 reverse transcriptase by pyridinone non-nucleoside derivatives.

    Science.gov (United States)

    Barzegar, Abolfazl; Zamani-Gharehchamani, Elham; Kadkhodaie-Ilkhchi, Ali

    2017-07-01

    Pyridinone derivatives have high potency against non-nucleoside reverse transcriptase inhibitor (NNRTI)-resistant human immunodeficiency virus type-1 strains. Quantitative structure-activity relationship (QSAR) studies on a series of pyridinone derivatives acting as NNRTIs are very important in designing the next generation of NNRTIs. Methodology & results: The QSAR models were developed using linear (single and forward stepwise) and combined nonlinear artificial neural network (ANN) approaches. ANN provided QSAR model with highly correlating values of 0.963, 0.964, 0.920 and 0.917, corresponding to the biological activity pIC 50 of the training, validation, testing and all samples, respectively. The nonlinear ANN-QSAR model based on the topological polarizability, geometrical steric, hydrophobicity and substituted benzene functional group indices might be able to help for designing novel pyridinone NNRTIs.

  18. Identification and Interrogation of the Herbicidin Biosynthetic Gene Cluster: First Insight into the Biosynthesis of a Rare Undecose Nucleoside Antibiotic.

    Science.gov (United States)

    Lin, Geng-Min; Romo, Anthony J; Liem, Priscilla H; Chen, Zhang; Liu, Hung-Wen

    2017-11-22

    Herbicidins are adenosine-based nucleoside antibiotics with an unusual tricyclic undecose core decorated with a (5-hydroxy)tiglyl moiety. Feeding studies are herein reported demonstrating that the tricyclic core is derived from d-glucose and d-ribose, whereas the tiglyl moiety is derived from an intermediate of l-isoleucine catabolism. Identification of the gene cluster for herbicidin A biosynthesis in Streptomyces sp. L-9-10 as well as its verification by heterologous expression in a nonproducing host are described, and the results of in vitro characterization of a carboxyl methyltransferase encoded in the cluster, Her8, are presented. Based on these observations, a biosynthetic pathway is proposed for herbicidins.

  19. The regulatory effect of nucleoside diphosphate kinase on G-protein and G-protein mediated phospholipase C.

    Science.gov (United States)

    Zhang, D; Chang, K

    1995-03-01

    The effect of nucleoside diphosphate kinase (NDPK) on the activity of guanine nucleotide regulatory protein (G-protein) mediated phospholipase C (PLC) and on the [35S] GTPT tau S binding of G-protein was investigated in this work in order to demonstrate the mechanism behind the regulation of G-protein and its effector PLC by NDPK. The stimulation of PLC in turkey erythrocyte membrane by both GTP and GTP tau S indicated that the PLC stimulation was mediated by G-protein. NDPK alone stimulated PLC activity, as well as the stimulation in the presence of GTP and GDP, in a dose-dependent manner. However, NDPK inhibited GTP tau S-stimulated PLC. Furthermore, NDPK inhibited [35S]GTP tau S binding of purified Gi-protein in a non-competitive manner. A hypothesis implying an important role of direct interaction of G-protein and NDPK in the regulation of their functions is suggested and discussed.

  20. Rilpivirine, a novel non-nucleoside reverse transcriptase inhibitor for the management of HIV-1 infection: a systematic review.

    Science.gov (United States)

    Schafer, Jason J; Short, William R

    2012-01-01

    Rilpivirine (RPV) is a second-generation non-nucleoside reverse transcriptase inhibitor (NNRTI). It remains active against HIV strains harbouring mutations that affect first-generation agents. RPV is dosed once daily with food and has been coformulated into a single tablet containing tenofovir and emtricitabine. Two Phase III studies of treatment-naive patients found RPV and efavirenz to have similar safety and efficacy. However, suboptimal virological suppression with RPV occurred more commonly in patients with higher baseline viral loads (>100,000 copies/ml). The most common mutation that emerged during RPV therapy was E138K, which often occurred in combination with M184I. E138K is likely to cause cross-resistance to other NNRTIs thereby limiting the further utilization of this class.

  1. N(3)-protection of thymidine with Boc for an easy synthetic access to sugar-alkylated nucleoside analogs.

    Science.gov (United States)

    Simeone, Luca; De Napoli, Lorenzo; Montesarchio, Daniela

    2012-03-01

    The use of Boc as a nucleobase-protecting group in the synthesis of sugar-modified thymidine analogs is reported. Boc was easily inserted at N(3) by a simple and high-yielding reaction and found to be stable to standard treatments for the removal of Ac and (t) BuMe(2) Si (TBDMS) groups, as well as to ZnBr(2) -mediated 4,4'-dimethoxytrityl (DMTr) deprotection. Boc Protection proved to be completely resistant to the strong basic conditions required to regioselectively achieve O-alkylation, therefore, providing synthetic access to a variety of sugar-alkylated nucleoside analogs. To demonstrate the feasibility of this approach, two 3'-O-alkylated thymidine analogs have been synthesized in high overall yields and fully characterized. Copyright © 2012 Verlag Helvetica Chimica Acta AG, Zürich.

  2. HIV-1 polymerase inhibition by nucleoside analogs: cellular- and kinetic parameters of efficacy, susceptibility and resistance selection.

    Directory of Open Access Journals (Sweden)

    Max von Kleist

    2012-01-01

    Full Text Available Nucleoside analogs (NAs are used to treat numerous viral infections and cancer. They compete with endogenous nucleotides (dNTP/NTP for incorporation into nascent DNA/RNA and inhibit replication by preventing subsequent primer extension. To date, an integrated mathematical model that could allow the analysis of their mechanism of action, of the various resistance mechanisms, and their effect on viral fitness is still lacking. We present the first mechanistic mathematical model of polymerase inhibition by NAs that takes into account the reversibility of polymerase inhibition. Analytical solutions for the model point out the cellular- and kinetic aspects of inhibition. Our model correctly predicts for HIV-1 that resistance against nucleoside analog reverse transcriptase inhibitors (NRTIs can be conferred by decreasing their incorporation rate, increasing their excision rate, or decreasing their affinity for the polymerase enzyme. For all analyzed NRTIs and their combinations, model-predicted macroscopic parameters (efficacy, fitness and toxicity were consistent with observations. NRTI efficacy was found to greatly vary between distinct target cells. Surprisingly, target cells with low dNTP/NTP levels may not confer hyper-susceptibility to inhibition, whereas cells with high dNTP/NTP contents are likely to confer natural resistance. Our model also allows quantification of the selective advantage of mutations by integrating their effects on viral fitness and drug susceptibility. For zidovudine triphosphate (AZT-TP, we predict that this selective advantage, as well as the minimal concentration required to select thymidine-associated mutations (TAMs are highly cell-dependent. The developed model allows studying various resistance mechanisms, inherent fitness effects, selection forces and epistasis based on microscopic kinetic data. It can readily be embedded in extended models of the complete HIV-1 reverse transcription process, or analogous processes

  3. Enzymatic properties of an ecto-nucleoside triphosphate diphosphohydrolase from Legionella pneumophila: substrate specificity and requirement for virulence.

    Science.gov (United States)

    Sansom, Fiona M; Riedmaier, Patrice; Newton, Hayley J; Dunstone, Michelle A; Müller, Christa E; Stephan, Holger; Byres, Emma; Beddoe, Travis; Rossjohn, Jamie; Cowan, Peter J; d'Apice, Anthony J F; Robson, Simon C; Hartland, Elizabeth L

    2008-05-09

    Legionella pneumophila is the predominant cause of Legionnaires disease, a severe and potentially fatal form of pneumonia. Recently, we identified an ecto-nucleoside triphosphate diphosphohydrolase (NTPDase) from L. pneumophila, termed Lpg1905, which enhances intracellular replication of L. pneumophila in eukaryotic cells. Lpg1905 is the first prokaryotic member of the CD39/NTPDase1 family of enzymes, which are characterized by the presence of five apyrase conserved regions and the ability to hydrolyze nucleoside tri- and diphosphates. Here we examined the substrate specificity of Lpg1905 and showed that apart from ATP and ADP, the enzyme catalyzed the hydrolysis of GTP and GDP but had limited activity against CTP, CDP, UTP, and UDP. Based on amino acid residues conserved in the apyrase conserved regions of eukaryotic NTPDases, we generated five site-directed mutants, Lpg1905E159A, R122A, N168A, Q193A, and W384A. Although the mutations E159A, R122A, Q193A, and W384A abrogated activity completely, N168A resulted in decreased activity caused by reduced affinity for nucleotides. When introduced into the lpg1905 mutant strain of L. pneumophila, only N168A partially restored the ability of L. pneumophila to replicate in THP-1 macrophages. Following intratracheal inoculation of A/J mice, none of the Lpg1905 mutants was able to restore virulence to an lpg1905 mutant during lung infection, thereby demonstrating the importance of NTPDase activity to L. pneumophila infection. Overall, the kinetic studies undertaken here demonstrated important differences to mammalian NTPDases and different sensitivities to NTPDase inhibitors that may reflect underlying structural variations.

  4. Discovery of novel inhibitors for Leishmania nucleoside diphosphatase kinase (NDK) based on its structural and functional characterization

    Science.gov (United States)

    Mishra, Arjun K.; Singh, Nidhi; Agnihotri, Pragati; Mishra, Shikha; Singh, Saurabh P.; Kolli, Bala K.; Chang, Kwang Poo; Sahasrabuddhe, Amogh A.; Siddiqi, M. I.; Pratap, J. Venkatesh

    2017-06-01

    Nucleoside diphosphate kinases (NDKs) are ubiquitous enzymes that catalyze the transfer of the γ-phosphate moiety from an NTP donor to an NDP acceptor, crucial for maintaining the cellular level of nucleoside triphosphates (NTPs). The inability of trypanosomatids to synthesize purines de novo and their dependence on the salvage pathway makes NDK an attractive target to develop drugs for the diseases they cause. Here we report the discovery of novel inhibitors for Leishmania NDK based on the structural and functional characterization of purified recombinant NDK from Leishmania amazonensis. Recombinant LaNDK possesses auto-phosphorylation, phosphotransferase and kinase activities with Histidine 117 playing an essential role. LaNDK crystals were grown by hanging drop vapour diffusion method in a solution containing 18% PEG-MME 500, 100 mM Bis-Tris propane pH 6.0 and 50 mM MgCl2. It belongs to the hexagonal space group P6322 with unit cell parameters a = b = 115.18, c = 62.18 Å and α = β = 90°, γ = 120°. The structure solved by molecular replacement methods was refined to crystallographic R-factor and Rfree values of 22.54 and 26.52%, respectively. Molecular docking and dynamics simulation -based virtual screening identified putative binding compounds. Protein inhibition studies of selected hits identified five inhibitors effective at micromolar concentrations. One of the compounds showed 45% inhibition of Leishmania promastigotes proliferation. Analysis of inhibitor-NDK complexes reveals the mode of their binding, facilitating design of new compounds for optimization of activities as drugs against leishmaniasis.

  5. Metabolomic profile and nucleoside composition of Cordyceps nidus sp. nov. (Cordycipitaceae: A new source of active compounds.

    Directory of Open Access Journals (Sweden)

    Juan Chiriví

    Full Text Available Cordyceps sensu lato is a genus of arthropod-pathogenic fungi, which have been used traditionally as medicinal in Asia. Within the genus, Ophiocordyceps sinensis is the most coveted and expensive species in China. Nevertheless, harvesting wild specimens has become a challenge given that natural populations of the fungus are decreasing and because large-scale culture of it has not yet been achieved. The worldwide demand for products derived from cultivable fungal species with medicinal properties has increased recently. In this study, we propose a new species, Cordyceps nidus, which parasitizes underground nests of trapdoor spiders. This species is phylogenetically related to Cordyceps militaris, Cordyceps pruinosa, and a sibling species of Cordyceps caloceroides. It is found in tropical rainforests from Bolivia, Brazil, Colombia and Ecuador. We also investigated the medicinal potential of this fungus based on its biochemical properties when grown on four different culture media. The metabolic profile particularly that of nucleosides, in polar and non-polar extracts was determined by UPLC, and then correlated to their antimicrobial activity and total phenolic content. The metabolome showed a high and significant dependency on the substrate used for fungal growth. The mass intensities of nucleosides and derivative compounds were higher in natural culture media in comparison to artificial culture media. Among these compounds, cordycepin was the predominant, showing the potential use of this species as an alternative to O. sinensis. Furthermore, methanol fractions showed antimicrobial activity against gram-positive bacteria, and less than 3.00 mg of gallic acid equivalents per g of dried extract were obtained when assessing its total phenolic content by modified Folin-Ciocalteu method. The presence of polyphenols opens the possibility of further exploring the antioxidant capacity and the conditions that may enhance this characteristic. The metabolic

  6. Quantifying the length and variance of the eukaryotic cell cycle phases by a stochastic model and dual nucleoside pulse labelling.

    Science.gov (United States)

    Weber, Tom Serge; Jaehnert, Irene; Schichor, Christian; Or-Guil, Michal; Carneiro, Jorge

    2014-07-01

    A fundamental property of cell populations is their growth rate as well as the time needed for cell division and its variance. The eukaryotic cell cycle progresses in an ordered sequence through the phases G1, S, G2, and M, and is regulated by environmental cues and by intracellular checkpoints. Reflecting this regulatory complexity, the length of each phase varies considerably in different kinds of cells but also among genetically and morphologically indistinguishable cells. This article addresses the question of how to describe and quantify the mean and variance of the cell cycle phase lengths. A phase-resolved cell cycle model is introduced assuming that phase completion times are distributed as delayed exponential functions, capturing the observations that each realization of a cycle phase is variable in length and requires a minimal time. In this model, the total cell cycle length is distributed as a delayed hypoexponential function that closely reproduces empirical distributions. Analytic solutions are derived for the proportions of cells in each cycle phase in a population growing under balanced growth and under specific non-stationary conditions. These solutions are then adapted to describe conventional cell cycle kinetic assays based on pulse labelling with nucleoside analogs. The model fits well to data obtained with two distinct proliferating cell lines labelled with a single bromodeoxiuridine pulse. However, whereas mean lengths are precisely estimated for all phases, the respective variances remain uncertain. To overcome this limitation, a redesigned experimental protocol is derived and validated in silico. The novelty is the timing of two consecutive pulses with distinct nucleosides that enables accurate and precise estimation of both the mean and the variance of the length of all phases. The proposed methodology to quantify the phase length distributions gives results potentially equivalent to those obtained with modern phase-specific biosensor

  7. Variability in non-nucleoside reverse transcriptase and protease inhibitor concentrations among HIV-infected adults in routine clinical practice

    Science.gov (United States)

    Moltó, José; Blanco, Asunción; Miranda, Cristina; Miranda, José; Puig, Jordi; Valle, Marta; DelaVarga, Meritxell; Fumaz, Carmina R; Barbanoj, Manuel José; Clotet, Bonaventura

    2006-01-01

    What is already known about this subject The concentration of protease and non-nucleoside reverse transcriptase inhibtors in plasma has been related to both efficacy and toxicity. Most antiretroviral concentration data come from selected populations of patients undergoing therapeutic drug monitoring programmes, which may overestimate interindividual variability. What this study adds Our study has demonstrated the large interindividual variability in antiretroviral drug concentrations in an unselected population of patients during routine clinical practice. These results may provide interesting information to clinicians for the management of antiretroviral therapy in HIV-infected patients. Aims The objective of this study was to assess interindividual variability in trough concentrations of plasma of non-nucleoside reverse transcriptase inhibitors (NNRTI) and protease inhibitors (PI) among HIV-infected adults in a routine outpatient setting. Methods One hundred and seventeen patients who attended our clinic for routine blood tests, and who were receiving antiretroviral therapy which included NNRTI or PI were studied. Patients were not informed that drug concentrations were going to be measured until blood sampling. The times of the last antiretroviral dose and of blood sampling were recorded. Drug concentrations were considered optimal if they were above the proposed minimum effective value. In addition, efavirenz, nevirapine and atazanavir concentrations were considered potentially toxic if they were >4.0 mg l−1, >6.0 mg l−1 and >0.85 mg l−1, respectively. Results Overall, interindividual variability of NNRTI and PI concentrations in plasma was approximately 50%, and only 68.4% of the patients had drug concentrations within the proposed therapeutic range. Poor adherence explained only 35% of subtherapeutic drug concentrations. Conclusion Interindividual variability in trough concentrations of NNRTI and PI among HIV-infected adults is large in routine clinical

  8. Metabolomic profile and nucleoside composition of Cordyceps nidus sp. nov. (Cordycipitaceae): A new source of active compounds.

    Science.gov (United States)

    Chiriví, Juan; Danies, Giovanna; Sierra, Rocio; Schauer, Nicolas; Trenkamp, Sandra; Restrepo, Silvia; Sanjuan, Tatiana

    2017-01-01

    Cordyceps sensu lato is a genus of arthropod-pathogenic fungi, which have been used traditionally as medicinal in Asia. Within the genus, Ophiocordyceps sinensis is the most coveted and expensive species in China. Nevertheless, harvesting wild specimens has become a challenge given that natural populations of the fungus are decreasing and because large-scale culture of it has not yet been achieved. The worldwide demand for products derived from cultivable fungal species with medicinal properties has increased recently. In this study, we propose a new species, Cordyceps nidus, which parasitizes underground nests of trapdoor spiders. This species is phylogenetically related to Cordyceps militaris, Cordyceps pruinosa, and a sibling species of Cordyceps caloceroides. It is found in tropical rainforests from Bolivia, Brazil, Colombia and Ecuador. We also investigated the medicinal potential of this fungus based on its biochemical properties when grown on four different culture media. The metabolic profile particularly that of nucleosides, in polar and non-polar extracts was determined by UPLC, and then correlated to their antimicrobial activity and total phenolic content. The metabolome showed a high and significant dependency on the substrate used for fungal growth. The mass intensities of nucleosides and derivative compounds were higher in natural culture media in comparison to artificial culture media. Among these compounds, cordycepin was the predominant, showing the potential use of this species as an alternative to O. sinensis. Furthermore, methanol fractions showed antimicrobial activity against gram-positive bacteria, and less than 3.00 mg of gallic acid equivalents per g of dried extract were obtained when assessing its total phenolic content by modified Folin-Ciocalteu method. The presence of polyphenols opens the possibility of further exploring the antioxidant capacity and the conditions that may enhance this characteristic. The metabolic composition and

  9. Trial-Based Functional Analysis and Functional Communication Training in an Early Childhood Setting

    Science.gov (United States)

    Lambert, Joseph M.; Bloom, Sarah E.; Irvin, Jennifer

    2012-01-01

    Problem behavior is common in early childhood special education classrooms. Functional communication training (FCT; Carr & Durand, 1985) may reduce problem behavior but requires identification of its function. The trial-based functional analysis (FA) is a method that can be used to identify problem behavior function in schools. We conducted…

  10. Meta-analysis of the clinical efficacy of combination therapy with traditional Chinese medicine and nucleoside analogue antiviral drugs for chronic hepatitis B

    Directory of Open Access Journals (Sweden)

    REN Shuang

    2013-04-01

    Full Text Available ObjectiveTo investigate whether the collected randomized controlled trials (RCTs published to date have shown that supplemental application of traditional Chinese medicine (TCM improves the outcome of chronic hepatitis B (CHB patients treated with nucleoside analogue antiviral drugs. MethodsLiterature collections on the Chinese Biomedical Literature Database, the Chinese Academic Journal Full-text Database, the Chinese Scientific Journals Database, PubMed, and Embase were searched from inception to July 2012 to identify publications of RCTs comparing the therapeutic efficacy of TCM combined with nucleoside analogues lamivudine (LDM, adefovir (ADV, entecavir (ETV to treat CHB. Inclusion criteria included: patients with CHB for more than six months, serum HbsAg-positivity, 2× upper normal limit (ULN≤alanine aminotransferase (ALT≤10× ULN, hepatitis B virus (HBV DNA-positivity, and no previous antiviral therapy for six months prior to study; a control group receiving a single nucleoside analogue; study treatment time of at least three months. The methodological quality of included RCTs was assessed using the Jadad scale. The meta-analysis was carried out with RevMan 5.0 software. Heterogeneity was examined by Chi-squared test. Pooled data was analyzed by the fixed effects model or random effects model, according to presence of heterogeneity. Publication bias was assessed by funnel plot asymmetry. ResultsA total of 14 RCTs were included in the analysis and comprised 1386 CHB patients with 703 in the trial groups and 683 in the control groups. Only one study had moderate-high methodological quality (Jadad score: 3, and the remaining studies had low methodological quality (Jadad score: 2, n=1; 1, n=12. The outcome measures included: ALT normalization rate, aspartate aminotransferase (AST normalization rate, hepatitis B e antigen (HbeAg-negativity rate, HBeAg seroconversion rate, HBV DNA-negativity rate, and incidence of the LAM-resistance YMDD

  11. Transient state kinetic studies of the MutT-catalyzed nucleoside triphosphate pyrophosphohydrolase reaction.

    Science.gov (United States)

    Xia, Zuyong; Azurmendi, Hugo F; Mildvan, Albert S

    2005-11-22

    The MutT pyrophosphohydrolase, in the presence of Mg2+, catalyzes the hydrolysis of nucleoside triphosphates by nucleophilic substitution at Pbeta, to yield the nucleotide and PP(i). The best substrate for MutT is the mutagenic 8-oxo-dGTP, on the basis of its Km being 540-fold lower than that of dGTP. Product inhibition studies have led to a proposed uni-bi-iso kinetic mechanism, in which PP(i) dissociates first from the enzyme-product complex (k3), followed by NMP (k4), leaving a product-binding form of the enzyme (F) which converts to the substrate-binding form (E) in a partially rate-limiting step (k5) [Saraswat, V., et al. (2002) Biochemistry 41, 15566-15577]. Single- and multiple-turnover kinetic studies of the hydrolysis of dGTP and 8-oxo-dGTP and global fitting of the data to this mechanism have yielded all of the nine rate constants. Consistent with an "iso" mechanism, single-turnover studies with dGTP and 8-oxo-dGTP hydrolysis showed slow apparent second-order rate constants for substrate binding similar to their kcat/Km values, but well below the diffusion limit (approximately 10(9) M(-1) s(-1)): k(on)app = 7.2 x 10(4) M(-1) s(-1) for dGTP and k(on)app = 2.8 x 10(7) M(-1) s(-1) for 8-oxo-dGTP. These low k(on)app values are fitted by assuming a slow iso step (k5 = 12.1 s(-1)) followed by fast rate constants for substrate binding: k1 = 1.9 x 10(6) M(-1) s(-1) for dGTP and k1 = 0.75 x 10(9) M(-1) s(-1) for 8-oxo-dGTP (the latter near the diffusion limit). With dGTP as the substrate, replacing Mg2+ with Mn2+ does not change k1, consistent with the formation of a second-sphere MutT-M2+-(H2O)-dGTP complex, but slows the iso step (k5) 5.8-fold, and its reverse (k(-5)) 25-fold, suggesting that the iso step involves a change in metal coordination, likely the dissociation of Glu-53 from the enzyme-bound metal so that it can function as the general base. Multiple-turnover studies with dGTP and 8-oxo-dGTP show bursts of product formation, indicating partially rate

  12. Structure-activity relationships of β-hydroxyphosphonate nucleoside analogues as cytosolic 5'-nucleotidase II potential inhibitors: synthesis, in vitro evaluation and molecular modeling studies.

    Science.gov (United States)

    Meurillon, Maïa; Marton, Zsuzsanna; Hospital, Audrey; Jordheim, Lars Petter; Béjaud, Jérôme; Lionne, Corinne; Dumontet, Charles; Périgaud, Christian; Chaloin, Laurent; Peyrottes, Suzanne

    2014-04-22

    The cytosolic 5'-nucleotidase II (cN-II) has been proposed as an attractive molecular target for the development of novel drugs circumventing resistance to cytotoxic nucleoside analogues currently used for treating leukemia and other malignant hemopathies. In the present work, synthesis of β-hydroxyphosphonate nucleoside analogues incorporating modifications either on the sugar residue or the nucleobase, and their in vitro evaluation towards the purified enzyme were carried out in order to determine their potency towards the inhibition of cN-II. In addition to the biochemical investigations, molecular modeling studies revealed important structural features for binding affinities towards the target enzyme. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  13. Synthesis and quantitative structure-activity relationship (QSAR) analysis of some novel oxadiazolo[3,4-d]pyrimidine nucleosides derivatives as antiviral agents.

    Science.gov (United States)

    Xu, Xiaojuan; Wang, Jun; Yao, Qizheng

    2015-01-15

    We have synthesized a series of 4H,6H-[1,2,5]oxadiazolo[3,4-d]pyrimidine-5,7-dione 1-oxide nucleoside and their anti-vesicular stomatitis virus (VSV) activities in Wish cell were also investigated in vitro. It was found that most compounds showed obvious anti-VSV activities and compound 9 with ribofuranoside improved the anti-VSV activity by approximately 10 times and 18 times compared to didanosine (DDI) and acyclovir, respectively. A quantitative structure-activity relationship (QSAR) study of these compounds as well as previous reported oxadiazolo[3,4-d]pyrimidine nucleoside derivatives indicated that compounds with high activity should have small values of logP(o/w), vsurf_G and a large logS value. These findings and results provide a base for further investigations. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. Determination of nucleosides and nucleotides in baby foods by hydrophilic interaction chromatography coupled to tandem mass spectrometry in the presence of hydrophilic ion-pairing reagents.

    Science.gov (United States)

    Mateos-Vivas, María; Rodríguez-Gonzalo, Encarnación; Domínguez-Álvarez, Javier; García-Gómez, Diego; Carabias-Martínez, Rita

    2016-11-15

    In this work we propose a rapid and efficient method for the joint determination of nucleosides and nucleotides in dairy and non-dairy baby foods based on hydrophilic interaction chromatography coupled to tandem mass spectrometry in the presence of diethylammonium (DEA) as a hydrophilic ion-pairing reagent (IP-HILIC-MS/MS). Sample treatment of the baby food included dilution with water and centrifugal ultrafiltration (CUF) with an additional washing step that notably improved the global performance of the process. Later dilution of the extract with acetonitrile allowed adequate separation in the HILIC system. With the proposed treatment, we obtained extraction recoveries higher than 80% and, additionally, no matrix effects were observed. The CUF-IP-HILIC-MS/MS method was validated according to the 2002/657/EC decision and was used for the quantification of nucleotides and nucleosides in sixteen samples of commercial baby foods. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Programmed cell death triggered by nucleotide pool damage and its prevention by MutT homolog-1 (MTH1) with oxidized purine nucleoside triphosphatase.

    Science.gov (United States)

    Nakabeppu, Yusaku; Oka, Sugako; Sheng, Zijing; Tsuchimoto, Daisuke; Sakumi, Kunihiko

    2010-11-28

    Accumulation of oxidized bases such as 8-oxoguanine in either nuclear or mitochondrial DNA triggers various cellular dysfunctions including mutagenesis, and programmed cell death or senescence. Recent studies have revealed that oxidized nucleoside triphosphates such as 8-oxo-dGTP in the nucleotide pool are the main source of oxidized bases accumulating in the DNA of cells under oxidative stress. To counteract such deleterious effects of nucleotide pool damage, mammalian cells possess MutT homolog-1 (MTH1) with oxidized purine nucleoside triphosphatase and related enzymes, thus minimizing the accumulation of oxidized bases in cellular DNA. Depletion or increased expression of the MTH1 protein have revealed its significant roles in avoiding programmed cell death or senescence as well as mutagenesis, and accumulating evidences indicate that MTH1 is involved in suppression of degenerative disorders such as neurodegeneration. 2010 Elsevier B.V. All rights reserved.

  16. Design of an adenosine phosphorylase by active-site modification of murine purine nucleoside phosphorylase. Enzyme kinetics and molecular dynamics simulation of Asn-243 and Lys-244 substitutions of purine nucleoside phosphorylase.

    Science.gov (United States)

    Maynes, J T; Yam, W; Jenuth, J P; Gang Yuan, R; Litster, S A; Phipps, B M; Snyder, F F

    1999-12-01

    Our objective was to alter the substrate specificity of purine nucleoside phosphorylase such that it would catalyse the phosphorolysis of 6-aminopurine nucleosides. We modified both Asn-243 and Lys-244 in order to promote the acceptance of the C6-amino group of adenosine. The Asn-243-Asp substitution resulted in an 8-fold increase in K(m) for inosine from 58 to 484 microM and a 1000-fold decrease in k(cat)/K(m). The Asn-243-Asp construct catalysed the phosphorolysis of adenosine with a K(m) of 45 microM and a k(cat)/K(m) 8-fold that with inosine. The Lys-244-Gln construct showed only marginal reduction in k(cat)/K(m), 83% of wild type, but had no activity with adenosine. The Asn-243-Asp;Lys-244-Gln construct had a 14-fold increase in K(m) with inosine and 7-fold decrease in k(cat)/K(m) as compared to wild type. This double substitution catalysed the phosphorolysis of adenosine with a K(m) of 42 microM and a k(cat)/K(m) twice that of the single Asn-243-Asp substitution. Molecular dynamics simulation of the engineered proteins with adenine as substrate revealed favourable hydrogen bond distances between N7 of the purine ring and the Asp-243 carboxylate at 2.93 and 2.88 A, for Asn-243-Asp and the Asn-243-Asp;Lys-244-Gln constructs respectively. Simulation also supported a favourable hydrogen bond distance between the purine C6-amino group and Asp-243 at 2.83 and 2.88 A for each construct respectively. The Asn-243-Thr substitution did not yield activity with adenosine and simulation gave unfavourable hydrogen bond distances between Thr-243 and both the C6-amino group and N7 of the purine ring. The substitutions were not in the region of phosphate binding and the apparent S(0.5) for phosphate with wild type and the Asn-243-Asp enzymes were 1.35+/-0.01 and 1.84+/-0.06 mM, respectively. Both proteins exhibited positive co-operativity with phosphate giving Hill coefficients of 7.9 and 3.8 respectively.

  17. Human N6-Methyl-AMP/dAMP aminohydrolase (abacavir 5’-monophosphate deaminase) is capable of metabolizing N6-substituted purine acyclic nucleoside phosphonates

    Czech Academy of Sciences Publication Activity Database

    Schinkmanová, Markéta; Votruba, Ivan; Shibata, R.; Han, B.; Liu, X.; Cihlař, T.; Holý, Antonín

    2008-01-01

    Roč. 73, č. 2 (2008), s. 275-291 ISSN 0010-0765 R&D Projects: GA MŠk 1M0508; GA AV ČR 1QS400550501 Institutional research plan: CEZ:AV0Z40550506 Keywords : guanine * acyclic nucleoside phosphonates * cPrPMEDAP * abacavir 5'-phosphate Subject RIV: CC - Organic Chemistry Impact factor: 0.784, year: 2008

  18. Kinetic analysis of ligand binding to the Ehrlich cell nucleoside transporter: Pharmacological characterization of allosteric interactions with the sup 3 Hnitrobenzylthioinosine binding site

    Energy Technology Data Exchange (ETDEWEB)

    Hammond, J.R. (Department of Pharmacology and Toxicology, University of Western Ontario, London (Canada))

    1991-06-01

    Kinetic analysis of the binding of {sup 3}Hnitrobenzylthioinosine ({sup 3}H NBMPR) to Ehrlich ascites tumor cell plasma membranes was conducted in the presence and absence of a variety of nucleoside transport inhibitors and substrates. The association of {sup 3}H NBMPR with Ehrlich cell membranes occurred in two distinct phases, possibly reflecting functional conformation changes in the {sup 3}HNBMPR binding site/nucleoside transporter complex. Inhibitors of the equilibrium binding of {sup 3}HNBMPR, tested at submaximal inhibitory concentrations, generally decreased the rate of association of {sup 3}HNBMPR, but the magnitude of this effect varied significantly with the agent tested. Adenosine and diazepam had relatively minor effects on the association rate, whereas dipyridamole and mioflazine slowed the rate dramatically. Inhibitors of nucleoside transport also decreased the rate of dissociation of {sup 3}HNBMPR, with an order of potency significantly different from their relative potencies as inhibitors of the equilibrium binding of {sup 3}HNBMPR. Dilazep, dipyridamole, and mioflazine were effective inhibitors of both {sup 3}HNBMPR dissociation and equilibrium binding. The lidoflazine analogue R75231, on the other hand, had no effect on the rate of dissociation of {sup 3}HNBMPR at concentrations below 300 microM, even though it was one of the most potent inhibitors of {sup 3}HNBMPR binding tested (Ki less than 100 nM). In contrast, a series of natural substrates for the nucleoside transport system enhanced the rate of dissociation of {sup 3}HNBMPR with an order of effectiveness that paralleled their relative affinities for the permeant site of the transporter. The most effective enhancers of {sup 3}HNBMPR dissociation, however, were the benzodiazepines diazepam, chlordiazepoxide, and triazolam.

  19. Maternal-Fetal Transfer and Amniotic Fluid Accumulation of Nucleoside Analogue Reverse Transcriptase Inhibitors in Human Immunodeficiency Virus-Infected Pregnant Women

    OpenAIRE

    Chappuy, Hélène; Tréluyer, Jean-Marc; Jullien, Vincent; Dimet, Jérôme; Rey, Elisabeth; Fouché, Maria; Firtion, Ghislaine; Pons, Gérard; Mandelbrot, Laurent

    2004-01-01

    This study was performed to investigate placental transfer of nucleoside analogue reverse transcriptase inhibitors (NRTIs) and their concentrations in amniotic fluid when given to human immunodeficiency virus (HIV)-infected pregnant women. A total of 100 HIV type 1-infected mothers receiving antiretroviral therapy, including one or more NRTIs, for clinical indications at the time of delivery were enrolled. Maternal blood samples and amniotic fluid were obtained during delivery or cesarean sec...

  20. FT-IR spectra of the anti-HIV nucleoside analogue d4T (Stavudine). Solid state simulation by DFT methods and scaling by different procedures

    Science.gov (United States)

    Alcolea Palafox, M.; Kattan, D.; Afseth, N. K.

    2018-04-01

    A theoretical and experimental vibrational study of the anti-HIV d4T (stavudine or Zerit) nucleoside analogue was carried out. The predicted spectra in the three most stable conformers in the biological active anti-form of the isolated state were compared. Comparison of the conformers with those of the natural nucleoside thymidine was carried out. The calculated spectra were scaled by using different scaling procedures and three DFT methods. The TLSE procedure leads to the lowest error and is thus recommended for scaling. With the population of these conformers the IR gas-phase spectra were predicted. The crystal unit cell of the different polymorphism forms of d4T were simulated through dimer forms by using DFT methods. The scaled spectra of these dimer forms were compared. The FT-IR spectrum was recorded in the solid state in the 400-4000 cm-1 range. The respective vibrational bands were analyzed and assigned to different normal modes of vibration by comparison with the scaled vibrational values of the different dimer forms. Through this comparison, the polymorphous form of the solid state sample was identified. The study indicates that d4T exist only in the ketonic form in the solid state. The results obtained were in agreement with those determined in related anti-HIV nucleoside analogues.

  1. Novel tentacle-type polymer stationary phase grafted with anion exchange polymer chains for open tubular CEC of nucleosides and proteins.

    Science.gov (United States)

    Aydoğan, Cemil; Çetin, Kemal; Denizli, Adil

    2014-08-07

    A novel and simple method for preparation of a tentacle-type polymer stationary phase grafted with polyethyleneimine (PEI) anion exchanger was developed for open tubular capillary electrochromatography (OT-CEC) of nucleosides and proteins. The polymeric stationary phase was prepared using 3-chloro-2-hydroxypropyl methacrylate (HPMA-Cl)-based reactive monomer. The preparation procedure included pretreatment of the capillary inner wall, silanization, in situ graft polymerization with HPMA-Cl and PEI modification. To compare with the tentacle-type capillary column with PEI functionalization, a monolayer capillary column without PEI functionalization was also prepared. The electrochromatographic characterization of the prepared open tubular column was performed using alkylbenzenes. The electroosmotic flow (EOF) with regard to PEI concentrations and the running buffer pH was investigated. The separation conditions of the nucleosides and the proteins were optimized. The modified tentacle-type column with high anion exchange capacity has proven to afford better retention and resolution for the separation of nucleosides and proteins. The PEI functionalization column can also provide long-term stable use for biomolecule separation using a single capillary with relative standard deviation values of retention times of less than 2%. The results indicate that the present method for open tubular capillary preparation with a HPMA-Cl-based reactive monomer is promising for OT-CEC biomolecule separation.

  2. Optimal allocation of selected T-series advanced base functional component equipment packages

    OpenAIRE

    Weitz, Jeffrey D.

    2000-01-01

    Advanced Base Functional Components (ABFCs) are pre-planned modular units of equipment and personnel designed to extend or create the logistic infrastructure required to support naval expeditionary operations. The ABFC program is structured to combine trained personnel with the equipment needed to perform a particular logistic mission such as seaport operations and cargo handling, warehousing, or freight terminal operations. If the ABFC must deploy to a theater of operations, individual equip...

  3. Isotope-specific and amino acid-specific heavy atom substitutions alter barrier crossing in human purine nucleoside phosphorylase.

    Science.gov (United States)

    Suarez, Javier; Schramm, Vern L

    2015-09-08

    Computational chemistry predicts that atomic motions on the femtosecond timescale are coupled to transition-state formation (barrier-crossing) in human purine nucleoside phosphorylase (PNP). The prediction is experimentally supported by slowed catalytic site chemistry in isotopically labeled PNP (13C, 15N, and 2H). However, other explanations are possible, including altered volume or bond polarization from carbon-deuterium bonds or propagation of the femtosecond bond motions into slower (nanoseconds to milliseconds) motions of the larger protein architecture to alter catalytic site chemistry. We address these possibilities by analysis of chemistry rates in isotope-specific labeled PNPs. Catalytic site chemistry was slowed for both [2H]PNP and [13C, 15N]PNP in proportion to their altered protein masses. Secondary effects emanating from carbon-deuterium bond properties can therefore be eliminated. Heavy-enzyme mass effects were probed for local or global contributions to catalytic site chemistry by generating [15N, 2H]His8-PNP. Of the eight His per subunit, three participate in contacts to the bound reactants and five are remote from the catalytic sites. [15N, 2H]His8-PNP had reduced catalytic site chemistry larger than proportional to the enzymatic mass difference. Altered barrier crossing when only His are heavy supports local catalytic site femtosecond perturbations coupled to transition-state formation. Isotope-specific and amino acid specific labels extend the use of heavy enzyme methods to distinguish global from local isotope effects.

  4. Switching from tenofovir and nucleoside analogue therapy to tenofovir monotherapy in virologically suppressed chronic hepatitis B patients with antiviral resistance.

    Science.gov (United States)

    Kim, Dong Yun; Lee, Hye Won; Song, Jeong Eun; Kim, Beom Kyung; Kim, Seung Up; Kim, Do Young; Ahn, Sang Hoon; Han, Kwang-Hyub; Park, Jun Yong

    2018-03-01

    It is unclear whether chronic hepatitis B (CHB) patients with antiviral resistance, who achieve a complete virologic response (CVR) with tenofovir disoproxil fumarate (TDF) and nucleoside analogue (NUC) combination therapy, maintain CVR if switched to TDF monotherapy. We investigated the persistence of CVR after cessation of NUC in virologically suppressed antiviral resistant CHB patients using TDF+NUC combination therapy. This study recruited 76 antiviral-resistant CHB patients showing CVR on TDF+entecavir (ETV) (n = 52), TDF+lamivudine (LAM; n = 14), and TDF+telbivudine (LdT; n = 10) combination therapy, who were switched to TDF monotherapy as step-down therapy. At baseline, 47 patients were male and the median age was 53.0 years (range: 30-78 years); 72.3% cases were hepatitis B e antigen-positive (HBeAg+) and 23.7% were of liver cirrhosis. The median duration of TDF+NUC combination therapy was 20.8 months (range: 3-46 months). At a median follow-up of 24.7 months (range: 12-48 months) after switching to TDF monotherapy, all 76 patients maintained CVR, regardless of the duration of combination therapy and the type of prior NUC and antiviral resistance. Renal dysfunction was not observed during the treatment period. The step-down strategy of switching from TDF+NUC combination therapy to TDF monotherapy in virologically suppressed CHB patients with antiviral resistance should be considered. © 2017 Wiley Periodicals, Inc.

  5. Functionalized tricyclic cytosine analogues provide nucleoside fluorophores with improved photophysical properties and a range of solvent sensitivities.

    Science.gov (United States)

    Rodgers, Brittney J; Elsharif, Nada A; Vashisht, Nisha; Mingus, Macy M; Mulvahill, Mark A; Stengel, Gudrun; Kuchta, Robert D; Purse, Byron W

    2014-02-10

    Tricyclic cytosines (tC and tC(O) frameworks) have emerged as a unique class of fluorescent nucleobase analogues that minimally perturb the structure of B-form DNA and that are not quenched in duplex nucleic acids. Systematic derivatization of these frameworks is a likely approach to improve on and diversify photophysical properties, but has not so far been examined. Synthetic methods were refined to improve on tolerance for electron-donating and electron-withdrawing groups, resulting in a series of eight new, fluorescent cytidine analogues. Photophysical studies show that substitution of the framework results in a pattern of effects largely consistent across tC and tC(O) and provides nucleoside fluorophores that are brighter than either parent. Moreover, a range of solvent sensitivities is observed, offering promise that this family of probes can be extended to new applications that require reporting on the local environment. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Target-triggered transcription machinery for ultra-selective and sensitive fluorescence detection of nucleoside triphosphates in one minute.

    Science.gov (United States)

    Dong, Jiantong; Wu, Tongbo; Xiao, Yu; Chen, Lu; Xu, Lei; Li, Mengyuan; Zhao, Meiping

    2018-02-15

    Nucleoside triphosphates (NTPs) play important roles in living organisms. However, no fluorescent assays are currently available to simply and rapidly detect multiple NTPs with satisfactory selectivity, sensitivity and low cost. Here we demonstrate for the first time a target-triggered in-vitro transcription machinery for ultra-selective, sensitive and instant fluorescence detection of multiple NTPs. The machinery assembles RNA polymerase, DNA template and non-target NTPs to convert the target NTP into equivalent RNA signal sequences which are monitored by the fluorescence enhancement of molecular beacon. The machinery offers excellent selectivity for the target NTP against NDP, NMP and dNTP. Notably, to accelerate the kinetics of the machinery while maintain its high specificity, we investigated the sequence of DNA templates systematically and established a set of guidelines for the design of the optimum DNA templates, which allowed for instant detection of the target NTP at fmol level in less than 1min. Furthermore, the machinery could be transformed into logic gates to study the coeffects of two NTPs in biosynthesis and real-time monitoring systems to reflect the distribution of NTP in nucleotide pools. These results provide very useful and low-cost tools for both biochemical tests and point-of-care analysis. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Hydrogen/Deuterium Exchange Mass Spectrometry Reveals Mechanistic Details of Activation of Nucleoside Diphosphate Kinases by Oligomerization.

    Science.gov (United States)

    Dautant, Alain; Meyer, Philippe; Georgescauld, Florian

    2017-06-13

    Most oligomeric proteins become active only after assembly, but why oligomerization is required to support function is not well understood. Here, we address this question using the wild type (WT) and a destabilized mutant (D93N) of the hexameric nucleoside diphosphate kinase from the pathogen Mycobacterium tuberculosis (Mt-NDPK). The conformational dynamics and oligomeric states of each were analyzed during unfolding and/or folding by hydrogen/deuterium exchange mass spectrometry (HDX-MS) at peptide resolution and by additional biochemical techniques. We found that WT and D93N native hexamers present a stable core and a flexible periphery, the latter being more flexible for the destabilized mutant. Stable but inactive species formed during unfolding of D93N and folding of WT were characterized. For the first time, we show that both of these species are nativelike dimers, each of its monomers having a major subdomain folded, while a minor subdomain (Kpn/α 0 ) remains unfolded. The Kpn/α 0 subdomain, which belongs to the catalytic site, becomes structured only upon hexamerization, explaining why oligomerization is required for NDPK activity. Further HDX-MS studies are necessary to establish the general activation mechanism for other homo-oligomers.

  8. 4-amino-5-aryl-6-arylethynylpyrimidines: structure-activity relationships of non-nucleoside adenosine kinase inhibitors.

    Science.gov (United States)

    Matulenko, Mark A; Paight, Ernest S; Frey, Robin R; Gomtsyan, Arthur; DiDomenico, Stanley; Jiang, Meiqun; Lee, Chih-Hung; Stewart, Andrew O; Yu, Haixia; Kohlhaas, Kathy L; Alexander, Karen M; McGaraughty, Steve; Mikusa, Joseph; Marsh, Kennan C; Muchmore, Steven W; Jakob, Clarissa L; Kowaluk, Elizabeth A; Jarvis, Michael F; Bhagwat, Shripad S

    2007-02-15

    A series of non-nucleoside adenosine kinase (AK) inhibitors is reported. These inhibitors originated from the modification of 5-(3-bromophenyl)-7-(6-morpholin-4-ylpyridin-3-yl)pyrido[2,3-d]pyrimidin-4-ylamine (ABT-702). The identification of a linker that would approximate the spatial arrangement found between the pyrimidine ring and the aryl group at C(7) in ABT-702 was a key element in this modification. A search of potential linkers led to the discovery of an acetylene moiety as a suitable scaffold. It was hypothesized that the aryl acetylenes, ABT-702, and adenosine bound to the active site of AK (closed form) in a similar manner with respect to the orientation of the heterocyclic base. Although potent acetylene analogs were discovered based on this assumption, an X-ray crystal structure of 5-(4-dimethylaminophenyl)-6-(6-morpholin-4-ylpyridin-3-ylethynyl)pyrimidin-4-ylamine (16a) revealed a binding orientation contrary to adenosine. In addition, this compound bound tightly to a unique open conformation of AK. The structure-activity relationships and unique ligand orientation and protein conformation are discussed.

  9. Diphenyl ether non-nucleoside reverse transcriptase inhibitors with excellent potency against resistant mutant viruses and promising pharmacokinetic properties.

    Science.gov (United States)

    Sweeney, Zachary K; Kennedy-Smith, Joshua J; Wu, Jeffrey; Arora, Nidhi; Billedeau, J Roland; Davidson, James P; Fretland, Jennifer; Hang, Julie Q; Heilek, Gabrielle M; Harris, Seth F; Hirschfeld, Donald; Inbar, Petra; Javanbakht, Hassan; Jernelius, Jesper A; Jin, Qingwu; Li, Yu; Liang, Weiling; Roetz, Ralf; Sarma, Keshab; Smith, Mark; Stefanidis, Dimitrio; Su, Guoping; Suh, Judy M; Villaseñor, Armando G; Welch, Michael; Zhang, Fang-Jie; Klumpp, Klaus

    2009-01-01

    Non-nucleoside reverse transcriptase inhibitors (NNRTIs) are part of the preferred treatment regimens for individuals infected with HIV. These NNRTI-based regimens are efficacious, but the most popular NNRTIs have a low genetic barrier to resistance and have been associated with adverse events. There is therefore still a need for efficacious antiviral medicines that facilitate patient adherence and allow durable suppression of viral replication. As part of an extensive program targeted toward the discovery of NNRTIs that have favorable pharmacokinetic properties, good potency against NNRTI-resistant viruses, and a high genetic barrier to drug resistance, we focused on the optimization of a series of diaryl ether NNRTIs. In the course of this effort, we employed molecular modeling to design a new set of NNRTIs that that are active against wild-type HIV and key NNRTI-resistant mutant viruses. The structure-activity relationships observed in this series of compounds provide insight into the structural features required for NNRTIs that inhibit the replication of a wide range of mutant viruses. Selected compounds have promising pharmacokinetic profiles.

  10. Variability in non-nucleoside reverse transcriptase and protease inhibitors concentrations among HIV-infected adults in routine clinical practice

    Science.gov (United States)

    Moltó, José; Blanco, Asunción; Miranda, Cristina; Miranda, José; Puig, Jordi; Valle, Marta; DelaVarga, Meritxell; Fumaz, Carmina R; Barbanoj, Manuel José; Clotet, Bonaventura

    2007-01-01

    Aims The objective of this study was to assess interindividual variability in plasma trough concentrations of non-nucleoside reverse transcriptase inhibitors (NNRTI) and protease inhibitors (PI) among HIV-infected adults in an outpatient routine clinical practice setting. Methods The study included 117 patients who attended our clinic for routine outpatient blood tests and who were receiving antiretroviral therapy which included NNRTI or PI. Patients were not informed that drug concentrations were going to be assessed until blood sampling. The time of the last antiretroviral treatment intake and blood sampling were recorded. Drug concentrations were considered optimal if they were above the proposed minimum effective concentration. In addition, efavirenz, nevirapine and atazanavir concentrations were considered potentially toxic if they were higher than 4.0 mg l−1, 6.0 mg l−1, and 0.85 mg l−1, respectively. Results Overall, interindividual variability in NNRTI and PI plasma concentrations was approximately 50%, and only 68.4% of the patients had drug concentrations within the proposed therapeutic range. Inappropriate adherence only explained 35% of subtherapeutic drug concentrations. Conclusion Interindividual variability in trough concentrations of NNRTI and PI among HIV-infected adults is large in routine clinical practice, with drug concentrations being outside the therapeutic window in a significant proportion of patients. Therapeutic drug monitoring may be useful to guide antiretroviral therapy in clinical practice. PMID:17223856

  11. Purine Nucleoside Phosphorylase (PNP) Activity of Lymphocytes and T Cell Subsets in Peripheral Blood in Thyroid Tumors

    International Nuclear Information System (INIS)

    Kim, Dong Soo

    1992-01-01

    To elucidate alteration of purine nucleoside phosphorylase (PNP) activity of peripheral lymphocytes and helper/inducer and suppressor/cytototxic T cells in patients with thyroid tumors, the author examined PNP activity, and CD4 + and CD8 + cells of peripheral blood in 20 cases of simple goiter, 9 cases of thyroid adenoma and 20 cases of thyroid cancer as well as 11 cases of adult healthy subjects as control. Diagnoses were established on the basis of commonly accepted clinical and biochemical criteria in simple goiter and were confirmed histopathologically in thyroid adenoma and cancer. All blood was obtained from veins of the patients and control subjects in Pusan National University Hospital during the period of January to August, 1991. The results obtained were summarized as follows: 1) The PNP activity was significantly decreased or tended to be decreased in thyroid adenomas and cancers as compared with control subjects and simple goiters. 2) The percentage of CD8 cells was significantly decreased or tended to be decreased in thyroid cancers as compared with simple goiters, thyroid adenomas and control subjects. 3) The CD4/CD8 ratio was significantly increased or tended to be increased in thyroid cancer as compared with simple goiters, thyroid adenomas and control subjects. On the basis of the results, it can be suggested that the immunodysfunction in thyroid cancer may be due to decreased suppressor/cytotoxic T cells, and the estimation of PNP activity of peripheral lymphocyte is a helpful test in detecting the immune status in thyroid tumors.

  12. Global warming, plant paraquat resistance, and light signal transduction through nucleoside diphosphate kinase as a paradigm for increasing food supply.

    Science.gov (United States)

    Hasunuma, Kohji; Yoshida, Yusuke; Haque, Mohamed Emdadul; Wang, Ni-yan; Fukamatsu, Yosuke; Miyoshi, Osamu; Lee, Bumkyu

    2011-10-01

    Light signal transduction was studied in extracts of mycelia of the fungus Neurospora crassa, and the third internodes of dark-grown Pisum sativum cv Alaska. Both processes increased the phosphorylation of nucleoside diphosphate kinase (NDPK). NDPK may function as a carrier of reduction equivalents, as it binds NADH, thereby providing electrons to transform singlet oxygen to superoxide by catalases (CAT). As the C-termini of NDPK interact with CAT which receive singlet oxygen, emitted from photoreceptors post light perception (which is transmitted to ambient triplet oxygen), we hypothesize that this may increase phospho-NDPK. Singlet oxygen, emitted from the photoreceptor, also reacts with unsaturated fatty acids in membranes thereby forming malonedialdehyde, which in turn could release ions from, e.g., the thylacoid membrane thereby reducing the rate of photosynthesis. A mutant of Alaska pea, which exhibited two mutations in chloroplast NDPK-2 and one mutation in mitochondrial localized NDPK-3, was resistant to reactive oxygen species including singlet oxygen and showed an increase in the production of carotenoids, anthocyanine, and thereby could reduce the concentration of singlet oxygen. The reduction of the concentration of singlet oxygen is predicted to increase the yield of crop plants, such as Alaska pea, soybean, rice, wheat, barley, and sugarcane. This approach to increase the yield of crop plants may contribute not only to enhance food supply, but also to reduce the concentration of CO(2) in the atmosphere.

  13. BCX4430 – a broad-spectrum antiviral adenosine nucleoside analog under development for the treatment of Ebola virus disease

    Science.gov (United States)

    Taylor, Raymond; Kotian, Pravin; Warren, Travis; Panchal, Rekha; Bavari, Sina; Julander, Justin; Dobo, Sylvia; Rose, Angela; El-Kattan, Yahya; Taubenheim, Brian; Babu, Yarlagadda; Sheridan, William P.

    2016-01-01

    The adenosine nucleoside analog BCX4430 is a direct-acting antiviral drug under investigation for the treatment of serious and life-threatening infections from highly pathogenic viruses, such as the Ebola virus. Cellular kinases phosphorylate BCX4430 to a triphosphate that mimics ATP; viral RNA polymerases incorporate the drug’s monophosphate nucleotide into the growing RNA chain, causing premature chain termination. BCX4430 is active in vitro against many RNA viral pathogens, including the filoviruses and emerging infectious agents such as MERS-CoV and SARS-CoV. In vivo, BCX4430 is active after intramuscular, intraperitoneal, and oral administration in a variety of experimental infections. In nonclinical studies involving lethal infections with Ebola virus, Marburg virus, Rift Valley fever virus, and Yellow Fever virus, BCX4430 has demonstrated pronounced efficacy. In experiments conducted in several models, both a reduction in the viral load and an improvement in survival were found to be related to the dose of BCX4430. A Phase 1 clinical trial of intramuscular administration of BCX4430 in healthy subjects is currently ongoing. PMID:27095300

  14. Hepatitis C virus NS3 protein polynucleotide-stimulated nucleoside triphosphatase and comparison with the related pestivirus and flavivirus enzymes.

    Science.gov (United States)

    Suzich, J A; Tamura, J K; Palmer-Hill, F; Warrener, P; Grakoui, A; Rice, C M; Feinstone, S M; Collett, M S

    1993-01-01

    Sequence motifs within the nonstructural protein NS3 of members of the Flaviviridae family suggest that this protein possesses nucleoside triphosphatase (NTPase) and RNA helicase activity. The RNA-stimulated NTPase activity of this protein from prototypic members of the Pestivirus and Flavivirus genera has recently been established and enzymologically characterized. Here, we experimentally demonstrate that the NS3 protein from a member of the third genus of Flaviviridae, human hepatitis C virus (HCV), also possesses a polynucleotide-stimulated NTPase activity. Characterization of the purified HCV NTPase activity showed that it exhibited reaction condition optima with respect to pH, MgCl2, and salt identical to those of the representative pestivirus and flavivirus enzymes. However, each NTPase also possessed several unique properties when compared with one another. Notably, the profile of polynucleotide stimulation of the NTPase activity was distinct for the three enzymes. The HCV NTPase was the only one whose activity was significantly enhanced by a deoxyribopolynucleotide. Additional distinguishing features among the three enzymes relating to the kinetic properties of their NTPase activities are discussed. These studies provide a foundation for investigation of the putative RNA helicase activity of these proteins and for further study of the role of the NS3 proteins of members of the Flaviviridae in the replication cycle of these viruses. Images PMID:8396675

  15. Synthesis of chromone, quinolone, and benzoxazinone sulfonamide nucleosides as conformationally constrained inhibitors of adenylating enzymes required for siderophore biosynthesis.

    Science.gov (United States)

    Engelhart, Curtis A; Aldrich, Courtney C

    2013-08-02

    MbtA catalyzes the first committed step of mycobactin biosynthesis in Mycobacterium tuberculosis (Mtb) and is responsible for the incorporation of salicylic acid into the mycobactin siderophores. 5'-O-[N-(Salicyl)sulfamoyl]adenosine (Sal-AMS) is an extremely potent nucleoside inhibitor of MbtA that possesses excellent activity against whole-cell Mtb but suffers from poor bioavailability. In an effort to improve the bioavailability, we have designed four conformationally constrained analogues of Sal-AMS that remove two rotatable bonds and the ionized sulfamate group on the basis of computational and structural studies. Herein we describe the synthesis, biochemical, and microbiological evaluation of chromone-, quinolone-, and benzoxazinone-3-sulfonamide derivatives of Sal-AMS. We developed new chemistry to assemble these three heterocycles from common β-ketosulfonamide intermediates. The synthesis of the chromone- and quinolone-3-sulfonamide intermediates features formylation of a β-ketosulfonamide employing dimethylformamide dimethyl acetal to afford an enaminone that can react intramolecularly with a phenol or intermolecularly with a primary amine via addition-elimination reaction(s). The benzoxazinone-3-sulfonamide was prepared by nitrosation of a β-ketosulfonamide followed by intramolecular nucleophilic aromatic substitution. Mitsunobu coupling of these bicyclic sulfonamides with a protected adenosine derivative followed by global deprotection provides a concise synthesis of the respective inhibitors.

  16. Transport mechanisms of a novel antileukemic and antiviral compound 9-norbornyl-6-chloropurine

    Czech Academy of Sciences Publication Activity Database

    Plačková, Pavla; Hřebabecký, Hubert; Šála, Michal; Nencka, Radim; Elbert, Tomáš; Mertlíková-Kaiserová, Helena

    2015-01-01

    Roč. 30, č. 1 (2015), s. 57-62 ISSN 1475-6366 R&D Projects: GA ČR GAP303/11/1297 Institutional support: RVO:61388963 Keywords : carbocyclic nucleoside analogs * CCRF-CEM cells * facilitated diffusion * MRP proteins * P-glycoprotein Subject RIV: CE - Biochemistry Impact factor: 3.428, year: 2015

  17. A New Analogue of Locked Cyclohexane Nucleic Acids

    Czech Academy of Sciences Publication Activity Database

    Šála, Michal; Dejmek, Milan; Procházková, Eliška; Hřebabecký, Hubert; Rybáček, Jiří; Dračínský, Martin; Novák, Pavel; Rosenbergová, Šárka; Rosenberg, Ivan; Nencka, Radim

    2015-01-01

    Roč. 47, č. 17 (2015), s. 2654-2662 ISSN 0039-7881 R&D Projects: GA ČR GPP207/12/P625; GA ČR GA13-26526S Institutional support: RVO:61388963 Keywords : nucleosides * bicyclic compounds * oligonucleotides * carbocycles * medicinal chemistry Subject RIV: CC - Organic Chemistry Impact factor: 2.652, year: 2015

  18. 9-Norbornyl-6-chloropurine Is a Novel Antileukemic Compound Interacting with Cellular GSH

    Czech Academy of Sciences Publication Activity Database

    Plačková, Pavla; Rozumová, N.; Hřebabecký, Hubert; Šála, Michal; Nencka, Radim; Elbert, Tomáš; Dvořáková, Alexandra; Votruba, Ivan; Mertlíková-Kaiserová, Helena

    2013-01-01

    Roč. 33, č. 8 (2013), s. 3163-3168 ISSN 0250-7005 R&D Projects: GA ČR GAP303/11/1297 Institutional support: RVO:61388963 Keywords : Substituted 6-chloropurines * carbocyclic nucleoside analogs * glutathione-S-transferase * glutathione depletion Subject RIV: FD - Oncology ; Hematology Impact factor: 1.872, year: 2013

  19. The growth and electronic structure of azobenzene-based functional molecules on layered crystals

    International Nuclear Information System (INIS)

    Iwicki, J; Ludwig, E; Buck, J; Kalläne, M; Kipp, L; Rossnagel, K; Köhler, F; Herges, R

    2012-01-01

    In situ ultraviolet photoelectron spectroscopy is used to study the growth of ultrathin films of azobenzene-based functional molecules (azobenzene, Disperse Orange 3 and a triazatriangulenium platform with an attached functional azo-group) on the layered metal TiTe 2 and on the layered semiconductor HfS 2 at liquid nitrogen temperatures. Effects of intermolecular interactions, of the substrate electronic structure, and of the thermal energy of the sublimated molecules on the growth process and on the adsorbate electronic structure are identified and discussed. A weak adsorbate-substrate interaction is particularly observed for the layered semiconducting substrate, holding the promise of efficient molecular photoswitching.

  20. The C8-2′-Deoxyguanosine Adduct of 2-Amino-3-methylimidazo[1,2-d]naphthalene, a Carbocyclic Analogue of the Potent Mutagen 2-Amino-3-methylimidazo[4,5-f]quinoline (IQ), is a Block to Replication in Vitro

    Science.gov (United States)

    Christov, Plamen P.; Chowdhury, Goutam; Garmendia, Craig A.; Wang, Feng; Stover, James S.; Elmquist, C. Eric; Kozekova, Albena; Angel, Karen C.; Turesky, Robert J.; Stone, Michael P.; Guengerich, F. Peter; Rizzo, Carmelo J.

    2010-01-01

    2-Amino-3-methylimidazo[4,5-f]naphthelene is a carbocyclic analogue of the dietary carcinogen 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) in which a naphthalene ring system replaces the quinoline unit of IQ. The activity of 2-amino-3-methylimidazo[4,5-f]naphthelene (cIQ) in Ames Salmonella typhimurium tester strain TA98 is known to be 4-5 orders of magnitude lower than IQ. 2-Amino-3-methylimidazo[4,5-f]naphthelene undergoes efficient bioactivation with rat liver microsomes. The C8-dGuo adduct was formed when calf thymus DNA was treated with the N-hydroxy-cIQ metabolite and either acetic anhydride or extracts from cells that overexpress N-acetyl transferase (NAT). These studies indicate that bioactivation, the stability of the N-hydroxylamine ester, and the reactivity of the nitrenium ion with DNA of cIQ are similar to IQ and that none of these factors account for the differences in mutagenic potency of these analogues in Ames assays. Oligonucleotides were synthesized that contain the C8-dGuo adduct of 2-amino-3-methylimidazo[4,5-f]naphthelene in the frameshift-prone CG-dinucleotide repeat unit of the NarI recognition sequence. We have examined the in vitro translesion synthesis of this adduct and find it to be a strong replication block to E. coli DNA polymerase I, Klenow fragment exo− (Kf−), E. coli DNA polymerase II exo− (pol II−), and Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4). Previous studies by Fuchs and coworkers identified E. coli pol II as the polymerase responsible for two-base deletions of the C8-dGuo adduct of N-acetyl-2-aminofluorene in the NarI sequence. Our observation that pol II is strongly inhibited by the C8-dGuo adduct of 2-amino-3-methylimidazo[4,5-f]naphthelene suggests that one of the other SOS inducible polymerases (E. coli pol IV or pol V) is required for its bypass and this accounts for the greatly attenuated mutagenicity in Ames assays compared with IQ. PMID:20377178

  1. Therapeutic evaluation of grain based functional food formulation in a geriatric animal model.

    Science.gov (United States)

    Teradal, Deepa; Joshi, Neena; Aladakatti, Ravindranath H

    2017-08-01

    This study investigates the effect of wholesome grain based functional food formulation, on clinical and biochemical parameters in 24-30 months old Wistar albino geriatric rats, corresponding to human age 60-75 years. Animals were randomly divided into five, groups. Experimental diets were compared to the basal rat diet (Group I). Four food, formulation were-wheat based (Group II), finger millet based (Group III), wheat based, diet + fenugreek seed powder (Group IV), finger millet based diet + fenugreek powder, (Group V). These five types of diets were fed to the experimental rats for 6 weeks. Hematological and biochemical parameters were evaluated. The results showed that, feed intake was influenced by the type of feed. Diets supplemented with, fenugreek (Group IV) caused a significant increase in serum hemoglobin. The total serum protein values were significantly highest in Group III. Total serum albumin was found to be lower in Group I and highest in Group II. The concentration of BUN was highest in Group I and the lowest in control diet. Serum cholesterol and glucose were significantly reduced in Group IV. Several hematological and serum mineral values were influenced by the type of diet. The type of diet did not influence the organs weight. A moderate hypoglycemic and hypercholesterolemic effect was observed in composite mix fed rats. This study clearly justifies the recommendation to use wholesome grain based functional foods for geriatric population.

  2. Structural and Preclinical Studies of Computationally Designed Non-Nucleoside Reverse Transcriptase Inhibitors for Treating HIV infection

    Energy Technology Data Exchange (ETDEWEB)

    Kudalkar, Shalley N.; Beloor, Jagadish; Chan, Albert H.; Lee, Won-Gil; Jorgensen, William L.; Kumar, Priti; Anderson, Karen S.

    2017-02-06

    The clinical benefits of HIV-1 non-nucleoside reverse transcriptase (RT) inhibitors (NNRTIs) are hindered by their unsatisfactory pharmacokinetic (PK) properties along with the rapid development of drug-resistant variants. However, the clinical efficacy of these inhibitors can be improved by developing compounds with enhanced pharmacological profiles and heightened antiviral activity. We used computational and structure-guided design to develop two next-generation NNRTI drug candidates, compounds I and II, which are members of a class of catechol diethers. We evaluated the preclinical potential of these compounds in BALB/c mice because of their high solubility (510 µg/ml for compound I and 82.9 µg/ml for compound II), low cytotoxicity, and enhanced antiviral activity against wild-type (WT) HIV-1 RT and resistant variants. Additionally, crystal structures of compounds I and II with WT RT suggested an optimal binding to the NNRTI binding pocket favoring the high anti-viral potency. A single intraperitoneal dose of compounds I and II exhibited a prolonged serum residence time of 48 hours and concentration maximum (Cmax) of 4000- to 15,000-fold higher than their therapeutic/effective concentrations. These Cmax values were 4- to 15-fold lower than their cytotoxic concentrations observed in MT-2 cells. Compound II showed an enhanced area under the curve (0–last) and decreased plasma clearance over compound I and efavirenz, the standard of care NNRTI. Hence, the overall (PK) profile of compound II was excellent compared with that of compound I and efavirenz. Furthermore, both compounds were very well tolerated in BALB/c mice without any detectable acute toxicity. Taken together, these data suggest that compounds I and II possess improved anti-HIV-1 potency, remarkable in vivo safety, and prolonged in vivo circulation time, suggesting strong potential for further development as new NNRTIs for the potential treatment of HIV infection.

  3. Diagnosis of immunodeficiency caused by a purine nucleoside phosphorylase defect by using tandem mass spectrometry on dried blood spots.

    Science.gov (United States)

    la Marca, Giancarlo; Canessa, Clementina; Giocaliere, Elisa; Romano, Francesca; Malvagia, Sabrina; Funghini, Silvia; Moriondo, Maria; Valleriani, Claudia; Lippi, Francesca; Ombrone, Daniela; Della Bona, Maria Luisa; Speckmann, Carsten; Borte, Stephan; Brodszki, Nicholas; Gennery, Andrew R; Weinacht, Katja; Celmeli, Fatih; Pagel, Julia; de Martino, Maurizio; Guerrini, Renzo; Wittkowski, Helmut; Santisteban, Ines; Bali, Pawan; Ikinciogullari, Aydan; Hershfield, Michael; Notarangelo, Luigi D; Resti, Massimo; Azzari, Chiara

    2014-07-01

    Purine nucleoside phosphorylase (PNP) deficiency is a rare form of autosomal recessive combined primary immunodeficiency caused by a enzyme defect leading to the accumulation of inosine, 2'-deoxy-inosine (dIno), guanosine, and 2'-deoxy-guanosine (dGuo) in all cells, especially lymphocytes. Treatments are available and curative for PNP deficiency, but their efficacy depends on the early approach. PNP-combined immunodeficiency complies with the criteria for inclusion in a newborn screening program. This study evaluate whether mass spectrometry can identify metabolite abnormalities in dried blood spots (DBSs) from affected patients, with the final goal of individuating the disease at birth during routine newborn screening. DBS samples from 9 patients with genetically confirmed PNP-combined immunodeficiency, 10,000 DBS samples from healthy newborns, and 240 DBSs from healthy donors of different age ranges were examined. Inosine, dIno, guanosine, and dGuo were tested by using tandem mass spectrometry (TMS). T-cell receptor excision circle (TREC) and kappa-deleting recombination excision circle (KREC) levels were evaluated by using quantitative RT-PCR only for the 2 patients (patients 8 and 9) whose neonatal DBSs were available. Mean levels of guanosine, inosine, dGuo, and dIno were 4.4, 133.3, 3.6, and 3.8 μmol/L, respectively, in affected patients. No indeterminate or false-positive results were found. In patient 8 TREC levels were borderline and KREC levels were abnormal; in patient 9 TRECs were undetectable, whereas KREC levels were normal. TMS is a valid method for diagnosis of PNP deficiency on DBSs of affected patients at a negligible cost. TMS identifies newborns with PNP deficiency, whereas TREC or KREC measurement alone can fail. Copyright © 2014 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

  4. FLT3 is implicated in cytarabine transport by human equilibrative nucleoside transporter 1 in pediatric acute leukemia.

    Science.gov (United States)

    Català, Albert; Pastor-Anglada, Marçal; Caviedes-Cárdenas, Liska; Malatesta, Roberta; Rives, Susana; Vega-García, Nerea; Camós, Mireia; Fernández-Calotti, Paula

    2016-08-02

    FLT3 abnormalities are negative prognostic markers in acute leukemia. Infant leukemias are a subgroup with frequent MLL (KMT2A) rearrangements, FLT3 overexpression and high sensitivity to cytarabine, but dismal prognosis. Cytarabine is transported into cells by Human Equilibrative Nucleoside Transporter-1 (hENT1, SLC29A1), but the mechanisms that regulate hENT1 in acute leukemia have been scarcely studied.We explored the expression and functional link between FLT3 and main cytarabine transporters in 50 pediatric patients diagnosed with acute lymphoblastic leukemia and MLL rearrangement (ALL-MLL+) and other subtypes of leukemia, and in leukemia cell lines.A significant positive correlation was found between FLT3 and hENT1 expression in patients. Cytarabine uptake into cells was mediated mainly by hENT1, hENT2 and hCNT1. hENT1-mediated uptake of cytarabine was transiently abolished by the FLT3 inhibitor PKC412, and this effect was associated with decreased hENT1 mRNA and protein levels. Noticeably, the cytotoxicity of cytarabine was lower when cells were first exposed to FLT3 inhibitors (PKC412 or AC220), probably due to decreased hENT1 activity, but we observed a higher cytotoxic effect if FLT3 inhibitors were administered after cytarabine.FLT3 regulates hENT1 activity and thereby affects cytarabine cytotoxicity. The sequence of administration of cytarabine and FLT3 inhibitors is important to maintain their efficacy.

  5. Human equilibrative nucleoside transporter 1 and carcinoma of the ampulla of Vater: expression differences in tumour histotypes

    Directory of Open Access Journals (Sweden)

    G. Perrone

    2010-09-01

    Full Text Available The human equilibrative nucleoside transporter 1 (hENT1 is the major means by which gemcitabine enters human cells; recent evidence exists that hENT1 is expressed in carcinoma of the ampulla of Vater and that it should be considered as a molecular prognostic marker for patients with resected ampullary cancer. Aim of the present study is to evaluate the variations of hENT1 expression in ampullary carcinomas and to correlate such variations with histological subtypes and clinicopathological parameters. Forty-one ampullary carcinomas were histologically classified into intestinal, pancreaticobiliary and unusual types. hENT1 and Ki67 expression were evaluated by immunohistochemistry, and apoptotic cells were identified by the terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate biotin nick end labelling (TUNEL method. hENT1 overexpression was detected in 63.4% ampullary carcinomas. A significant difference in terms of hENT1 and Ki67 expression was found between intestinal vs. pancreaticobiliary types (P=0.03 and P=0.009 respectively. Moreover, a significant statistical positive correlation was found between apoptotic and proliferative Index (P=0.036, while no significant correlation was found between hENT1 and apoptosis. Our results on hENT1 expression suggest that classification of ampullary carcinoma by morphological subtypes may represent an additional tool in prospective clinical trials aimed at examining treatment efficacy; in addition, data obtained from Ki67 and TUNEL suggest a key role of hENT1 in tumour growth of ampullary carcinoma.

  6. Human equilibrative nucleoside transporter 1 gene expression is associated with gemcitabine efficacy in advanced leiomyosarcoma and angiosarcoma.

    Science.gov (United States)

    Vincenzi, Bruno; Stacchiotti, Silvia; Collini, Paola; Pantano, Francesco; Rabitti, Carla; Perrone, Giuseppe; Iuliani, Michele; Baldi, Alfonso; Badalamenti, Giuseppe; Sanfilippo, Roberta; Santini, Daniele; Muda, Andrea Onetti; Gronchi, Alessandro; Casali, Paolo; Dei Tos, Angelo Paolo; Tonini, Giuseppe

    2017-07-25

    The expression of human equilibrative nucleoside transporter 1 (hENT1), the major gemcitabine transporter into cells, has been thoroughly investigated as a predictive marker of response to gemcitabine in pancreatic cancer and biliary tract cancers. Since gemcitabine is widely used in the treatment of leiomyosarcoma and angiosarcoma, we investigated the correlation between hENT1 expression and gemcitabine efficacy in these sarcoma subtypes. We retrospectively identified 71 patients affected by advanced angiosarcoma (26) or leiomyosarcoma (45) treated within five Italian referral centres for sarcoma; among them, 49 patients (15 angiosarcoma, 34 leiomyosarcoma) were treated with gemcitabine. All tumour samples were analysed for hENT1 expression by real-time PCR. Median 2 -ΔCt value was used as the cutoff to dichotomise patients into 'high' expression and 'low' expression groups. Kaplan-Meier analysis was performed to estimate progression-free survival (PFS) and overall survival (OS). We found a significant association between high hENT1 expression levels and favourable outcome in terms of PFS and OS compared to cases with low hENT1 expression in leiomyosarcoma treated with gemcitabine (PFS: 6.8 vs 3.2 months, P=0.004; OS: 14.9 vs 8.5 months, P=0.007). In addition, hENT1 overexpression correlated with a significant improvement in PFS (9.3 vs 4.5 months; P=0.02) and OS (20.6 vs 10.8 months; P=0.001) in angiosarcoma patients treated with gemcitabine. Our study suggests that higher hENT1 expression are associated to gemcitabine efficacy both in patients with advanced leiomyosarcoma and angiosarcoma.

  7. Multimers formed by the rotavirus nonstructural protein NSP2 bind to RNA and have nucleoside triphosphatase activity.

    Science.gov (United States)

    Taraporewala, Z; Chen, D; Patton, J T

    1999-12-01

    The nonstructural protein NSP2 is a component of rotavirus replication intermediates and accumulates in cytoplasmic inclusions (viroplasms), sites of genome RNA replication and the assembly of subviral particles. To better understand the structure and function of the protein, C-terminally His-tagged NSP2 was expressed in bacteria and purified to homogeneity. In its purified form, the protein did not exist as a monomer but rather was present as an 8S-10S homomultimer consisting of 6 +/- 2 subunits of recombinant NSP2 (rNSP2). As shown by gel mobility shift assays, the rNSP2 multimers bound to RNA in discrete cooperative steps to form higher-order RNA-protein complexes. The RNA-binding activity of the rNSP2 multimers was determined to be nonspecific and to have a strong preference for single-stranded RNA over double-stranded RNA, for which it displayed little affinity. Enzymatic analysis revealed that rNSP2 possessed an associated nucleoside triphosphatase (NTPase) activity in vitro, which in the presence of Mg(2+) catalyzed the hydrolysis of each of the four NTPs to NDPs with equal efficiency. Evidence indicating that the hydrolysis of NTP resulted in the covalent linkage of the gamma-phosphate to rNSP2 was obtained. Additional experiments showed that NSP2 expressed transiently in MA014 cells is phosphorylated. We propose that NSP2 functions as a molecular motor, catalyzing the packaging of viral mRNA into core-like replication intermediates through the energy derived from its NTPase activity.

  8. The effects of nucleoside analogue prophylactic treatment on HBV activation in HBcAb+ patients undergoing immunosuppressive therapy.

    Science.gov (United States)

    Liu, H-L; Zhao, Z; Yang, H; Liu, F-F; Liu, Q; Luo, Q; Yuan, Q; Chen, L-M; Zeng, A-Z

    2013-09-01

    We investigated the effects of prophylactic nucleoside analogue treatment on HBV activation in patients with antibodies against core antigen (HBcAb+) patients undergoing immunosuppressive therapy. Patients (113), who were HBcAb+, with various autoimmune diseases, undergoing immunosuppressive therapy, were divided into two groups. The control group, not treated with antivirals, and the prophylactic group, treated with antiviral drugs. The two groups were evaluated for changes in serum biochemical marker (alanine aminotransferase ALT), virological marker (HBV DNA) and for seroconversion. In the control group, the number of patients with an increase in ALT in patients with isolated HBcAb and HBcAb and antibodies against HBsAg (HBsAb +) were five (20.0%) and one (2.8%), respectively (P HBcAb+ subgroup and one case (2.8%) in HBsAb+/HBcAb+ subgroup (P HBcAb+ only population, six patients (24.0%) in the control group had an increase in HBV DNA compared with none in the antiviral prophylactic group (P HBcAb+ in the control group underwent an HBsAg seroconversion when receiving immunosuppressive therapy for 18 months, while none in the antiviral prophylactic group underwent reversion to HBsAg positivity (P = 0.4949). Under immunosuppressive condition, the risk of HBV activation was much higher in patients with HBcAb than in patients with both HBcAb and antibodies to HBsAb group. Antiviral prophylactic therapy could significantly reduce the risk of HBV reactivation. © 2013 John Wiley & Sons Ltd.

  9. The role of nucleoside/nucleotide transport and metabolism in the uptake and retention of 3'-fluoro-3'-deoxythymidine in human B-lymphoblast cells

    International Nuclear Information System (INIS)

    Plotnik, David A.; McLaughlin, Lena J.; Chan, Jenny; Redmayne-Titley, Joshua N.; Schwartz, Jeffrey L.

    2011-01-01

    Introduction: Recent studies in the human adenocarcinoma cell line A549 have identified cell growth-dependent equilibrative nucleoside transporter-1 (hENT1) as a modifier of 3'-fluoro-3'-deoxythymidine (FLT) uptake and retention. In the present study, we used the ability to isolate human lymphoblastoid clones deficient in thymidine kinase 1 (TK1) to study how metabolism and nucleoside transport influence FLT uptake and retention. Methods: Transport and metabolism of FLT were measured in the human lymphoblastoid cell line TK6 and in eight clones isolated from TK6. Four clones were TK1-proficient, while four were TK1-deficient. Both influx and efflux of FLT were measured under conditions where concentrative and equilibrative transport could be distinguished. Results: Sodium-dependent concentrative FLT transport dominated over equilibrative transport mechanisms and while inhibition of hENT1 reduced FLT uptake, there were no correlations between clonal variations in hENT1 levels and FLT uptake. There was an absolute requirement of TK1 for concentration of FLT in TK6 cells. FLT uptake reached a peak after 60 min of incubation with FLT after which intracellular levels of FLT and FLT metabolites declined. Efflux was rapid and was associated with reductions in FLT and each of its metabolites. Both FLT and FLT-monophosphate were found in the efflux buffer. Conclusions: Initial rates of FLT uptake were a function of both concentrative and equilibrative transporters. TK1 activity was an absolute requirement for the accumulation of FLT. Retention was dependent on nucleoside/nucleotide efflux and retrograde metabolism of FLT nucleotides.

  10. Determination of nucleoside analog mono-, di-, and tri-phosphates in cellular matrix by solid phase extraction and ultra-sensitive LC-MS/MS detection

    Science.gov (United States)

    Bushman, Lane R.; Kiser, Jennifer J.; Rower, Joseph E.; Klein, Brandon; Zheng, Jia-Hua; Ray, Michelle L.; Anderson, Peter L.

    2011-01-01

    An ultra-sensitive liquid chromatography tandem mass spectrometry (LC-MS/MS) assay was developed and validated to facilitate the assessment of clinical pharmacokinetics of nucleotide analogs from lysed intracellular matrix. The method utilized a strong anion exchange isolation of mono-(MP), di-(DP), and tri-phosphates (TP) from intracellular matrix. Each fraction was then dephosphorylated to the parent moiety yielding a molar equivalent to the original nucleotide analog intracellular concentration. The analytical portion of the methodology was optimized in specific nucleoside analog centric modes (i.e. tenofovir (TFV) centric, zidovudine (ZDV) centric), which included desalting/concentration by solid phase extraction and detection by LC-MS/MS. Nucleoside analog MP-, DP-, and TP- determined on the TFV centric mode of analysis include TFV, lamivudine (3TC), and emtricitibine (FTC). The quantifiable linear range for TFV was 2.50 to 2000 fmol/sample, and that for 3TC/FTC was 0.10 to 200 pmol/sample. Nucleoside analog MP-, DP-, and TP- determined on the ZDV centric mode of analysis included 3TC and ZDV. The quantifiable linear range for 3TC was 0.10 to 100 pmol/sample, and 5.00 to 2000 fmol/sample for ZDV. Stable labeled isotopic internal standards facilitated accuracy and precision in alternative cell matrices, which supported the intended use of the method for MP, DP, and TP determinations in various cell types. The method was successfully applied to clinical research samples generating novel intracellular information for TFV, FTC, ZDV, and 3TC nucleotides. This document outlines method development, validation, and application to clinical research. PMID:21715120

  11. Demonstration in vitro of inhibition in normal rat tissues yet stimulation in Jensen sarcoma cells of 5-fluorouracil anabolism by purine nucleosides

    International Nuclear Information System (INIS)

    Beltz, R.E.; Haddad-Zackrison, L.

    1986-01-01

    It has been shown previously that the ability of tumor cells to anabolize 5-fluorouracil (FUra) to nucleotides can often be enhanced by exposing the cells to various purine nucleosides. Increases in FUra cytotoxicity have been observed to accompany this enhancement. In the present study the effects of purine nucleosides on FUra anabolism in rat tumor cells and in normal rat tissues sensitive to FUra were compared. Pieces of small intestine (SI), bone marrow suspensions (BM) and Jensen tumor cells were incubated in culture medium at 37 0 for 1 hr in the presence (or absence) of a selected purine nucleoside, then (2- 14 C)FUra was added and the incubation was continued for another hr. Incorporation of radioactivity into the trichloroacetic acid-insoluble fraction in each case was determined as a measure of FUra anabolism. Inosine, adenosine and N 6 -methyl-adenosine, 1 mM, stimulated FUra incorporation into the acid-insoluble fraction 2-3 fold in the tumor cells but inhibited this incorporation 59-70% in SI and 31-70% in BM. Attempts to further suppress FUra anabolism in the normal tissues resulted in a maximal inhibition of 92% in SI, using 1 mM alloxanthine, and a maximal inhibition of 84% in BM, employing combined 1 mM alloxanthine and 1 mM 5-aminoimidazole-4-carbox-amide ribonucleoside. These data suggest ways of selectively altering FUra anabolism in normal tissue and in tumor tissue of the tumor-bearing rat to improve the therapeutic index of FUra

  12. Clustering of nucleosides in the presence of alkali metals: Biologically relevant quartets of guanosine, deoxyguanosine and uridine observed by ESI-MS/MS.

    Science.gov (United States)

    Aggerholm, Tenna; Nanita, Sergio C; Koch, Kim J; Cooks, R Graham

    2003-01-01

    Electrospray ionization (ESI) mass spectra of nucleosides, recorded in the presence of alkali metals, display alkali metal ion-bound quartets and other clusters that may have implications for understanding non-covalent interactions in DNA and RNA. The tetramers of guanosine and deoxyguanosine and also their metaclusters (clusters of clusters), cationized by alkali metals, were observed as unusually abundant magic number clusters. The observation of these species in the gas phase parallels previous condensed-phase studies, which show that guanine derivatives can form quartets and metaclusters of quartets in solution in the presence of metal cations. This parallel behavior and also internal evidence suggest that bonding in the guanosine tetramers involves the bases rather than the sugar units. The nucleobases thymine and uracil are known to form magic number pentameric adducts with K+, Cs+ and NH4+ in the gas phase. In sharp contrast, we now show that the nucleosides uridine and deoxythymidine do not form the pentameric clusters characteristic of the corresponding bases. More subtle effects of the sugars are evident in the fact that adenosine and cytidine form numerous higher order clusters with alkali metals, whereas deoxyadenosine and deoxycytidine show no clustering. It is suggested that hydrogen bonding between the bases in the tetramers of dG and rG are the dominant interactions in the clusters, hence changing the ribose group to deoxyribose (and vice versa) generally has little effect. However, the additional hydroxyl group of RNA nucleosides enhances the non-selective formation of higher-order aggregates for adenosine and cytidine and results in the lack of highly stable magic number clusters. Some clusters are the result of aggregation in the course of ionization (ESI) whereas others appear to be intrinsic to the solution being examined. Copyright 2003 John Wiley & Sons, Ltd.

  13. Introduction of 2-O-benzyl abasic nucleosides to the 3'-overhang regions of siRNAs greatly improves nuclease resistance.

    Science.gov (United States)

    Nagaya, Yuki; Kitamura, Yoshiaki; Shibata, Aya; Ikeda, Masato; Akao, Yukihiro; Kitade, Yukio

    2017-12-15

    Chemically modified siRNAs containing 2-O-benzyl-1-deoxy-d-ribofuranose (R H OBn ) in their 3'-overhang region were significantly more resistant towards serum nucleases than siRNAs possessing the natural nucleoside in this region. The knockdown efficacies and binding affinities of these modified siRNAs to the recombinant human Argonaute protein 2 (hAgo2) PAZ domain were comparable with that of siRNA with a thymidine dimer at the 3'-end. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Lactic Acidosis Complicating Metformin and Non-Nucleoside Reverse Transcriptase Inhibitor Combination Therapy: A Smoldering Threat in the Post-HAART Era.

    Science.gov (United States)

    Ortiz-Brizuela, Edgar; Pérez-Patrigeon, Santiago; Recillas-Gispert, Claudia; Gómez-Pérez, Francisco Javier

    2015-01-01

    Dear Editor, The prevalence of type 2 diabetes (DM-2) in HIV-infected patients and the concomitant use of metformin (MTF) and non-nucleoside reverse transcriptase inhibitors (NRTI) is rising. Through inhibition of NADH dehydrogenase and DNA pol-γ, both drugs hinder oxidative phosphorylation that may lead to lactic acidosis (LA). Among NRTIs, abacavir and tenofovir have the lowest mitochondrial toxicity, with only a few LA cases reported2-4. We describe here a case of MTF-associated LA (MALA) secondary to the interaction with NRTI.

  15. On the regularization of regional gravity field solutions in spherical radial base functions

    Science.gov (United States)

    Naeimi, Majid; Flury, Jakob; Brieden, Phillip

    2015-08-01

    Regional refinement of the gravity field models from satellite data using spherical radial base functions (SRBF) is an ill-posed problem. This is mainly due to the regional confinement of the data and the base functions, which leads to severe instabilities in the solutions. Here, this ill-posedness as well as the related regularization process are investigated. We compare three methods for the choice of the regularization parameter, which have been frequently used in gravity modelling. These methods are (1) the variance component estimation (VCE), (2) the generalized cross validation (GCV) and (3) the L-curve criterion. A particular emphasis is put on the impact of the SRBF type on the regularization parameter. To do this, we include two types of SRBF which are often used for regional gravity field modelling. These are the Shannon SRBF or the reproducing kernel and the Spline SRBF. The investigations are performed on two months of the real GOCE ultrasensitive gravity gradients over Central Africa and Amazon. The solutions are validated against a state-of-the-art global gravity solution. We conclude that if a proper regularization method is applied, both SRBF deliver more or less the same accuracy. We show that when the Shannon wavelet is used, the L-curve method gives the best results, while with the Spline kernel, the GCV outperforms the other two methods. Moreover, we observe that the estimated coefficients for the Spline kernel cannot be spatially interpreted. In contrast, the coefficients obtained for the Shannon wavelet reflect the energy of the recovered gravity field with a correlation factor of above 95 per cent. Therefore, when combined with the L-curve method, the Shannon SRBF is advantageous for regional gravity field estimation, since it is one of the simplest band-limited SRBF. In addition, it delivers promising solutions and the estimated coefficients represent the characteristics of the gravity field within the target region.

  16. Simultaneous Quantitation of Free Amino Acids, Nucleosides and Nucleobases in Sipunculus nudus by Ultra-High Performance Liquid Chromatography with Triple Quadrupole Mass Spectrometry.

    Science.gov (United States)

    Ge, Yahui; Tang, Yuping; Guo, Sheng; Liu, Xin; Zhu, Zhenhua; Zhang, Lili; Liu, Pei; Ding, Shaoxiong; Lin, Xiangzhi; Lin, Rurong; Duan, Jin-ao

    2016-03-25

    To evaluate the nutritional and functional value of Sipunculus nudus, a rapid, simple and sensitive analytical method was developed using ultra-high performance liquid chromatography coupled with a triple quadrupole mass detection in multiple-reaction monitoring mode for the simultaneous quantitative determination of 25 free amino acids and 16 nucleosides and nucleobases in S. nudus within 20 min, which was confirmed to be reproducible and accurate. The limits of detection (LODs) and quantification (LOQs) were between 0.003-0.229 μg/mL and 0.008-0.763 μg/mL for the 41 analytes, respectively. The established method was applied to analyze 19 batches of S. nudus samples from four habitats with two different processing methods. The results showed that S. nudus contained a variety of free amino acids, nucleosides and nucleobases in sufficient quantity and reasonable proportion. They also demonstrated that the contents of these compounds in different parts of S. nudus were significantly discriminating, which were in the order: (highest) coelomic fluid > body wall > intestine (lowest). The method is simple and accurate, and could serve as a technical support for establishing quality control of S. nudus and other functional seafoods. Moreover, the research results also laid foundation for further exploitation and development of S. nudus.

  17. [Efficacy of initial antiretroviral therapy based on lopinavir/ritonavir plus 2 nucleoside/nucleotide analogs in patients with human immunodeficiency virus type 1 infection].

    Science.gov (United States)

    Zamora, Laura; Gatell, José M

    2014-11-01

    Triple combination regimens consisting of lopinavir/ritonavir (LPV/r) plus 2 nucleoside/nucleotide analogs continue to be a valid option in initial antiretroviral therapy. Other protease inhibitors boosted with ritonavir (and in future with cobicistat) have been introduced, as well as other non-nucleoside analogs (rilpivirin) and 3 integrase inhibitors. None of the new regimens have shown superiority over LPV/r or comparisons are lacking. Therefore, regimens including LPV/r continue to be recommended as initial first-line or alternative strategies in most treatment guidelines. Dual combinations with LPV/r (plus raltegravir or lamivudine) are described in another article and can provide a similar response rate to triple combinations, better tolerance, and an improved cost-efficacy ratio, both for initial therapy and in simplification strategies. In contrast, LPV/r or darunavir/r monotherapy does not seem an acceptable option in treatment-naïve patients and is becoming increasingly less acceptable in simplification strategies. Copyright © 2014 Elsevier España, S.L.U. All rights reserved.

  18. Simultaneous Quantitation of Free Amino Acids, Nucleosides and Nucleobases in Sipunculus nudus by Ultra-High Performance Liquid Chromatography with Triple Quadrupole Mass Spectrometry

    Directory of Open Access Journals (Sweden)

    Yahui Ge

    2016-03-01

    Full Text Available To evaluate the nutritional and functional value of Sipunculus nudus, a rapid, simple and sensitive analytical method was developed using ultra-high performance liquid chromatography coupled with a triple quadrupole mass detection in multiple-reaction monitoring mode for the simultaneous quantitative determination of 25 free amino acids and 16 nucleosides and nucleobases in S. nudus within 20 min, which was confirmed to be reproducible and accurate. The limits of detection (LODs and quantification (LOQs were between 0.003–0.229 μg/mL and 0.008–0.763 μg/mL for the 41 analytes, respectively. The established method was applied to analyze 19 batches of S. nudus samples from four habitats with two different processing methods. The results showed that S. nudus contained a variety of free amino acids, nucleosides and nucleobases in sufficient quantity and reasonable proportion. They also demonstrated that the contents of these compounds in different parts of S. nudus were significantly discriminating, which were in the order: (highest coelomic fluid > body wall > intestine (lowest. The method is simple and accurate, and could serve as a technical support for establishing quality control of S. nudus and other functional seafoods. Moreover, the research results also laid foundation for further exploitation and development of S. nudus.

  19. A new sensitive 32P-postlabeling assay based on the specific enzymatic conversion of bulky DNA lesions to radiolabeled dinucleotides and nucleoside 5'-monophosphates

    International Nuclear Information System (INIS)

    Randerath, Kurt; Randerath, Erika; Danna, T.F.; Van Golen, K.L.; Putman, K.L.

    1989-01-01

    A new sensitive 32 P-postlabelling assay for DNA adducts has been developed. When DNA containing bulky adducts, X 1 , X 2 , .....X n , is digested with nuclease P1 at pH 5, normal nucleotides are released as 5'-monophosphates, pN, while adducts are excised as 5'-phosphorylated dinucleotides, pX i pN, because inter-nucleotide linkages on the 3' side of X resist attack by nuclease P1. Addition of prostatic acid phosphatase to such a digest results in 5'-dephosphorylation of the nucleotides to normal nucleosides, N, and adducted dinucleotides, X i pN, carrying a 5'-terminal free hydroxyl group. The dinucleotides but not nucleosides are converted to 5'- 32 P-labeled dinucleotides,[ 32 P]pX i pN, by T4 polynucleotide kinase-catalyzed [ 32 P]posphate transfer from [γ- 32 P]ATP. Upon mapping on polyethyleneimine-cellulose anion-exchange TLC, the labeled dinucleotide adducts produce characteristic autoradiographic fingerprints. Alternatively, they are further digested with snake venom phosphodiesterase to yield 5'-monophosphates, [ 32 P]pX i and pN. TLC profiles of the monophosphate adducts are distinct from those of the dinucleotides. These reactions provide the basis of the new 32 P-postlabeling scheme, which is compared in this paper with a previously reported protocol yielding adducts in the form of 5'- 32 P-labeled 3',5'-bisphosphates, [ 32 P]pX i p. (author)

  20. Capillary electrophoresis coupled to mass spectrometry employing hexafluoro-2-propanol for the determination of nucleosides and nucleotide mono-, di- and tri-phosphates in baby foods.

    Science.gov (United States)

    Mateos-Vivas, María; Domínguez-Álvarez, Javier; Rodríguez-Gonzalo, Encarnación; Carabias-Martínez, Rita

    2017-10-15

    The present work describes a method for the simultaneous determination of unmodified nucleosides and nucleotide mono-, di- and tri-phosphates by capillary electrophoresis coupled to mass spectrometry (CE-MS). The use of hexafluoro-2-propanol (HFIP) in the separation medium, and as an additive to the sheath liquid of the electrospray interface (ESI), generated a highly efficient and sensitive method. Instrumental limits of detection in the range of 14-53ngmL -1 for nucleosides and 7-23, 20-49 and 64-124ngmL -1 for nucleotide mono-, di-, and tri-phosphates, respectively, were found. Sample treatment involved diluting an aliquot of baby food with ultra-high quality water and applying centrifugation-assisted ultrafiltration (CUF). The proposed method was validated and used to analyse a variety of baby food samples (16 in total) such as fish, meat, fruits, and baby dairy desserts that may endogenously contain these analytes. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Expression, purification, immunogenicity and protective efficacy of a recombinant nucleoside hydrolase from Leishmania donovani, a vaccine candidate for preventing cutaneous leishmaniasis.

    Science.gov (United States)

    McAtee, C Patrick; Seid, Christopher A; Hammond, Molly; Hudspeth, Elissa; Keegan, Brian P; Liu, Zhuyun; Wei, Junfei; Zhan, Bin; Arjona-Sabido, Raul; Cruz-Chan, Vladimir; Dumonteil, Eric; Hotez, Peter J; Bottazzi, Maria Elena

    2017-02-01

    The nucleoside hydrolase gene from Leishmania donovani was cloned and expressed in Escherichia coli as a full length 36-kDa protein (LdNH36). Following lysis and extraction, the protein was purified by anion exchange and gel filtration chromatography. The purified protein had a molecular mass of approximately 36-kDa and was confirmed to be >99% pure. Using a nucleoside hydrolase assay, the protein was found to exhibit a Km of 741 ± 246 μM. Protein integrity was confirmed by lithium dodecyl sulfate polyacrylamide gel electrophoresis (LDS-PAGE), mass spectrometry (MS), and enzymatic assay. Analysis of antibody levels from immunized mice indicated that LdNH36 alone or in a stable emulsion with the Toll-like receptor-4 ligand glucopyranosyl lipid adjuvant (GLA-SE) as immunostimulant induced high levels of antigen-specific IgG antibodies. The cellular immune response indicated a T h 1 response in mice immunized with LdNH36, but only when formulated with GLA-SE. Mice immunized with the LdNH36 antigen in combination with the GLA-SE adjuvant and challenged with Leishmania mexicana showed significant reductions (>20 fold) in parasite burden, confirming the protective efficacy of this vaccine candidate. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. Synthesis and antiviral properties of novel 7-heterocyclic substituted 7-deaza-adenine nucleoside inhibitors of Hepatitis C NS5B polymerase.

    Science.gov (United States)

    Di Francesco, M Emilia; Avolio, Salvatore; Pompei, Marco; Pesci, Silvia; Monteagudo, Edith; Pucci, Vincenzo; Giuliano, Claudio; Fiore, Fabrizio; Rowley, Michael; Summa, Vincenzo

    2012-08-01

    Previous investigations in our laboratories resulted in the discovery of a novel series of potent nucleoside inhibitors of Hepatitis C virus (HCV) NS5B polymerase bearing tetracyclic 7-substituted 7-deaza-adenine nucleobases. The planarity of such modified systems was suggested to play a role in the high inhibitory potency observed. This paper describes how we envisaged to maintain the desired planarity of the modified nucleobase by means of an intra-molecular H-bond, engaging a H-bond donor atom on an appropriately substituted 7-heterocyclic residue with the adjacent amino group of the nucleobase. The success of this strategy is reflected by the identification of several novel potent nucleoside inhibitors of HCV NS5B bearing a 7-heterocyclic substituted 7-deaza-adenine nucleobase. Amongst these, the 1,2,4-oxadiazole analog 11 showed high antiviral potency against HCV replication in replicon cells and efficient conversion to the corresponding NTP in vivo, with high and sustained levels of NTP measured in rat liver following intravenous and oral administration. Copyright © 2012 Elsevier Ltd. All rights reserved.

  3. Selective fluorescent and fluorogenic substrates for purine-nucleoside phosphorylases from various sources, and direct fluorimetric determination of enzyme levels in human and animal blood

    International Nuclear Information System (INIS)

    Wierzchowski, Jacek; Ogiela, Maria; Iwanska, Beata; Shugar, David

    2002-01-01

    The decrease in fluorescence of 7-methylguanosine (m 7 Guo) accompanying its phosphorolysis is readily applicable to monitoring the level of purine nucleoside phosphorylase (PNP) in mammalian whole blood lysates, without prepurification of the latter. A 10-fold higher sensitivity is attained with the use of 8-azaguanine (8-azaGua), a non-reversible, highly fluorogenic, substrate in the reverse, synthetic pathway. Although its affinity for human PNP is low (K m =250 μM at pH 7), its high reaction rate and fluorescence quantum yield of the product, 8-azaguanosine (φ=0.55 at pH 9), permit direct fluorometric monitoring of nucleoside synthesis in 1000-fold diluted hemolysates. It is also quite selective for mammalian enzymes, 8-azaGua being a poor substrate for bacterial (E. coli) PNP. Specificity of the assay was confirmed by the competitive effects of the natural purines hypoxanthine and adenine, with IC 50 values comparable to their K m values. PNP levels in animal blood, assayed with both substrates, gave comparable results. With the exception of mice, PNP blood levels in animals were lower than in humans. We have also characterized specific fluorescent/fluorogenic substrates for bacterial (E. coli) PNP. The best of these is 2,6-diaminopurine riboside. Kinetic parameters for all the foregoing substrates are listed, and shown to be favorable for analytical applications

  4. Efficacy and safety of twice daily first-line ritonavir/indinavir plus double nucleoside combination therapy in HIV-infected individuals. German Ritonavir/Indinavir Study Group.

    Science.gov (United States)

    Rockstroh, J K; Bergmann, F; Wiesel, W; Rieke, A; Thiesen, A; Fätkenheuer, G; Oette, M; Carls, H; Fenske, S; Nadler, M; Knechten, H

    2000-06-16

    To evaluate the virological efficacy and safety of quadruple therapy with two nucleoside analogues and ritonavir (400 mg twice daily) plus indinavir (400 mg twice daily) combination in antiretroviral therapy-naive patients. An open-label, uncontrolled multicentre trial. Antiretroviral therapy-naive patients (n = 90) with high median baseline HIV RNA levels of 220,000 copies/ml (range, 36,000-2,943,000 copies/ml) and median CD4 cell count of 189 x 10(6)/l (range, 4-656 x 10(6)/l) were started on a twice daily regimen of either zidovudine/lamivudine (49%), stavudine/lamivudine (38%) or stavudine/didanosine (13%) plus ritonavir 400 mg twice daily and indinavir 400 mg twice daily combination therapy. CD4 cell counts and HIV RNA were determined at weeks 0, 4, 8, 12, 16, 20, and 24. Statistical analysis was performed on treatment as well as intent-to-treat, where missing values were accounted for as failure. In the intent-to-treat analysis at week 24, the proportion of patients with HIV RNA of preliminary data suggest that the protease inhibitor combination ritonavir/indinavir plus double nucleoside therapy appears to be effective and safe in short-term treatment (up to 24 weeks).

  5. AICA riboside both activates AMP-activated protein kinase and competes with adenosine for the nucleoside transporter in the CA1 region of the rat hippocampus.

    Science.gov (United States)

    Gadalla, Anne E; Pearson, Tim; Currie, Ailsa J; Dale, Nicholas; Hawley, Simon A; Sheehan, Mike; Hirst, Warren; Michel, Anton D; Randall, Andrew; Hardie, D Grahame; Frenguelli, Bruno G

    2004-03-01

    5-Aminoimidazole-4-carboxamide riboside (AICA riboside; Acadesine) activates AMP-activated protein kinase (AMPK) in intact cells, and is reported to exert protective effects in the mammalian CNS. In rat cerebrocortical brain slices, AMPK was activated by metabolic stress (ischaemia > hypoxia > aglycaemia) and AICA riboside (0.1-10 mm). Activation of AMPK by AICA riboside was greatly attenuated by inhibitors of equilibrative nucleoside transport. AICA riboside also depressed excitatory synaptic transmission in area CA1 of the rat hippocampus, which was prevented by an adenosine A1 receptor antagonist and reversed by application of adenosine deaminase. However, AICA riboside was neither a substrate for adenosine deaminase nor an agonist at adenosine receptors. We conclude that metabolic stress and AICA riboside both stimulate AMPK activity in mammalian brain, but that AICA riboside has an additional effect, i.e. competition with adenosine for uptake by the nucleoside transporter. This results in an increase in extracellular adenosine and subsequent activation of adenosine receptors. Neuroprotection by AICA riboside could be mediated by this mechanism as well as, or instead of, by AMPK activation. Caution should therefore be exercised in ascribing an effect of AICA riboside to AMPK activation, especially in systems where inhibition of adenosine re-uptake has physiological consequences.

  6. Design, synthesis, anticancer, antimicrobial activities and molecular docking studies of theophylline containing acetylenes and theophylline containing 1,2,3-triazoles with variant nucleoside derivatives.

    Science.gov (United States)

    Ruddarraju, Radhakrishnam Raju; Murugulla, Adharvana Chari; Kotla, Ravindar; Chandra Babu Tirumalasetty, Muni; Wudayagiri, Rajendra; Donthabakthuni, Shobha; Maroju, Ravichandar; Baburao, K; Parasa, Lakshmana Swamy

    2016-11-10

    A new series of theophylline containing acetylene derivatives (6a-6b and 7-13) and theophylline containing 1,2,3-triazoles with variant nucleoside derivatives (20-32) have been designed and synthesized. These compounds were screened for anticancer and antimicrobial activity. Further the computational docking and 2D QSAR were performed using MOE software to identify novel scaffolds. The results showed that compound 29 and 30 exhibit significant cytotoxic effect on all four cancer cells such as lung (A549), colon (HT-29), breast (MCF-7) and melanoma (A375) with IC50 values of 2.56, 2.19, 1.89, 4.89 μM and 3.57, 2.90, 2.10, 5.81 μM respectively. Whereas quite different results were observed for these compounds in antimicrobial studies. Compounds 11, 21 and 26 have exhibited significant minimum inhibitory concentrations (MIC) against Staphylococcus aureus, Bacillus cereus, Escherichia coli and Pseudomonas aeruginosa. The docking studies demonstrate that compound 27, 28, 29 and 30 have good dock score and binding affinities with various therapeutic targets in cancer cell proliferation. In addition these compounds have shown acceptable correlation with bioassay results in the regression plots generated in 2D QSAR models. This is the first report to demonstrate the theophylline containing acetylene derivatives and theophylline containing 1,2,3-triazole nucleoside hybrids as potential anticancer and antimicrobial agents with comprehensive in silico analysis. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  7. The nucleoside diphosphate kinase gene Nme3 acts as quantitative trait locus promoting non-Mendelian inheritance.

    Directory of Open Access Journals (Sweden)

    Hermann Bauer

    Full Text Available The t-haplotype, a variant form of the t-complex region on mouse chromosome 17, acts as selfish genetic element and is transmitted at high frequencies (> 95% from heterozygous (t/+ males to their offspring. This phenotype is termed transmission ratio distortion (TRD and is caused by the interaction of the t-complex responder (Tcr with several quantitative trait loci (QTL, the t-complex distorters (Tcd1 to Tcd4, all located within the t-haplotype region. Current data suggest that the distorters collectively impair motility of all sperm derived from t/+ males; t-sperm is rescued by the responder, whereas (+-sperm remains partially dysfunctional. Recently we have identified two distorters as regulators of RHO small G proteins. Here we show that the nucleoside diphosphate kinase gene Nme3 acts as a QTL on TRD. Reduction of the Nme3 dosage by gene targeting of the wild-type allele enhanced the transmission rate of the t-haplotype and phenocopied distorter function. Genetic and biochemical analysis showed that the t-allele of Nme3 harbors a mutation (P89S that compromises enzymatic activity of the protein and genetically acts as a hypomorph. Transgenic overexpression of the Nme3 t-allele reduced t-haplotype transmission, proving it to be a distorter. We propose that the NME3 protein interacts with RHO signaling cascades to impair sperm motility through hyperactivation of SMOK, the wild-type form of the responder. This deleterious effect of the distorters is counter-balanced by the responder, SMOK(Tcr, a dominant-negative protein kinase exclusively expressed in t-sperm, thus permitting selfish behaviour and preferential transmission of the t-haplotype. In addition, the previously reported association of NME family members with RHO signaling in somatic cell motility and metastasis, in conjunction with our data involving RHO signaling in sperm motility, suggests a functional conservation between mechanisms for motility control in somatic cells and

  8. Development and characterization of a soymilk Kefir-based functional beverage

    Directory of Open Access Journals (Sweden)

    Camilla Fernanda Godinho da SILVA

    2018-03-01

    Full Text Available Abstract Kefir is a fermented beverage that deserves special attention, since it has probiotic activity and unique sensory, nutritional, and therapeutic properties. Given that both kefir and soymilk are beneficial to human health, this study aimed to assess the physicochemical characteristics and acceptability of soymilk Kefir-based functional beverages (SKB properly inoculated with lactobacilli strain after 16h of incubation at 37 °C. It was monitored lactobacilli cell viability, yeasts count, pH, titratable acidity, lipids, proteins, ash, total solid, carbohydrates, caloric values and acceptability of the products. Additionally was conducted a shelf-life study of SKB added of peach-flavor. The lactobacilli cell count ranged from 7.0 to 8.0 Log10 CFUmL-1 and pH values from 4.5 to 4.6. SKB samples with higher soymilk kefir percent presented higher lactobacilli cell count and lower lipid, ash, total solid, carbohydrate and caloric value. Results showed similar preferential rates for the SKB up to 30% of added soymilk kefir. The functional peach-flavored beverage presented appropriate pH value (4.3 and high viable cells count (7.0 Log10 CFUmL-1 up to the 28th day of cold storage, showed high acceptability (94.5% and positive purchase intention (83.4% among consumers.

  9. Partial covariance based functional connectivity computation using Ledoit-Wolf covariance regularization.

    Science.gov (United States)

    Brier, Matthew R; Mitra, Anish; McCarthy, John E; Ances, Beau M; Snyder, Abraham Z

    2015-11-01

    Functional connectivity refers to shared signals among brain regions and is typically assessed in a task free state. Functional connectivity commonly is quantified between signal pairs using Pearson correlation. However, resting-state fMRI is a multivariate process exhibiting a complicated covariance structure. Partial covariance assesses the unique variance shared between two brain regions excluding any widely shared variance, hence is appropriate for the analysis of multivariate fMRI datasets. However, calculation of partial covariance requires inversion of the covariance matrix, which, in most functional connectivity studies, is not invertible owing to rank deficiency. Here we apply Ledoit-Wolf shrinkage (L2 regularization) to invert the high dimensional BOLD covariance matrix. We investigate the network organization and brain-state dependence of partial covariance-based functional connectivity. Although RSNs are conventionally defined in terms of shared variance, removal of widely shared variance, surprisingly, improved the separation of RSNs in a spring embedded graphical model. This result suggests that pair-wise unique shared variance plays a heretofore unrecognized role in RSN covariance organization. In addition, application of partial correlation to fMRI data acquired in the eyes open vs. eyes closed states revealed focal changes in uniquely shared variance between the thalamus and visual cortices. This result suggests that partial correlation of resting state BOLD time series reflect functional processes in addition to structural connectivity. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. A Robust, Water-Based, Functional Binder Framework for High-Energy Lithium-Sulfur Batteries.

    Science.gov (United States)

    Lacey, Matthew J; Österlund, Viking; Bergfelt, Andreas; Jeschull, Fabian; Bowden, Tim; Brandell, Daniel

    2017-07-10

    We report here a water-based functional binder framework for the lithium-sulfur battery systems, based on the general combination of a polyether and an amide-containing polymer. These binders are applied to positive electrodes optimised towards high-energy electrochemical performance based only on commercially available materials. Electrodes with up to 4 mAh cm -2 capacity and 97-98 % coulombic efficiency are achievable in electrodes with a 65 % total sulfur content and a poly(ethylene oxide):poly(vinylpyrrolidone) (PEO:PVP) binder system. Exchange of either binder component for a different polymer with similar functionality preserves the high capacity and coulombic efficiency. The improvement in coulombic efficiency from the inclusion of the coordinating amide group was also observed in electrodes where pyrrolidone moieties were covalently grafted to the carbon black, indicating the role of this functionality in facilitating polysulfide adsorption to the electrode surface. The mechanical properties of the electrodes appear not to significantly influence sulfur utilisation or coulombic efficiency in the short term but rather determine retention of these properties over extended cycling. These results demonstrate the robustness of this very straightforward approach, as well as the considerable scope for designing binder materials with targeted properties. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Isospecific Propylene Polymerization Behavior of Lewis Base Functionalized Unbridged Zirconocences under Bulk Conditions

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Seung Woong; Kim, Hwa Kyu; Kim, Seong Kyun; Do, Young Kyu [KAIST, Daejeon (Korea, Republic of); Kim, Tae Won; Lee, Min Hyung [Univ. of Ulsan, Ulsan (Korea, Republic of)

    2014-01-15

    Isospecific propylene polymerization behavior of meta- and para-Lewis base (E) functionalized unbridged zirconocenes ([1-(E{sub n}-Ph)-3,4-Me{sub 2}C{sub 5}H{sub 2}]{sub 2}ZrCl{sub 2}, E = NMe{sub 2}, OMe; n = 1 or 2) was investigated under bulk conditions. Catalytic activity of the zirconocenes, and molecular weight and isotacticity of polypropylenes are found to be dependent on the position and number of the Lewis base functional groups in the zirconocenes. All the crude polypropylenes possess a broad molecular weight distribution and multi-melting transitions, indicating an involvement of multi-catalytic active species in the polymerization. The highest [mmmm] value of an isotactic portion of the polypropylenes reached 89%, which is higher than that (85%) from the well-known C{sub 2}-symmetric EBIZr (rac-Et(Ind){sub 2}ZrCl{sub 2}) catalyst. These results support that the in situ generated, rigid rac-like cation-anion pair through the Lewis acid-base interactions between the functional groups of zirconocenes and methylaluminoxane anion is effective in the formation of isotactic polypropylene under bulk propylene polymerization conditions.

  12. Dynamic chest radiography: flat-panel detector (FPD) based functional X-ray imaging.

    Science.gov (United States)

    Tanaka, Rie

    2016-07-01

    Dynamic chest radiography is a flat-panel detector (FPD)-based functional X-ray imaging, which is performed as an additional examination in chest radiography. The large field of view (FOV) of FPDs permits real-time observation of the entire lungs and simultaneous right-and-left evaluation of diaphragm kinetics. Most importantly, dynamic chest radiography provides pulmonary ventilation and circulation findings as slight changes in pixel value even without the use of contrast media; the interpretation is challenging and crucial for a better understanding of pulmonary function. The basic concept was proposed in the 1980s; however, it was not realized until the 2010s because of technical limitations. Dynamic FPDs and advanced digital image processing played a key role for clinical application of dynamic chest radiography. Pulmonary ventilation and circulation can be quantified and visualized for the diagnosis of pulmonary diseases. Dynamic chest radiography can be deployed as a simple and rapid means of functional imaging in both routine and emergency medicine. Here, we focus on the evaluation of pulmonary ventilation and circulation. This review article describes the basic mechanism of imaging findings according to pulmonary/circulation physiology, followed by imaging procedures, analysis method, and diagnostic performance of dynamic chest radiography.

  13. Synthesis and antiviral activity of cyclopropyl-spirocarbocyclic adenosine, (4R,5S,6R,7R)-4-(6-amino-9H-purin-9-yl)-7-(hydroxymethyl)spiro[2.4]heptane-5,6-diol against hepatitis C virus.

    Science.gov (United States)

    Gadthula, Srinivas; Rawal, Ravindra K; Sharon, Ashoke; Wu, Dong; Korba, Brent; Chu, Chung K

    2011-07-01

    An efficient method was developed for the synthesis of 6-exocyclic methylene carbocyclic intermediate 4. The Simmons-Smith cyclopropanation protocol was applied on the 6-exocyclic methylene of intermediate 4 and demonstrated its utility for the synthesis of novel class of a spiro-carbocyclic nucleoside analog 8. The titled compound 8 demonstrated a significant antiviral activity against HCV with EC(50) values of 0.273 and 0.368 μM in genotypes 1A and 1B, respectively. Copyright © 2011 Elsevier Ltd. All rights reserved.

  14. Evaluation of Novel Acyclic Nucleoside Phosphonates against Human and Animal Gammaherpesviruses Revealed an Altered Metabolism of Cyclic Prodrugs upon Epstein-Barr Virus Reactivation in P3HR-1 Cells

    Czech Academy of Sciences Publication Activity Database

    Coen, N.; Duraffour, S.; Naesens, L.; Krečmerová, Marcela; Van Den Oord, J.; Snoeck, R.; Andrei, G.

    2013-01-01

    Roč. 87, č. 22 (2013), s. 12422-12432 ISSN 0022-538X R&D Projects: GA MPO FR-TI4/625 Institutional support: RVO:61388963 Keywords : acyclic nucleoside phosphonate * gammaherpesvirus * Epstein-Barr virus * Kaposi's sarcoma * HPMP-5-azaC * cidofovir Subject RIV: EE - Microbiology, Virology Impact factor: 4.648, year: 2013

  15. Use of nucleoside reverse transcriptase inhibitors and risk of myocardial infarction in HIV-infected patients enrolled in the D:A:D study: a multi-cohort collaboration

    DEFF Research Database (Denmark)

    Sabin, Caroline A; Worm, Signe W; Weber, Rainer

    2008-01-01

    BACKGROUND: Whether nucleoside reverse transcriptase inhibitors increase the risk of myocardial infarction in HIV-infected individuals is unclear. Our aim was to explore whether exposure to such drugs was associated with an excess risk of myocardial infarction in a large, prospective observational...

  16. A Modular Approach to Aryl-C-ribonucleosides via the Allylic Substitution and Ring-Closing Metathesis Sequence. A Stereocontrolled Synthesis of All Four alpha-/beta- and D-/L-C-Nucleoside Stereoisomers

    Czech Academy of Sciences Publication Activity Database

    Štambaský, J.; Kapras, V.; Štefko, Martin; Kysilka, O.; Hocek, Michal; Malkov, A. V.; Kočovský, P.

    2011-01-01

    Roč. 76, č. 19 (2011), s. 7781-7803 ISSN 0022-3263 R&D Projects: GA MŠk LC512; GA AV ČR IAA400550902 Institutional research plan: CEZ:AV0Z40550506 Keywords : C-nucleosides * allylic substitution * metathesis * dihydroxylation Subject RIV: CC - Organic Chemistry Impact factor: 4.450, year: 2011

  17. Synthesis of piperidine nucleosides

    Czech Academy of Sciences Publication Activity Database

    Kovačková, Soňa; Dračínský, Martin; Rejman, Dominik

    2007-01-01

    Roč. 101, č. 11 (2007), s. 950 ISSN 0009-2770. [Pokroky v organické, bioorganické a farmaceutické chemii /42./. 16.11.2007-18.11.2007, Liblice] R&D Projects: GA MŠk LC512; GA MŠk(CZ) LC06061; GA MŠk(CZ) LC06077; GA MŠk 2B06065; GA ČR GP203/02/D150; GA ČR GP204/05/P510 Institutional research plan: CEZ:AV0Z40550506 Keywords : piperidine * alkylation * nucleobase Subject RIV: CC - Organic Chemistry

  18. Statistically significant contrasts between EMG waveforms revealed using wavelet-based functional ANOVA

    Science.gov (United States)

    McKay, J. Lucas; Welch, Torrence D. J.; Vidakovic, Brani

    2013-01-01

    We developed wavelet-based functional ANOVA (wfANOVA) as a novel approach for comparing neurophysiological signals that are functions of time. Temporal resolution is often sacrificed by analyzing such data in large time bins, increasing statistical power by reducing the number of comparisons. We performed ANOVA in the wavelet domain because differences between curves tend to be represented by a few temporally localized wavelets, which we transformed back to the time domain for visualization. We compared wfANOVA and ANOVA performed in the time domain (tANOVA) on both experimental electromyographic (EMG) signals from responses to perturbation during standing balance across changes in peak perturbation acceleration (3 levels) and velocity (4 levels) and on simulated data with known contrasts. In experimental EMG data, wfANOVA revealed the continuous shape and magnitude of significant differences over time without a priori selection of time bins. However, tANOVA revealed only the largest differences at discontinuous time points, resulting in features with later onsets and shorter durations than those identified using wfANOVA (P < 0.02). Furthermore, wfANOVA required significantly fewer (∼¼×; P < 0.015) significant F tests than tANOVA, resulting in post hoc tests with increased power. In simulated EMG data, wfANOVA identified known contrast curves with a high level of precision (r2 = 0.94 ± 0.08) and performed better than tANOVA across noise levels (P < <0.01). Therefore, wfANOVA may be useful for revealing differences in the shape and magnitude of neurophysiological signals (e.g., EMG, firing rates) across multiple conditions with both high temporal resolution and high statistical power. PMID:23100136

  19. Applications of nucleoside-based molecular probes for the in vivo assessment of tumour biochemistry using positron emission tomography (PET

    Directory of Open Access Journals (Sweden)

    Leonard I. Wiebe

    2007-05-01

    Full Text Available Positron emission tomography (PET is a non-invasive nuclear imaging technique. In PET, radiolabelled molecules decay by positron emission. The gamma rays resulting from positron annihilation are detected in coincidence and mapped to produce three dimensional images of radiotracer distribution in the body. Molecular imaging with PET refers to the use of positron-emitting biomolecules that are highly specific substrates for target enzymes, transport proteins or receptor proteins. Molecular imaging with PET produces spatial and temporal maps of the target-related processes. Molecular imaging is an important analytical tool in diagnostic medical imaging, therapy monitoring and the development of new drugs. Molecular imaging has its roots in molecular biology. Originally, molecular biology meant the biology of gene expression, but now molecular biology broadly encompasses the macromolecular biology and biochemistry of proteins, complex carbohydrates and nucleic acids. To date, molecular imaging has focused primarily on proteins, with emphasis on monoclonal antibodies and their derivative forms, small-molecule enzyme substrates and components of cell membranes, including transporters and transmembrane signalling elements. This overview provides an introduction to nucleosides, nucleotides and nucleic acids in the context of molecular imaging.A tomografia por emissão de pósitrons (TEP é uma técnica de imagem não invasiva da medicina nuclear. A TEP utiliza moléculas marcadas com emissores de radiação beta positiva (pósitrons. As radiações gama medidas que resultam do aniquilamento dos pósitrons são detectadas por um sistema de coincidência e mapeadas para produzir uma imagem tridimensional da distribuição do radiotraçador no corpo. A imagem molecular com TEP refere-se ao uso de biomoléculas marcadas com emissor de pósitron que são substratos altamente específicos para alvos como enzimas, proteínas transportadoras ou receptores prot

  20. Evaluation of technetium-99m/rhenium labelled nucleoside analogues as potential radiotracers in oncology[Dissertation 17173

    Energy Technology Data Exchange (ETDEWEB)

    Desbouis, D

    2007-07-01

    Over the last decade, suicide gene therapy has emerged as a very promising method to treat cancer. This therapy consists of introducing new genetic material into the nucleus of cancer cells so that they express a therapeutic protein. The protein leads to a therapeutic effect upon interaction with a prodrug. The most widely used system is the combination of the enzyme Herpes Simplex Virus Thymidine Kinase of type 1 (HSV1-TK) with the nontoxic antiviral prodrug gancyclovir. The efficiency of protein expression is crucial for a successful therapy. Several Positron Emission Tomography (PET) tracers such as 9-[4-[{sup 18}F]fluoro-3-(hydroxymethyl)butyl]guanine [{sup 18}F]FHBG) or [{sup 124}I]iodo-2'-fluoro-2'-deoxy-l-{beta}-D-arabino-furanosyluracil ({sup 124}I]FIAU) have been shown to be suitable probes for reporting HSV1-TK expression. One aspect of this work was to develop Single Photon Emission Tomography (SPET) reporter probes based on the inexpensive technetium-99m whose physical properties are well-suited for diagnosis (t{sub 1/2} = 6.02 h; E{sub {gamma}} = 140 keV). A series of complexes was prepared by derivatizing the precursor 5'-amino-5'-deoxythymidine at position N5' in order to introduce spacers of various lengths ({approx} 0-30 aangstroem) carrying tridentate metal chelating entities such as iminodiacetic acid and picolylamine-N-monoacetic acid. The nucleoside derivatives were reacted with the precursors [ReBr{sub 3}(CO){sub 3}]{sup 2-} and [{sup 99m}Tc(OH{sub 2}){sub 3}(CO){sub 3}]{sup +} to form water-stable organometallic thymidine complexes. Unexpectedly, most of the compounds showed no inhibition of HSV1-TK but a mixed inhibition of the human cytosolic thymidine kinase with K{sub iu} values ranging from 4.4-334 {mu}M. Competitive inhibition of HSV1-TK was only observed for the thymidine analogue in which the base and the metal core were separated by a spacer of approximately 30 aangstroem length (K{sub i} = 16.3 {+-} 4.6 {mu

  1. New Palladium-Catalyzed Approaches to Heterocycles and Carbocycles

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Qinhua [Iowa State Univ., Ames, IA (United States)

    2004-12-19

    The tert-butylimines of o-(1-alkynyl)benzaldehydes and analogous pyridinecarbaldehydes have been cyclized under very mild reaction conditions in the presence of I2, ICl, PhSeCl, PhSCl and p-O2NC6H4SCl to give the corresponding halogen-, selenium- and sulfur-containing disubstituted isoquinolines and naphthyridines, respectively. Monosubstituted isoquinolines and naphthyridines have been synthesized by the metal-catalyzed ring closure of these same iminoalkynes. This methodology accommodates a variety of iminoalkynes and affords the anticipated heterocycles in moderate to excellent yields. The Pd(II)-catalyzed cyclization of 2-(1-alkynyl)arylaldimines in the presence of various alkenes provides an efficient way to synthesize a variety of 4-(1-alkenyl)-3-arylisoquinolines in moderate to excellent yields. The introduction of an ortho-methoxy group on the arylaldimine promotes the Pd-catalyzed cyclization and stabilizes the resulting Pd(II) intermediate, improving the yields of the isoquinoline products. Highly substituted naphthalenes have been synthesized by the palladium-catalyzed annulation of a variety of internal alkynes, in which two new carbon-carbon bonds are formed in a single step under relatively mild reaction conditions. This method has also been used to synthesize carbazoles, although a higher reaction temperature is necessary. The process involves arylpalladation of the alkyne, followed by intramolecular Heck olefination and double bond isomerization. This method accommodates a variety of functional groups and affords the anticipated highly substituted naphthalenes and carbazoles in good to excellent yields. Novel palladium migratiodarylation methodology for the synthesis of complex fused polycycles has been developed, in which one or more sequential Pd-catalyzed intramolecular migration processes involving C-H activation are employed. The chemistry works best with electron-rich aromatics, which is in agreement with the idea that these palladium-catalyzed C-H activation reactions parallel electrophilic aromatic substitution. A relatively efficient synthesis of cyclopropanes has been developed using palladium-catalyzed C-H activation chemistry, in which two new carbon-carbon bonds are formed in a single step. This method involves the palladium-catalyzed activation of relatively unreactive C-H bonds, and provides a very efficient way to synthesize cyclopropapyrrolo[1,2-a]indoles, analogues of the mitomycin antibiotics.

  2. Decomposition pathways and in vitro HIV inhibitory effects of isoddA pronucleotides: toward a rational approach for intracellular delivery of nucleoside 5'-monophosphates.

    Science.gov (United States)

    Valette, G; Pompon, A; Girardet, J L; Cappellacci, L; Franchetti, P; Grifantini, M; La Colla, P; Loi, A G; Périgaud, C; Gosselin, G; Imbach, J L

    1996-05-10

    The decomposition pathways and kinetics in various biological media and the in vitro anti-HIV-1 and anti-HIV-2 activities of four derivatives of the 5'-mononucleotide of isoddA incorporating carboxylate esterase-labile transient phosphate protecting groups are reported and compared: namely, two mononucleoside aryl phosphoramidate derivatives 1a,b and two mononucleoside phosphotriester derivatives incorporating two S-acyl-2-thioethyl groups 2a,b. All four compounds show better antiviral activity, compared to the parent nucleoside analog isoddA. The results highlight that both types of compounds act as pronucleotides, i.e. they exert their antiviral effect via intracellular delivery of the 5'-mononucleotide of isoddA. The results may give insights for the design of new more efficient pronucleotides.

  3. Evaluation of intensity and energy interaction parameters for the complexation of Pr(III) with selected nucleoside and nucleotide through absorption spectral studies

    Science.gov (United States)

    Bendangsenla, N.; Moaienla, T.; David Singh, Th.; Sumitra, Ch.; Rajmuhon Singh, N.; Indira Devi, M.

    2013-02-01

    The interactions of Pr(III) with nucleosides and nucleotides have been studied in different organic solvents employing absorption difference and comparative absorption spectrophotometry. The magnitudes of the variations in both energy and intensity interaction parameters were used to explore the degree of outer and inner sphere co-ordination, incidence of covalency and the extent of metal 4f-orbital involvement in chemical bonding. Various electronic spectral parameters like Slater-Condon (Fk), Racah (Ek), Lande parameter (ξ4f), Nephelauxatic ratio (β), bonding (b1/2), percentage covalency (δ) and intensity parameters like oscillator strength (P) and Judd Ofelt electronic dipole intensity parameter (Tλ, λ = 2, 4, 6) have been evaluated. The variation of these evaluated parameters were employed to interpret the nature of binding of Pr(III) with different ligands i.e. Adenosine/ATP in presence and absence of Ca2+.

  4. Enzymic determination of inorganic phosphates, organic phosphates and phosphate-liberating enzymes by use of nucleoside phosphorylase-xanthine oxidase (dehydrogenase)-coupled reactions.

    Science.gov (United States)

    de Groot, H; de Groot, H; Noll, T

    1985-01-01

    Coupled enzyme assays are described for measuring inorganic phosphates, organic phosphates and phosphate-liberating enzymes in biological material. The assays all determine Pi by its reaction with inosine, catalysed by nucleoside phosphorylase; this yields ribose 1-phosphate and hypoxanthine. The hypoxanthine is oxidized to uric acid by xanthine oxidase, and may be measured either by the absorbance of the uric acid, or by the formazan formed when a tetrazolium salt is used as the oxidant. The coupled enzyme assays are characterized by high sensitivity, quantitative utilization of phosphates and stoichiometric formation of the measurable products, measurement at pH 6.0-8.5, determination of phosphates within a single analytical step, and continuous measurement of phosphohydrolase activity in a corresponding rate assay. Examples include determinations of substrates such as Pi, PPi and AMP, and of enzymes such as 5'-nucleotidase, inorganic pyrophosphatase and glucose-6-phosphatase. Directions for further examples are given. PMID:2996493

  5. Synthesis of purin-2-yl and purin-6-yl-aminoglucitols as C-nucleosidic ATP mimics and biological evaluation as FGFR3 inhibitors.

    Science.gov (United States)

    Tak-Tak, Lotfi; Barbault, Florent; Maurel, François; Busca, Patricia; Le Merrer, Yves

    2011-04-01

    Two new series of C-nucleosidic ATP mimics have been synthesized using an efficient and versatile synthetic pathway. These compounds were designed as FGFR3 inhibitors using purine as a central scaffold. The two substituents, a polyhydroxylated ribose mimic and a lipophilic moiety, were linked either in position 2 or 6 of the purine ring in order to explore any possible binding mode. All the compounds were able to inhibit FGFR3 kinase activity at a concentration of 50 μM. Unexpectedly, the best inhibitor was found to be one of the synthetic intermediates 13 bearing an iodine atom in position 2. Docking studies have confirmed its location in the ATP binding site and revealed halogen bonding among key interactions. Copyright © 2011. Published by Elsevier Masson SAS.

  6. Synthesis of enantiomerically pure (purin-6-yl)phenylalanines and their nucleosides, a novel type of purine-amino acid conjugates.

    Science.gov (United States)

    Capek, Petr; Pohl, Radek; Hocek, Michal

    2005-09-30

    [Chemical reaction: See text] Enantiomerically or diastereomerically pure 4-(purin-6-yl)phenylalanines, a novel type of stable amino acid-purine conjugates, were synthesized by palladium-catalyzed cross-coupling reactions of protected 4-boronophenylalanines or 4-(trimethylstanyl)phenylalanines with diverse 6-halopurines (9-benzyl-6-halopurines and 9-(tetrahydropyran-2-yl)-6-halopurines as well as acyl- and silyl-protected 6-halopurine ribonucleosides and 2-deoxyribonucleosides). Free purine bases and nucleosides bearing (S)- or (R)-phenylalanine in position 6 were obtained after complete deprotection of the products of cross-coupling reactions. Reactivity trends for both of these cross-coupling and deprotection protocols have been compared in terms of practicability, efficiency, and stereoselectivity.

  7. Dietary nucleosides and nucleotides do not affect tumor incidence but reduce amyloidosis incidence in B6C3F1 mice irradiated with californium-252.

    Science.gov (United States)

    Yokoyama, Hiroomi; Fujiwara, Hiroshi; Watanabe, Hiromitsu

    2004-04-01

    We investigated the effects of a dietary mixture of nucleosides and nucleotides (NS) on the systemic incidence rates of postirradiation carcinogenesis and non-neoplastic lesions in mice. Five-week-old male B6C3F1 mice were fed AIN-76B Purified Diet supplemented with NS for 1 wk and 13 mo before and after irradiation of neutron with californium-252 ((252)Cf); specifically NS was added to the AIN-76B Purified Diet (without nucleotide) to obtain a final concentration of 0%, 0.5%, or 2.5% NS. A commercial stock diet was also given to mice, and half of the mice were irradiated. Both irradiated and non-irradiated mice were used for reference controls. The incidence of liver tumors in each NS group was lower than that in the reference control group (P 252)Cf irradiation.

  8. A multi-step process of viral adaptation to a mutagenic nucleoside analogue by modulation of transition types leads to extinction-escape.

    Science.gov (United States)

    Agudo, Rubén; Ferrer-Orta, Cristina; Arias, Armando; de la Higuera, Ignacio; Perales, Celia; Pérez-Luque, Rosa; Verdaguer, Nuria; Domingo, Esteban

    2010-08-26

    Resistance of viruses to mutagenic agents is an important problem for the development of lethal mutagenesis as an antiviral strategy. Previous studies with RNA viruses have documented that resistance to the mutagenic nucleoside analogue ribavirin (1-β-D-ribofuranosyl-1-H-1,2,4-triazole-3-carboxamide) is mediated by amino acid substitutions in the viral polymerase that either increase the general template copying fidelity of the enzyme or decrease the incorporation of ribavirin into RNA. Here we describe experiments that show that replication of the important picornavirus pathogen foot-and-mouth disease virus (FMDV) in the presence of increasing concentrations of ribavirin results in the sequential incorporation of three amino acid substitutions (M296I, P44S and P169S) in the viral polymerase (3D). The main biological effect of these substitutions is to attenuate the consequences of the mutagenic activity of ribavirin -by avoiding the biased repertoire of transition mutations produced by this purine analogue-and to maintain the replicative fitness of the virus which is able to escape extinction by ribavirin. This is achieved through alteration of the pairing behavior of ribavirin-triphosphate (RTP), as evidenced by in vitro polymerization assays with purified mutant 3Ds. Comparison of the three-dimensional structure of wild type and mutant polymerases suggests that the amino acid substitutions alter the position of the template RNA in the entry channel of the enzyme, thereby affecting nucleotide recognition. The results provide evidence of a new mechanism of resistance to a mutagenic nucleoside analogue which allows the virus to maintain a balance among mutation types introduced into progeny genomes during replication under strong mutagenic pressure.

  9. A multi-step process of viral adaptation to a mutagenic nucleoside analogue by modulation of transition types leads to extinction-escape.

    Directory of Open Access Journals (Sweden)

    Rubén Agudo

    2010-08-01

    Full Text Available Resistance of viruses to mutagenic agents is an important problem for the development of lethal mutagenesis as an antiviral strategy. Previous studies with RNA viruses have documented that resistance to the mutagenic nucleoside analogue ribavirin (1-β-D-ribofuranosyl-1-H-1,2,4-triazole-3-carboxamide is mediated by amino acid substitutions in the viral polymerase that either increase the general template copying fidelity of the enzyme or decrease the incorporation of ribavirin into RNA. Here we describe experiments that show that replication of the important picornavirus pathogen foot-and-mouth disease virus (FMDV in the presence of increasing concentrations of ribavirin results in the sequential incorporation of three amino acid substitutions (M296I, P44S and P169S in the viral polymerase (3D. The main biological effect of these substitutions is to attenuate the consequences of the mutagenic activity of ribavirin -by avoiding the biased repertoire of transition mutations produced by this purine analogue-and to maintain the replicative fitness of the virus which is able to escape extinction by ribavirin. This is achieved through alteration of the pairing behavior of ribavirin-triphosphate (RTP, as evidenced by in vitro polymerization assays with purified mutant 3Ds. Comparison of the three-dimensional structure of wild type and mutant polymerases suggests that the amino acid substitutions alter the position of the template RNA in the entry channel of the enzyme, thereby affecting nucleotide recognition. The results provide evidence of a new mechanism of resistance to a mutagenic nucleoside analogue which allows the virus to maintain a balance among mutation types introduced into progeny genomes during replication under strong mutagenic pressure.

  10. MD SIMULATION STUDIES TO INVESTIGATE ISO-ENERGETIC CONFORMATIONAL BEHAVIOUR OF MODIFIED NUCLEOSIDES M2G AND M22G PRESENT IN tRNA

    Directory of Open Access Journals (Sweden)

    Rohit S Bavi

    2013-02-01

    Full Text Available Modified nucleic acid bases are most commonly found in tRNA. These may contain modifications from simple methylation to addition of bulky groups. Methylation of the four canonical nucleotide bases at a wide variety of positions is particularly prominent among the known modification. Methylation of N2 group of guanine is a relatively common modification in tRNA and rRNA. N2-methylguanosine (m2G is the second most often encountered nucleoside in E. coli tRNAs. N2, N2-dimethylguanosine (m22G is found in the majority of eukaryotic tRNAs and involved in forming base pair interactions with adjacent bases. Hence, in order to understand the structural significance of these methylated nucleic acid bases we have carried out molecular dynamics simulation to see the salvation effect. The results obtained shows iso-energetic conformational behaviors for m2G and m22G. The simulation trajectory of m2G shows regular periodical fluctuations suggesting that m2G is equally stable as either s-cis or s-trans rotamers. The two rotamers of m2G may interact canonically or non-canonically with opposite base as s-trans m2G26:C/A/U44 and s-cis m2G26:A/U44. The free rotations around the C-N bond could be the possible reason for these iso-energetic conformations. Dimethylation of G has almost no influence on base pairing with either A or U. Thus, these results reveal that modified nucleosides m2G and m22G may play an important role to prevent tRNA from adopting the unusual mitochondrial like conformation.

  11. Structure of a mutant human purine nucleoside phosphorylase with the prodrug, 2-fluoro-2-deoxyadenosine and the cytotoxic drug, 2-fluoroadenine

    Energy Technology Data Exchange (ETDEWEB)

    Afshar, Sepideh; Sawaya, Michael R.; Morrison, Sherie L.; (UCLA); (HHMI); (UCL)

    2009-06-30

    A double mutant of human purine nucleoside phosphorylase (hDM) with the amino acid mutations Glu201Gln:Asn243Asp cleaves adenosine-based prodrugs to their corresponding cytotoxic drugs. When fused to an anti-tumor targeting component, hDM is targeted to tumor cells, where it effectively catalyzes phosphorolysis of the prodrug, 2-fluoro-2'-deoxyadenosine (F-dAdo) to the cytotoxic drug, 2-fluoroadenine (F-Ade). This cytotoxicity should be restricted only to the tumor microenvironment, because the endogenously expressed wild type enzyme cannot use adenosine-based prodrugs as substrates. To gain insight into the interaction of hDM with F-dAdo, we have determined the crystal structures of hDM with F-dAdo and F-Ade. The structures reveal that despite the two mutations, the overall fold of hDM is nearly identical to the wild type enzyme. Importantly, the residues Gln201 and Asp243 introduced by the mutation form hydrogen bond contacts with F-dAdo that result in its binding and catalysis. Comparison of substrate and product complexes suggest that the side chains of Gln201 and Asp243 as well as the purine base rotate during catalysis possibly facilitating cleavage of the glycosidic bond. The two structures suggest why hDM, unlike the wild-type enzyme, can utilize F-dAdo as substrate. More importantly, they provide a critical foundation for further optimization of cleavage of adenosine-based prodrugs, such as F-dAdo by mutants of human purine nucleoside phosphorylase.

  12. Guanine α-carboxy nucleoside phosphonate (G-α-CNP) shows a different inhibitory kinetic profile against the DNA polymerases of human immunodeficiency virus (HIV) and herpes viruses.

    Science.gov (United States)

    Balzarini, Jan; Menni, Michael; Das, Kalyan; van Berckelaer, Lizette; Ford, Alan; Maguire, Nuala M; Liekens, Sandra; Boehmer, Paul E; Arnold, Eddy; Götte, Matthias; Maguire, Anita R

    2017-07-15

    α-Carboxy nucleoside phosphonates (α-CNPs) are modified nucleotides that represent a novel class of nucleotide-competing reverse transcriptase (RT) inhibitors (NcRTIs). They were designed to act directly against HIV-1 RT without the need for prior activation (phosphorylation). In this respect, they differ from the nucleoside or nucleotide RTIs [N(t)RTIs] that require conversion to their triphosphate forms before being inhibitory to HIV-1 RT. The guanine derivative (G-α-CNP) has now been synthesized and investigated for the first time. The (L)-(+)-enantiomer of G-α-CNP directly and competitively inhibits HIV-1 RT by interacting with the substrate active site of the enzyme. The (D)-(-)-enantiomer proved inactive against HIV-1 RT. In contrast, the (+)- and (-)-enantiomers of G-α-CNP inhibited herpes (i.e. HSV-1, HCMV) DNA polymerases in a non- or uncompetitive manner, strongly indicating interaction of the (L)-(+)- and the (D)-(-)-G-α-CNPs at a location different from the polymerase substrate active site of the herpes enzymes. Such entirely different inhibition profile of viral polymerases is unprecedented for a single antiviral drug molecule. Moreover, within the class of α-CNPs, subtle differences in their sensitivity to mutant HIV-1 RT enzymes were observed depending on the nature of the nucleobase in the α-CNP molecules. The unique properties of the α-CNPs make this class of compounds, including G-α-CNP, direct acting inhibitors of multiple viral DNA polymerases. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Discovery of biphenyl-substituted diarylpyrimidines as non-nucleoside reverse transcriptase inhibitors with high potency against wild-type and mutant HIV-1.

    Science.gov (United States)

    Jin, KaiJun; Yin, Hong; De Clercq, Erik; Pannecouque, Christophe; Meng, Ge; Chen, FenEr

    2018-02-10

    A novel series of diarylpyrimidine (DAPY) derivatives bearing the biphenyl motif with multiple substituted groups was synthesized as human immunodeficiency virus (HIV)-1 non-nucleoside reverse transcriptase inhibitors. All of the target compounds were evaluated for their in vitro activity against HIV in MT-4 cells. Most of the compounds exhibited excellent activity with low nanomolar EC 50 values against wild-type, single and double mutant HIV-1 strains. Compound 4b displayed an EC 50 value of 1 nM against HIV-1 IIIB, 1.3 nM against L100I, 0.84 nM against K103 N, 1.5 nM against Y181C, 11 nM against Y188L, 2 nM against E138K, 10 nM against K103 N + Y181C, and almost 110 nM against F227L + V106. The improvement in the selectivity and potency of the target molecules against the wild-type and mutant HIV-1 strains validated our hypothesis. The biphenyl ring in the DAPY derivatives could strengthen the π-π stacking effect between the target molecule and the non-nucleoside inhibitor-binding pocket in the reverse transcriptase by extending the conjugating systems. This research represented a significant step toward the discovery of novel therapeutic DAPYs for treating acquired immunodeficiency syndrome in patients infected with HIV-1. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

  14. Using Trial-Based Functional Analysis to Design Effective Interventions for Students Diagnosed with Autism Spectrum Disorder

    Science.gov (United States)

    Larkin, Wallace; Hawkins, Renee O.; Collins, Tai

    2016-01-01

    Functional behavior assessments and function-based interventions are effective methods for addressing the challenging behaviors of children; however, traditional functional analysis has limitations that impact usability in applied settings. Trial-based functional analysis addresses concerns relating to the length of time, level of expertise…

  15. Using a Modified Pyramidal Training Model to Teach Special Education Teachers to Conduct Trial-Based Functional Analyses

    Science.gov (United States)

    Kunnavatana, S. Shanun; Bloom, Sarah E.; Samaha, Andrew L.; Lignugaris/Kraft, Benjamin; Dayton, Elizabeth; Harris, Shannon K.

    2013-01-01

    Functional behavioral assessments are commonly used in school settings to assess and develop interventions for problem behavior. The trial-based functional analysis is an approach that teachers can use in their classrooms to identify the function of problem behavior. The current study evaluates the effectiveness of a modified pyramidal training…

  16. Biochar-based functional materials in the purification of agricultural wastewater: Fabrication, application and future research needs.

    Science.gov (United States)

    Wei, Dongning; Li, Bingyu; Huang, Hongli; Luo, Lin; Zhang, Jiachao; Yang, Yuan; Guo, Jiajun; Tang, Lin; Zeng, Guangming; Zhou, Yaoyu

    2018-04-01

    Nowadays, agricultural contamination is becoming more and more serious due to the rapid growth of agricultural industry, which discharged antibiotics, pesticides or toxic metals into farmlands. A large number of researchers have applied biochar-based functional materials to the treatment of agricultural wastewater contamination. Meanwhile, biochar has also proved to be a very promising and effective technology in water purification field due to its various beneficial properties (e.g., cost effective, high specific surface area, and surface reactive groups). The focus of this review is to highlight the fabrication methods and application of biochar-based functional materials with the removal of different agricultural contaminants, and discuss the underlying mechanisms. However, the application of biochar-based functional materials is currently under its infancy, with the main hindrance is identified as the gap between laboratory scale and field application, immaturity of engineered biochar production technologies, and lack of quality standards. In order to fill these knowledge gaps, more efforts should be made to pay for the relevant research in future studies. Copyright © 2018 Elsevier Ltd. All rights reserved.

  17. Unsaturated fluoro-ketopyranosyl nucleosides: synthesis and biological evaluation of 3-fluoro-4-keto-beta-d-glucopyranosyl derivatives of N(4)-benzoyl cytosine and N(6)-benzoyl adenine.

    Science.gov (United States)

    Manta, Stella; Agelis, George; Botić, Tanja; Cencic, Avrelija; Komiotis, Dimitri

    2008-02-01

    The protected beta-nucleosides 1-(2,4,6-tri-O-acetyl-3-deoxy-3-fluoro-beta-d-glucopyranosyl)-N(4)-benzoyl cytosine (2a) and 9-(2,4,6-tri-O-acetyl-3-deoxy-3-fluoro-beta-d-glucopyranosyl)-N(6)-benzoyl adenine (2b), were synthesized by the coupling of peracetylated 3-deoxy-3-fluoro-d-glucopyranose (1) with silylated N(4)-benzoyl cytosine and N(6)-benzoyl adenine, respectively. The nucleosides were deacetylated and several subsequent protection and deprotection steps afforded the partially acetylated nucleosides of cytosine 7a and adenine 7b, respectively. Finally, direct oxidation of the free hydroxyl group at 4'-position of 7a and 7b, and simultaneous elimination reaction of the beta-acetoxyl group, afforded the desired unsaturated 3-fluoro-4-keto-beta-d-glucopyranosyl derivatives. These newly synthesized compounds were evaluated for their potential antitumor and antiviral activities. Compared to 5FU, the newly synthesized derivatives showed to be more efficient as antitumor growth inhibitors and they exhibited direct antiviral effect toward rotavirus.

  18. Structures of bacterial polynucleotide kinase in a michaelis complex with nucleoside triphosphate (NTP)-Mg2+ and 5'-OH RNA and a mixed substrate-product complex with NTP-Mg2+ and a 5'-phosphorylated oligonucleotide.

    Science.gov (United States)

    Das, Ushati; Wang, Li Kai; Smith, Paul; Munir, Annum; Shuman, Stewart

    2014-12-01

    Clostridium thermocellum polynucleotide kinase (CthPnk), the 5'-end-healing module of a bacterial RNA repair system, catalyzes reversible phosphoryl transfer from a nucleoside triphosphate (NTP) donor to a 5'-OH polynucleotide acceptor, either DNA or RNA. Here we report the 1.5-Å crystal structure of CthPnk-D38N in a Michaelis complex with GTP-Mg(2+) and a 5'-OH RNA oligonucleotide. The RNA-binding mode of CthPnk is different from that of the metazoan RNA kinase Clp1. CthPnk makes hydrogen bonds to the ribose 2'-hydroxyls of the 5' terminal nucleoside, via Gln51, and the penultimate nucleoside, via Gln83. The 5'-terminal nucleobase is sandwiched by Gln51 and Val129. Mutating Gln51 or Val129 to alanine reduced kinase specific activity 3-fold. Ser37 and Thr80 donate functionally redundant hydrogen bonds to the terminal phosphodiester; a S37A-T80A double mutation reduced kinase activity 50-fold. Crystallization of catalytically active CthPnk with GTP-Mg(2+) and a 5'-OH DNA yielded a mixed substrate-product complex with GTP-Mg(2+) and 5'-PO4 DNA, wherein the product 5' phosphate group is displaced by the NTP γ phosphate and the local architecture of the acceptor site is perturbed. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  19. Functional and Structural Characterization of Purine Nucleoside Phosphorylase from Kluyveromyces lactis and Its Potential Applications in Reducing Purine Content in Food.

    Science.gov (United States)

    Mahor, Durga; Priyanka, Anu; Prasad, Gandham S; Thakur, Krishan Gopal

    2016-01-01

    Consumption of foods and beverages with high purine content increases the risk of hyperuricemia, which causes gout and can lead to cardiovascular, renal, and other metabolic disorders. As patients often find dietary restrictions challenging, enzymatically lowering purine content in popular foods and beverages offers a safe and attractive strategy to control hyperuricemia. Here, we report structurally and functionally characterized purine nucleoside phosphorylase (PNP) from Kluyveromyces lactis (KlacPNP), a key enzyme involved in the purine degradation pathway. We report a 1.97 Å resolution crystal structure of homotrimeric KlacPNP with an intrinsically bound hypoxanthine in the active site. KlacPNP belongs to the nucleoside phosphorylase-I (NP-I) family, and it specifically utilizes 6-oxopurine substrates in the following order: inosine > guanosine > xanthosine, but is inactive towards adenosine. To engineer enzymes with broad substrate specificity, we created two point variants, KlacPNPN256D and KlacPNPN256E, by replacing the catalytically active Asn256 with Asp and Glu, respectively, based on structural and comparative sequence analysis. KlacPNPN256D not only displayed broad substrate specificity by utilizing both 6-oxopurines and 6-aminopurines in the order adenosine > inosine > xanthosine > guanosine, but also displayed reversal of substrate specificity. In contrast, KlacPNPN256E was highly specific to inosine and could not utilize other tested substrates. Beer consumption is associated with increased risk of developing gout, owing to its high purine content. Here, we demonstrate that KlacPNP and KlacPNPN256D could be used to catalyze a key reaction involved in lowering beer purine content. Biochemical properties of these enzymes such as activity across a wide pH range, optimum activity at about 25°C, and stability for months at about 8°C, make them suitable candidates for food and beverage industries. Since KlacPNPN256D has broad substrate specificity, a

  20. Etravirine and rilpivirine resistance in HIV-1 subtype CRF01_AE-infected adults failing non-nucleoside reverse transcriptase inhibitor-based regimens.

    Science.gov (United States)

    Bunupuradah, Torsak; Ananworanich, Jintanat; Chetchotisakd, Ploenchan; Kantipong, Pacharee; Jirajariyavej, Supunnee; Sirivichayakul, Sunee; Munsakul, Warangkana; Prasithsirikul, Wisit; Sungkanuparph, Somnuek; Bowonwattanuwong, Chureeratana; Klinbuayaem, Virat; Petoumenos, Kathy; Hirschel, Bernard; Bhakeecheep, Sorakij; Ruxrungtham, Kiat

    2011-01-01

    We studied prevalence of etravirine (ETR) and rilpivirine (RPV) resistance in HIV-1 subtype CRF01_AE infection with first-line non-nucleoside reverse transcriptase inhibitor (NNRTI) failure. A total of 225 adults failing two nucleoside reverse transcriptase inhibitors (NRTIs) plus 1 NNRTI in Thailand with HIV RNA>1,000 copies/ml were included. Genotypic resistance results and HIV-1 subtype were interpreted by Stanford DR database. ETR resistance was calculated by the new Monogram weighted score (Monogram WS; ≥ 4 indicating high-level ETR resistance) and by DUET weighted score (DUET WS; 2.5-3.5 and ≥ 4 resulted in intermediate and reduce ETR response, respectively). RPV resistance interpretation was based on previous reports. Median (IQR) age was 38 (34-42) years, 41% were female and CDC A:B:C were 22%:21%:57%. HIV subtypes were 96% CRF01_AE and 4% B. Antiretrovirals at failure were lamivudine (100%), stavudine (93%), nevirapine (90%) and efavirenz (10%) with a median (IQR) duration of 3.4 (1.8-4.5) years. Median (IQR) CD4(+) T-cell count and HIV RNA were 194 (121-280) cells/mm³ and 4.1 (3.6-4.6) log₁₀ copies/ml, respectively. The common NNRTI mutations were Y181C (41%), G190A (22%) and K103N (19%). The proportion of patients with Monogram WS score ≥ 4 was 61.3%. By DUET WS, 49.8% and 7.5% of patients were scored 2.5-3.5 and ≥4, respectively. Only HIV RNA ≥ 4 log₁₀ copies/ml at failure was associated with both Monogram WS ≥ 4 (OR 2.3, 95% CI 1.3-3.9; P=0.003) and DUET WS ≥ 2.5 (OR 1.9, 95% CI 1.1-3.3; P=0.02). The RVP resistance-associated mutations (RAMs) detected were K101P (1.8%), Y181I (2.7%) and Y181V (3.6%). All patients with RPV mutation had ETR resistance. No E138R/E138K mutations were detected. Approximately 60% of patients had high-level ETR resistance. The role of ETR in second-line therapy is limited in late NNRTI failure settings. RVP RAMs were uncommon, but cross-resistance between ETR and RVP was high.

  1. Organometallic nucleosides induce non-classical leukemic cell death that is mitochondrial-ROS dependent and facilitated by TCL1-oncogene burden.

    Science.gov (United States)

    Prinz, Christian; Vasyutina, Elena; Lohmann, Gregor; Schrader, Alexandra; Romanski, Steffen; Hirschhäuser, Christoph; Mayer, Petra; Frias, Corazon; Herling, Carmen D; Hallek, Michael; Schmalz, Hans-Günther; Prokop, Aram; Mougiakakos, Dimitrios; Herling, Marco

    2015-06-04

    Redox stress is a hallmark of the rewired metabolic phenotype of cancer. The underlying dysregulation of reactive oxygen species (ROS) is interconnected with abnormal mitochondrial biogenesis and function. In chronic lymphocytic leukemia (CLL), elevated ROS are implicated in clonal outgrowth and drug resistance. The pro-survival oncogene T-cell leukemia 1 (TCL1) is causally linked to the high threshold towards classical apoptosis in CLL. We investigated how aberrant redox characteristics and bioenergetics of CLL are impacted by TCL1 and if this is therapeutically exploitable. Bio-organometallic chemistry provided compounds containing a cytosine nucleobase, a metal core (ferrocene, ruthenocene, Fe(CO)3), and a 5'-CH2O-TDS substituent. Four of these metal-containing nucleoside analogues (MCNA) were tested for their efficacy and mode of action in CLL patient samples, gene-targeted cell lines, and murine TCL1-transgenic splenocytes. The MCNA showed a marked and selective cytotoxicity towards CLL cells. MCNA activity was equally observed in high-risk disease groups, including those of del11q/del17p cytogenetics and of clinical fludarabine resistance. They overcame protective stromal cell interactions. MCNA-evoked PARP-mediated cell death was non-autophagic and non-necrotic as well as caspase- and P53-independent. This unconventional apoptosis involved early increases of ROS, which proved indispensible based on mitigation of MCNA-triggered death by various scavengers. MCNA exposure reduced mitochondrial respiration (oxygen consumption rate; OCR) and induced a rapid membrane depolarization (∆ΨM). These characteristics distinguished the MCNA from the alkylator bendamustine and from fludarabine. Higher cellular ROS and increased MCNA sensitivity were linked to TCL1 expression. The presence of TCL1 promoted a mitochondrial release of in part caspase-independent apoptotic factors (AIF, Smac, Cytochrome-c) in response to MCNA. Although basal mitochondrial respiration (OCR

  2. Synthesis and biological evaluation of nucleoside analogues than contain silatrane on the basis of the structure of acyclovir (ACV) as novel inhibitors of hepatitis B virus (HBV).

    Science.gov (United States)

    Han, Anyue; Li, Lingna; Qing, Kuiyou; Qi, Xiaolu; Hou, Leping; Luo, Xintong; Shi, Shaohua; Ye, Faqing

    2013-03-01

    Hepatitis B virus (HBV) infection causes major public health problems worldwide. Acyclovir (ACV) is mainly used to inhibit herpes simplex virus (HSV) rather than HBV. In this study, we used the combination principle to design and synthesize nucleoside analogues that contain silatrane on the basis of the structure of ACV. We found that the compounds were effective inhibitors of HBV, both in vitro and in vivo. All of the compounds showed suppressive activity on the expression of HBV surface antigen (HBsAg) and HBV e antigen (HBeAg) in the HepG2.2.15 cell line with low cytotoxicity. One of compounds was studied in HBV transgenic mice model, and the test results showed its ability to reduce the levels of HBsAg, HBeAg and HBV DNA by ELASE and qPCR. Furthermore, significant improvement of T lymphocyte was observed after treatment, as evaluated by flow cytometry (FCM). Copyright © 2013 Elsevier Ltd. All rights reserved.

  3. Identification of 3,N(4)-etheno-5-methyl-2'-deoxycytidine in human DNA: a new modified nucleoside which may perturb genome methylation.

    Science.gov (United States)

    Nair, Jagadeesan; Godschalk, Roger W; Nair, Urmila; Owen, Robert W; Hull, William E; Bartsch, Helmut

    2012-01-13

    Methylation of cytidine at dCpdG sequences regulates gene expression and is altered in many chronic inflammatory diseases. Inflammation generates lipid peroxidation (LPO) products which can react with deoxycytidine, deoxyadenosine, and deoxyguanosine in DNA to form pro-mutagenic exocyclic etheno-nucleoside residues. Since 5-methyl-2'-deoxycytidine (5mdC) residues exhibit increased nucleophilicity at N3, they should be even better targets for LPO products. We synthesized and characterized 3,N(4)-etheno-5-methyl-2'-deoxycytidine-3'-phosphate and showed that LPO products can indeed form the corresponding etheno-5mdC (ε5mdC) lesion in DNA in vitro. Our newly developed (32)P-postlabeling method was subsequently used to detect ε5mdC lesions in DNA from human white blood cells, lung, and liver at concentrations 4-10 times higher than that observed for etheno adducts on nonmethylated cytidine. Our new detection method can now be used to explore the hypothesis that this DNA lesion perturbs the DNA methylation status.

  4. Design, synthesis, and biological evaluation of new 1,2,3-triazolo-2'-deoxy-2'-fluoro- 4'-azido nucleoside derivatives as potent anti-HBV agents.

    Science.gov (United States)

    Liu, Yuan; Peng, Youmei; Lu, Jingjing; Wang, Jingwen; Ma, Haoran; Song, Chuanjun; Liu, Bingjie; Qiao, Yan; Yu, Wenquan; Wu, Jie; Chang, Junbiao

    2018-01-01

    Novel drugs are urgently needed to combat hepatitis B virus (HBV) infection due to drug-resistant virus. In this paper, a series of novel 4-monosubstituted 2'-deoxy-2'-β-fluoro-4'-azido-β-d-arabinofuranosyl 1,2,3-triazole nucleoside analogues (1a-g) were designed, synthesized and screened for in vitro anti-HBV activity. At 5.0 μM in the cellular model, all the synthetic compounds display activities comparable to that of the positive control, lamivudine at 20 μM. Of the compounds tested, the amide-substituted analogue (1a) shows the most promising anti-HBV activity and low cytotoxicity in the cell model. In particular, it retains excellent activity against lamivudine-resistant HBV mutants. In duck HBV (DHBV)-infected duck models, both the serum and liver DHBV DNA levels (67.4% and 53.3%, respectively) were reduced markedly by the treatment with 1a. Analysis of the structure of HBV polymer/1a-triphosphate (1a-TP) complex shows that 1a-TP is stabilized by specific van der Waals interactions with the enzyme residues arising from 4-amino-1,2,3-triazole and the 4'-azido group. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  5. Effect of dietary nucleosides and yeast extracts on composition and metabolic activity of infant gut microbiota in PolyFermS colonic fermentation models.

    Science.gov (United States)

    Doo, Eun-Hee; Chassard, Christophe; Schwab, Clarissa; Lacroix, Christophe

    2017-08-01

    Nucleotides (NT) and nucleosides (NS) are added to infant formula to mimic the content of breast milk, but little is known about their impact on infant gut microbiota. In this study, we tested the effect of NS and of yeast extracts (YE) with different NT content using PolyFermS continuous fermentation models mimicking formula-fed, healthy and enteropathogen-contaminated infant gut microbiota. Microbiota composition, short-chain fatty acid (SCFA) formation and gene expression were determined. NS, and to a larger extend YE modulated microbiota composition and increased metabolic activity in both models. Anaerococcus, Peptoniphilus, Fusobacterium, Lactobacillus/Pediococcus/Leuconostoc and Veillonella were enhanced when YE and/or NS were added. The production of SCFA increased with the level of supplied NT equivalents. Addition of NS and YE reduced colonization of Salmonella compared to control periods. Gene expression analysis confirmed taxonomical changes and indicated functional responses to YE. Transcripts related to NT and sulfur metabolism and iron acquisition increased while biosynthesis of co-factors and vitamins decreased after YE addition. Elevated butyrate formation correlated with increased transcripts encoding key enzymes of the two major butyrate synthesis pathways. Our results uncover a strong dose-dependent modulation of NS and YE on infant gut microbiota composition and metabolic activity. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  6. Study on Suitability of KOD DNA Polymerase for Enzymatic Production of Artificial Nucleic Acids Using Base/Sugar Modified Nucleoside Triphosphates

    Directory of Open Access Journals (Sweden)

    Satoshi Obika

    2010-11-01

    Full Text Available Recently, KOD and its related DNA polymerases have been used for preparing various modified nucleic acids, including not only base-modified nucleic acids, but also sugar-modified ones, such as bridged/locked nucleic acid (BNA/LNA which would be promising candidates for nucleic acid drugs. However, thus far, reasons for the effectiveness of KOD DNA polymerase for such purposes have not been clearly elucidated. Therefore, using mutated KOD DNA polymerases, we studied here their catalytic properties upon enzymatic incorporation of nucleotide analogues with base/sugar modifications. Experimental data indicate that their characteristic kinetic properties enabled incorporation of various modified nucleotides. Among those KOD mutants, one achieved efficient successive incorporation of bridged nucleotides with a 2′-ONHCH2CH2-4′ linkage. In this study, the characteristic kinetic properties of KOD DNA polymerase for modified nucleoside triphosphates were shown, and the effectiveness of genetic engineering in improvement of the enzyme for modified nucleotide polymerization has been demonstrated.

  7. Structure-based design, synthesis, and biological evaluation of novel pyrrolyl aryl sulfones: HIV-1 non-nucleoside reverse transcriptase inhibitors active at nanomolar concentrations.

    Science.gov (United States)

    Artico, M; Silvestri, R; Pagnozzi, E; Bruno, B; Novellino, E; Greco, G; Massa, S; Ettorre, A; Loi, A G; Scintu, F; La Colla, P

    2000-05-04

    Pyrrolyl aryl sulfones (PASs) have been recently reported as a new class of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) inhibitors acting at the non-nucleoside binding site of this enzyme (Artico, M.; et al. J. Med. Chem. 1996, 39, 522-530). Compound 3, the most potent inhibitor within the series (EC(50) = 0.14 microM, IC(50) = 0.4 microM, and SI > 1429), was then selected as a lead compound for a synthetic project based on molecular modeling studies. Using the three-dimensional structure of RT cocrystallized with the alpha-APA derivative R95845, we derived a model of the RT/3 complex by taking into account previously developed structure-activity relationships. Inspection of this model and docking calculations on virtual compounds prompted the design of novel PAS derivatives and related analogues. Our computational approach proved to be effective in making qualitative predictions, that is in discriminating active versus inactive compounds. Among the compounds synthesized and tested, 20 was the most active one, with EC(50) = 0.045 microM, IC(50) = 0.05 microM, and SI = 5333. Compared with the lead 3, these values represent a 3- and 8-fold improvement in the cell-based and enzyme assays, respectively, together with the highest selectivity achieved so far in the PAS series.

  8. Quantitative assessment of the use of modified nucleoside triphosphates in expression profiling: differential effects on signal intensities and impacts on expression ratios

    Directory of Open Access Journals (Sweden)

    Dorris David

    2002-07-01

    Full Text Available Abstract Background The power of DNA microarrays derives from their ability to monitor the expression levels of many genes in parallel. One of the limitations of such powerful analytical tools is the inability to detect certain transcripts in the target sample because of artifacts caused by background noise or poor hybridization kinetics. The use of base-modified analogs of nucleoside triphosphates has been shown to increase complementary duplex stability in other applications, and here we attempted to enhance microarray hybridization signal across a wide range of sequences and expression levels by incorporating these nucleotides into labeled cRNA targets. Results RNA samples containing 2-aminoadenosine showed increases in signal intensity for a majority of the sequences. These results were similar, and additive, to those seen with an increase in the hybridization time. In contrast, 5-methyluridine and 5-methylcytidine decreased signal intensities. Hybridization specificity, as assessed by mismatch controls, was dependent on both target sequence and extent of substitution with the modified nucleotide. Concurrent incorporation of modified and unmodified ATP in a 1:1 ratio resulted in significantly greater numbers of above-threshold ratio calls across tissues, while preserving ratio integrity and reproducibility. Conclusions Incorporation of 2-aminoadenosine triphosphate into cRNA targets is a promising method for increasing signal detection in microarrays. Furthermore, this approach can be optimized to minimize impact on yield of amplified material and to increase the number of expression changes that can be detected.

  9. Discovery of novel purine nucleoside derivatives as phosphodiesterase 2 (PDE2) inhibitors: Structure-based virtual screening, optimization and biological evaluation.

    Science.gov (United States)

    Qiu, Xiaoxia; Huang, Yiyou; Wu, Deyan; Mao, Fei; Zhu, Jin; Yan, Wenzhong; Luo, Hai-Bin; Li, Jian

    2018-01-01

    Phosphodiesterase 2 (PDE2) has received much attention for the potential treatment of the central nervous system (CNS) disorders and pulmonary hypertension. Herein, we identified that clofarabine (4), an FDA-approved drug, displayed potential PDE2 inhibitory activity (IC 50  = 3.12 ± 0.67 μM) by structure-based virtual screening and bioassay. Considering the potential therapeutic benefit of PDE2, a series of purine nucleoside derivatives based on the structure and binding mode of 4 were designed, synthesized and evaluated, which led to the discovery of the best compound 14e with a significant improvement of inhibitory potency (IC 50  = 0.32 ± 0.04 μM). Further molecular docking and molecular dynamic (MD) simulations studies revealed that 5'-benzyl group of 14e could interact with the unique hydrophobic pocket of PDE2 by forming extra van der Waals interactions with hydrophobic residues such as Leu770, Thr768, Thr805 and Leu809, which might contribute to its enhancement of PDE2 inhibition. These potential compounds reported in this article and the valuable structure-activity relationships (SARs) might bring significant instruction for further development of potent PDE2 inhibitors. Copyright © 2017. Published by Elsevier Ltd.

  10. A novel biological role for nsLTP2 from Oriza sativa: Potential incorporation with anticancer agents, nucleosides and their analogues.

    Science.gov (United States)

    Tousheh, Mojtaba; Darvishi, Fatemeh Zahra; Miroliaei, Mehran

    2015-10-01

    Development of a protein-based drug delivery system has major impact on the efficacy and bioavailability of unstable and water insoluble drugs. In the present study, the binding modes of a nonspecific lipid transfer protein (nsLTP2) from Oryza sativa with various nucleosides and analogous molecules were identified. The 3-D structure of the protein was designed and validated using modeler 9.13, Molegro virtual docker and procheck tool, respectively. The binding affinity and strength of interactions, key contributing residues and specificity toward the substrates were accomplished by computational docking and model prediction. The protein presented high affinity to acyclovir and vidarabine as purine-analogous drugs. Binding affinity is influenced by the core template and functional groups of the ligands which are structurally different cause the variation of interaction energies with nsLTP2. Nonetheless, all the evaluated analogous drugs occupy the proximity space at the nsLTP active site with high similarity in their binding modes. Our findings hold great promise for the future applications of nsLTPs in various aspects of pharmaceutical science and molecular biology. Copyright © 2015. Published by Elsevier Ltd.

  11. Simultaneous determination of nucleosides and their bases in Cordyceps sinensis and its substitutes by matrix solid-phase dispersion extraction and HPLC.

    Science.gov (United States)

    Wang, Zhibing; Li, Na; Wang, Min; Wang, Yue; Du, Lin; Ji, Xiaofeng; Yu, Aimin; Zhang, Hanqi; Qiu, Fangping

    2013-07-01

    Nine nucleosides and nucleobases, including uracil, adenine, thymine, uridine, adenosine, thymidine, cytidine, guanosine, and cordycepin in natural Cordyceps sinensis, cultured Cordyceps mycelia, and Cordyceps fruiting bodies were extracted by matrix solid-phase dispersion (MSPD) and determined by HPLC. The experimental conditions for the MSPD extraction were optimized. Florisil was used as dispersant, petroleum ether as washing solvent, and methanol as elution solvent. The Florisil-to-sample ratio was selected to be 4:1 and no additional clean-up sorbent was needed. The calibration curves had good linear relationships (r > 0.9997). The LOD and LOQ were in the range of 12~79 and 41~265 ng/mL, respectively. The intra- and interday precision were lower than 8.3%. The recoveries were between 61.5 and 93.2%. The present method consumed less sample compared with ultrasonic extraction and heating reflux extraction (HRE). The extraction yields obtained by using the present method are much higher than those obtained by UE and comparable to those obtained by HRE. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Design, Synthesis, and Evaluation of Thiophene[3,2-d]pyrimidine Derivatives as HIV-1 Non-nucleoside Reverse Transcriptase Inhibitors with Significantly Improved Drug Resistance Profiles.

    Science.gov (United States)

    Kang, Dongwei; Fang, Zengjun; Li, Zhenyu; Huang, Boshi; Zhang, Heng; Lu, Xueyi; Xu, Haoran; Zhou, Zhongxia; Ding, Xiao; Daelemans, Dirk; De Clercq, Erik; Pannecouque, Christophe; Zhan, Peng; Liu, Xinyong

    2016-09-08

    We designed and synthesized a series of human immunodeficiency virus type 1 (HIV-1) non-nucleoside reverse transcriptase inhibitors (NNRTIs) with a piperidine-substituted thiophene[3,2-d]pyrimidine scaffold, employing a strategy of structure-based molecular hybridization and substituent decorating. Most of the synthesized compounds exhibited broad-spectrum activity with low (single-digit) nanomolar EC50 values toward a panel of wild-type (WT), single-mutant, and double-mutant HIV-1 strains. Compound 27 was the most potent; compared with ETV, its antiviral efficacy was 3-fold greater against WT, 5-7-fold greater against Y181C, Y188L, E138K, and F227L+V106A, and nearly equipotent against L100I and K103N, though somewhat weaker against K103N+Y181C. Importantly, 27 has lower cytotoxicity (CC50 > 227 μM) and a huge selectivity index (SI) value (ratio of CC50/EC50) of >159101. 27 also showed favorable, drug-like pharmacokinetic and safety properties in rats in vivo. Molecular docking studies and the structure-activity relationships provide important clues for further molecular elaboration.

  13. Synthesis, structure-activity relationship and molecular docking of cyclohexenone based analogous as potent non-nucleoside reverse-transcriptase inhibitors

    Science.gov (United States)

    Nazar, Muhammad Faizan; Abdullah, Muhammad Imran; Badshah, Amir; Mahmood, Asif; Rana, Usman Ali; Khan, Salah Ud-Din

    2015-04-01

    The chalcones core in compounds is advantageously chosen effective synthons, which offer exciting perspectives in biological and pharmacological research. The present study reports the successful development of eight new cyclohexenone based anti-reverse transcriptase analogous using rational drug design synthesis principles. These new cyclohexenone derivatives (CDs) were synthesized by following a convenient route of Robinson annulation, and the molecular structure of these CDs were later confirmed by various analytical techniques such as 1H NMR, 13C NMR, FT-IR, UV-Vis spectroscopy and mass spectrometry. All the synthesized compounds were screened theoretically and experimentally against reverse transcriptase (RT) and found potentially active reverse transcriptase (RT) inhibitors. Of the compounds studied, the compound 2FC4 showed high interaction with RT at non-nucleoside binding site, contributing high free binding energy (ΔG -8.01 Kcal) and IC50 (0.207 μg/ml), respectively. Further results revealed that the compounds bearing more halogen groups, with additional hydrophobic character, offered superior anti-reverse transcriptase activity as compared to rest of compounds. It is anticipate that the present study would be very useful for the selection of potential reverse transcriptase inhibitors featuring inclusive pharmacological profiles.

  14. Development of a New Structural Class of Broadly Acting HCV Non-Nucleoside Inhibitors Leading to the Discovery of MK-8876

    Energy Technology Data Exchange (ETDEWEB)

    McComas, Casey C.; Palani, Anandan; Chang, Wei; Holloway, M. Katharine; Lesburg, Charles A.; Li, Peng; Liverton, Nigel; Meinke, Peter T.; Olsen, David B.; Peng, Xuanjia; Soll, Richard M.; Ummat, Ajay; Wu, Jie; Wu, Jin; Zorn, Nicolas; Ludmerer, Steven W. (Merck); (WuXi App Tec)

    2017-07-25

    Studies directed at developing a broadly acting non-nucleoside inhibitor of HCV NS5B led to the discovery of a novel structural class of 5-aryl benzofurans that simultaneously interact with both the palm I and palm II binding regions. An initial candidate was potent in vitro against HCV GT1a and GT1b replicons, and induced multi-log reductions in HCV viral load when orally dosed to chronic GT1 infected chimpanzees. However, in vitro potency losses against clinically relevant GT1a variants prompted a further effort to develop compounds with sustained potency across a broader array of HCV genotypes and mutants. Ultimately, a biology and medicinal chemistry collaboration led to the discovery of the development candidate MK-8876. MK-8876 demonstrated a pan-genotypic potency profile and maintained potency against clinically relevant mutants. It demonstrated moderate bioavailability in rats and dogs, but showed low plasma clearance characteristics consistent with once-daily dosing. Herein we describe the efforts which led to the discovery of MK-8876, which advanced into Phase 1 monotherapy studies for evaluation and characterization as a component of an all-oral direct-acting drug regimen for the treatment of chronic HCV infection.

  15. EEG-based functional networks evoked by acupuncture at ST 36: A data-driven thresholding study

    Science.gov (United States)

    Li, Huiyan; Wang, Jiang; Yi, Guosheng; Deng, Bin; Zhou, Hexi

    2017-10-01

    This paper investigates how acupuncture at ST 36 modulates the brain functional network. 20 channel EEG signals from 15 healthy subjects are respectively recorded before, during and after acupuncture. The correlation between two EEG channels is calculated by using Pearson’s coefficient. A data-driven approach is applied to determine the threshold, which is performed by considering the connected set, connected edge and network connectivity. Based on such thresholding approach, the functional network in each acupuncture period is built with graph theory, and the associated functional connectivity is determined. We show that acupuncturing at ST 36 increases the connectivity of the EEG-based functional network, especially for the long distance ones between two hemispheres. The properties of the functional network in five EEG sub-bands are also characterized. It is found that the delta and gamma bands are affected more obviously by acupuncture than the other sub-bands. These findings highlight the modulatory effects of acupuncture on the EEG-based functional connectivity, which is helpful for us to understand how it participates in the cortical or subcortical activities. Further, the data-driven threshold provides an alternative approach to infer the functional connectivity under other physiological conditions.

  16. Synthesis of carbasugars from aldonolactones, part III - A study on the allylic substitution of (1R,5R,8R)- and (1R,5R,8S)-8-hydroxy-2-oxabicyclo[3.3.0]oct-6-en-3-one derivatives - Preparation of (1S,2R,3R)-9-[2-hydroxy-3-(2-hydroxyethyl)cyclopent-4-en-1-yl]-9H-adenine

    DEFF Research Database (Denmark)

    Johansen, Steen Karsk; Lundt, Inge

    2001-01-01

    with these nucleophiles. Additionally, Mitsunobu substitution of (1R,5R,8R)-8-hydroxy-2-oxabicyclo[3.3.0]oct-B-en-3-one (3) with 6-chloropurine, followed by reduction of the lactone moiety and treatment with Liquid ammonia, gave the carbocyclic nucleoside (1S,2R,3R)-9-[2-hydroxy-3-(2-hydroxyethyl)cyclopent-4-en-1-yl]-9H...

  17. Ter-dependent stress response systems: novel pathways related to metal sensing, production of a nucleoside-like metabolite, and DNA-processing.

    Science.gov (United States)

    Anantharaman, Vivek; Iyer, Lakshminarayan M; Aravind, L

    2012-10-30

    The mode of action of the bacterial ter cluster and TelA genes, implicated in natural resistance to tellurite and other xenobiotic toxic compounds, pore-forming colicins and several bacteriophages, has remained enigmatic for almost two decades. Using comparative genomics, sequence-profile searches and structural analysis we present evidence that the ter gene products and their functional partners constitute previously underappreciated, chemical stress response and anti-viral defense systems of bacteria. Based on contextual information from conserved gene neighborhoods and domain architectures, we show that the ter gene products and TelA lie at the center of membrane-linked metal recognition complexes with regulatory ramifications encompassing phosphorylation-dependent signal transduction, RNA-dependent regulation, biosynthesis of nucleoside-like metabolites and DNA processing. Our analysis suggests that the multiple metal-binding and non-binding TerD paralogs and TerC are likely to constitute a membrane-associated complex, which might also include TerB and TerY, and feature several, distinct metal-binding sites. Versions of the TerB domain might also bind small molecule ligands and link the TerD paralog-TerC complex to biosynthetic modules comprising phosphoribosyltransferases (PRTases), ATP grasp amidoligases, TIM-barrel carbon-carbon lyases, and HAD phosphoesterases, which are predicted to synthesize novel nucleoside-like molecules. One of the PRTases is also likely to interact with RNA by means of its Pelota/Ribosomal protein L7AE-like domain. The von Willebrand factor A domain protein, TerY, is predicted to be part of a distinct phosphorylation switch, coupling a protein kinase and a PP2C phosphatase. We show, based on the evidence from numerous conserved gene neighborhoods and domain architectures, that both the TerB and TelA domains have been linked to diverse lipid-interaction domains, such as two novel PH-like and the Coq4 domains, in different bacteria

  18. A Versatile Strategy for Production of Membrane Proteins with Diverse Topologies: Application to Investigation of Bacterial Homologues of Human Divalent Metal Ion and Nucleoside Transporters.

    Science.gov (United States)

    Ma, Cheng; Hao, Zhenyu; Huysmans, Gerard; Lesiuk, Amelia; Bullough, Per; Wang, Yingying; Bartlam, Mark; Phillips, Simon E; Young, James D; Goldman, Adrian; Baldwin, Stephen A; Postis, Vincent L G

    2015-01-01

    Membrane proteins play key roles in many biological processes, from acquisition of nutrients to neurotransmission, and are targets for more than 50% of current therapeutic drugs. However, their investigation is hampered by difficulties in their production and purification on a scale suitable for structural studies. In particular, the nature and location of affinity tags introduced for the purification of recombinant membrane proteins can greatly influence their expression levels by affecting their membrane insertion. The extent of such effects typically depends on the transmembrane topologies of the proteins, which for proteins of unknown structure are usually uncertain. For example, attachment of oligohistidine tags to the periplasmic termini of membrane proteins often interferes with folding and drastically impairs expression in Escherichia coli. To circumvent this problem we have employed a novel strategy to enable the rapid production of constructs bearing a range of different affinity tags compatible with either cytoplasmic or periplasmic attachment. Tags include conventional oligohistidine tags compatible with cytoplasmic attachment and, for attachment to proteins with a periplasmic terminus, either tandem Strep-tag II sequences or oligohistidine tags fused to maltose binding protein and a signal sequence. Inclusion of cleavage sites for TEV or HRV-3C protease enables tag removal prior to crystallisation trials or a second step of purification. Together with the use of bioinformatic approaches to identify members of membrane protein families with topologies favourable to cytoplasmic tagging, this has enabled us to express and purify multiple bacterial membrane transporters. To illustrate this strategy, we describe here its use to purify bacterial homologues of human membrane proteins from the Nramp and ZIP families of divalent metal cation transporters and from the concentrative nucleoside transporter family. The proteins are expressed in E. coli in a

  19. Mutations in SLC29A3, encoding an equilibrative nucleoside transporter ENT3, cause a familial histiocytosis syndrome (Faisalabad histiocytosis and familial Rosai-Dorfman disease.

    Directory of Open Access Journals (Sweden)

    Neil V Morgan

    2010-02-01

    Full Text Available The histiocytoses are a heterogeneous group of disorders characterised by an excessive number of histiocytes. In most cases the pathophysiology is unclear and treatment is nonspecific. Faisalabad histiocytosis (FHC (MIM 602782 has been classed as an autosomal recessively inherited form of histiocytosis with similarities to Rosai-Dorfman disease (RDD (also known as sinus histiocytosis with massive lymphadenopathy (SHML. To elucidate the molecular basis of FHC, we performed autozygosity mapping studies in a large consanguineous family and identified a novel locus at chromosome 10q22.1. Mutation analysis of candidate genes within the target interval identified biallelic germline mutations in SLC29A3 in the FHC kindred and in two families reported to have familial RDD. Analysis of SLC29A3 expression during mouse embryogenesis revealed widespread expression by e14.5 with prominent expression in the central nervous system, eye, inner ear, and epithelial tissues including the gastrointestinal tract. SLC29A3 encodes an intracellular equilibrative nucleoside transporter (hENT3 with affinity for adenosine. Recently germline mutations in SLC29A3 were also described in two rare autosomal recessive disorders with overlapping phenotypes: (a H syndrome (MIM 612391 that is characterised by cutaneous hyperpigmentation and hypertrichosis, hepatomegaly, heart anomalies, hearing loss, and hypogonadism; and (b PHID (pigmented hypertrichosis with insulin-dependent diabetes mellitus syndrome. Our findings suggest that a variety of clinical diagnoses (H and PHID syndromes, FHC, and familial RDD can be included in a new diagnostic category of SLC29A3 spectrum disorder.

  20. Screening and characterization of purine nucleoside degrading lactic acid bacteria isolated from Chinese sauerkraut and evaluation of the serum uric acid lowering effect in hyperuricemic rats.

    Directory of Open Access Journals (Sweden)

    Ming Li

    Full Text Available Hyperuricemia is well known as the cause of gout. In recent years, it has also been recognized as a risk factor for arteriosclerosis, cerebrovascular and cardiovascular diseases, and nephropathy in diabetic patients. Foods high in purine compounds are more potent in exacerbating hyperuricemia. Therefore, the development of probiotics that efficiently degrade purine compounds is a promising potential therapy for the prevention of hyperuricemia. In this study, fifty-five lactic acid bacteria isolated from Chinese sauerkraut were evaluated for the ability to degrade inosine and guanosine, the two key intermediates in purine metabolism. After a preliminary screening based on HPLC, three candidate strains with the highest nucleoside degrading rates were selected for further characterization. The tested biological characteristics of candidate strains included acid tolerance, bile tolerance, anti-pathogenic bacteria activity, cell adhesion ability, resistance to antibiotics and the ability to produce hydrogen peroxide. Among the selected strains, DM9218 showed the best probiotic potential compared with other strains despite its poor bile resistance. Analysis of 16S rRNA sequences showed that DM9218 has the highest similarity (99% to Lactobacillus plantarum WCFS1. The acclimated strain DM9218-A showed better resistance to 0.3% bile salt, and its survival in gastrointestinal tract of rats was proven by PCR-DGGE. Furthermore, the effects of DM9218-A in a hyperuricemia rat model were evaluated. The level of serum uric acid in hyperuricemic rat can be efficiently reduced by the intragastric administration of DM9218-A (P<0.05. The preventive treatment of DM9218-A caused a greater reduction in serum uric acid concentration in hyperuricemic rats than the later treatment (P<0.05. Our results suggest that DM9218-A may be a promising candidate as an adjunctive treatment in patients with hyperuricemia during the onset period of disease. DM9218-A also has potential