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Alan G Marshall - One of the best experts on this subject based on the ideXlab platform.
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gas phase rna and dna ions 1 h d exchange of the m h anions of nucleoside 5 monophosphates gmp dgmp amp damp cmp dCMP ump dtmp ribose 5 monophosphate and 2 deoxyribose 5 monophosphate with d2o and d2s
Journal of the American Chemical Society, 1998Co-Authors: Michael A Freitas, And Christopher L Hendrickson, Alan G MarshallAbstract:H/D exchange from D2O and D2S to electrosprayed [M − H]- nucleoside 5‘-monophosphate anions (GMP, dGMP, AMP, dAMP, CMP, dCMP, UMP, TMP) is examined by Fourier transform ion cyclotron resonance mass spectrometry at 9.4 T, along with sugar phosphate controls (ribose 5-monophosphate (R5P) and 2-deoxyribose 5-monophosphate (dR5P)). The relative exchange rates of the nucleotides with D2O were dR5P > dCMP > R5P > CMP > dAMP > UMP > AMP > dTMP ≫ dGMP ≫ GMP, and with D2S were CMP > UMP ≈ dTMP > dCMP > dAMP > AMP > R5P > dR5P ≫ dGMP ≫ GMP. All exchange rates increase dramatically on changing from D2O to D2S, due to the smaller gas-phase acidity difference between exchange reagent and the nucleotide: Δ(ΔHacid) > 60 kcal mol-1 for D2O vs Δ(ΔHacid) > 20 kcal mol-1 for D2S. Ab initio calculations on model compounds at the MP2/6-31+G*//HF/6-31+G* level yield the following order of calculated acidities for each of the exchangeable hydrogens: R2O3PO-H > R2N-H > (R2O-H on ribose) > RN-H2 > (R2O-H on 2-deoxyribose). The ...
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Gas-Phase RNA and DNA Ions. 1. H/D Exchange of the [M − H]- Anions of Nucleoside 5‘-Monophosphates (GMP, dGMP, AMP, dAMP, CMP, dCMP, UMP, dTMP), Ribose 5-Monophosphate, and 2-Deoxyribose 5-Monophosphate with D2O and D2S
Journal of the American Chemical Society, 1998Co-Authors: Michael A Freitas, And Christopher L Hendrickson, Alan G MarshallAbstract:H/D exchange from D2O and D2S to electrosprayed [M − H]- nucleoside 5‘-monophosphate anions (GMP, dGMP, AMP, dAMP, CMP, dCMP, UMP, TMP) is examined by Fourier transform ion cyclotron resonance mass spectrometry at 9.4 T, along with sugar phosphate controls (ribose 5-monophosphate (R5P) and 2-deoxyribose 5-monophosphate (dR5P)). The relative exchange rates of the nucleotides with D2O were dR5P > dCMP > R5P > CMP > dAMP > UMP > AMP > dTMP ≫ dGMP ≫ GMP, and with D2S were CMP > UMP ≈ dTMP > dCMP > dAMP > AMP > R5P > dR5P ≫ dGMP ≫ GMP. All exchange rates increase dramatically on changing from D2O to D2S, due to the smaller gas-phase acidity difference between exchange reagent and the nucleotide: Δ(ΔHacid) > 60 kcal mol-1 for D2O vs Δ(ΔHacid) > 20 kcal mol-1 for D2S. Ab initio calculations on model compounds at the MP2/6-31+G*//HF/6-31+G* level yield the following order of calculated acidities for each of the exchangeable hydrogens: R2O3PO-H > R2N-H > (R2O-H on ribose) > RN-H2 > (R2O-H on 2-deoxyribose). The ...
Michael A Freitas - One of the best experts on this subject based on the ideXlab platform.
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gas phase rna and dna ions 1 h d exchange of the m h anions of nucleoside 5 monophosphates gmp dgmp amp damp cmp dCMP ump dtmp ribose 5 monophosphate and 2 deoxyribose 5 monophosphate with d2o and d2s
Journal of the American Chemical Society, 1998Co-Authors: Michael A Freitas, And Christopher L Hendrickson, Alan G MarshallAbstract:H/D exchange from D2O and D2S to electrosprayed [M − H]- nucleoside 5‘-monophosphate anions (GMP, dGMP, AMP, dAMP, CMP, dCMP, UMP, TMP) is examined by Fourier transform ion cyclotron resonance mass spectrometry at 9.4 T, along with sugar phosphate controls (ribose 5-monophosphate (R5P) and 2-deoxyribose 5-monophosphate (dR5P)). The relative exchange rates of the nucleotides with D2O were dR5P > dCMP > R5P > CMP > dAMP > UMP > AMP > dTMP ≫ dGMP ≫ GMP, and with D2S were CMP > UMP ≈ dTMP > dCMP > dAMP > AMP > R5P > dR5P ≫ dGMP ≫ GMP. All exchange rates increase dramatically on changing from D2O to D2S, due to the smaller gas-phase acidity difference between exchange reagent and the nucleotide: Δ(ΔHacid) > 60 kcal mol-1 for D2O vs Δ(ΔHacid) > 20 kcal mol-1 for D2S. Ab initio calculations on model compounds at the MP2/6-31+G*//HF/6-31+G* level yield the following order of calculated acidities for each of the exchangeable hydrogens: R2O3PO-H > R2N-H > (R2O-H on ribose) > RN-H2 > (R2O-H on 2-deoxyribose). The ...
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Gas-Phase RNA and DNA Ions. 1. H/D Exchange of the [M − H]- Anions of Nucleoside 5‘-Monophosphates (GMP, dGMP, AMP, dAMP, CMP, dCMP, UMP, dTMP), Ribose 5-Monophosphate, and 2-Deoxyribose 5-Monophosphate with D2O and D2S
Journal of the American Chemical Society, 1998Co-Authors: Michael A Freitas, And Christopher L Hendrickson, Alan G MarshallAbstract:H/D exchange from D2O and D2S to electrosprayed [M − H]- nucleoside 5‘-monophosphate anions (GMP, dGMP, AMP, dAMP, CMP, dCMP, UMP, TMP) is examined by Fourier transform ion cyclotron resonance mass spectrometry at 9.4 T, along with sugar phosphate controls (ribose 5-monophosphate (R5P) and 2-deoxyribose 5-monophosphate (dR5P)). The relative exchange rates of the nucleotides with D2O were dR5P > dCMP > R5P > CMP > dAMP > UMP > AMP > dTMP ≫ dGMP ≫ GMP, and with D2S were CMP > UMP ≈ dTMP > dCMP > dAMP > AMP > R5P > dR5P ≫ dGMP ≫ GMP. All exchange rates increase dramatically on changing from D2O to D2S, due to the smaller gas-phase acidity difference between exchange reagent and the nucleotide: Δ(ΔHacid) > 60 kcal mol-1 for D2O vs Δ(ΔHacid) > 20 kcal mol-1 for D2S. Ab initio calculations on model compounds at the MP2/6-31+G*//HF/6-31+G* level yield the following order of calculated acidities for each of the exchangeable hydrogens: R2O3PO-H > R2N-H > (R2O-H on ribose) > RN-H2 > (R2O-H on 2-deoxyribose). The ...
Wojciech Rode - One of the best experts on this subject based on the ideXlab platform.
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phosphorylation of thymidylate synthase affects slow binding inhibition by 5 fluoro dump and n4 hydroxy dCMP
Molecular BioSystems, 2016Co-Authors: Jan Ludwiczak, Borys Kierdaszuk, Piotr Wilk, Tomasz Frączyk, Tomasz Ruman, Adam Jarmula, Wojciech RodeAbstract:Endogenous thymidylate synthases, isolated from tissues or cultured cells of the same specific origin, have been reported to show differing slow-binding inhibition patterns. These were reflected by biphasic or linear dependence of the inactivation rate on time and accompanied by differing inhibition parameters. Considering its importance for chemotherapeutic drug resistance, the possible effect of thymidylate synthase inhibition by post-translational modification was tested, e.g. phosphorylation, by comparing sensitivities to inhibition by two slow-binding inhibitors, 5-fluoro-dUMP and N4-hydroxy-dCMP, of two fractions of purified recombinant mouse enzyme preparations, phosphorylated and non-phosphorylated, separated by metal oxide/hydroxide affinity chromatography on Al(OH)3 beads. The modification, found to concern histidine residues and influence kinetic properties by lowering Vmax, altered both the pattern of dependence of the inactivation rate on time from linear to biphasic, as well as slow-binding inhibition parameters, with each inhibitor studied. Being present on only one subunit of at least a great majority of phosphorylated enzyme molecules, it probably introduced dimer asymmetry, causing the altered time dependence of the inactivation rate pattern (biphasic with the phosphorylated enzyme) and resulting in asymmetric binding of each inhibitor studied. The latter is reflected by the ternary complexes, stable under denaturing conditions, formed by only the non-phosphorylated subunit of the phosphorylated enzyme with each of the two inhibitors and N5,10-methylenetetrahydrofolate. Inhibition of the phosphorylated enzyme by N4-hydroxy-dCMP was found to be strongly dependent on [Mg2+], cations demonstrated previously to also influence the activity of endogenous mouse TS isolated from tumour cells.
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crystal structures of complexes of mouse thymidylate synthase crystallized with n4 oh dCMP alone or in the presence of n5 10 methylenetetrahydrofolate
Pteridines, 2013Co-Authors: A Dowiercial, Borys Kierdaszuk, Piotr Wilk, Adam Jarmula, W Rypniewski, Wojciech RodeAbstract:To solve the inhibition mechanism of thymidylate synthase (TS) by N 4 -hydroxy-dCMP (N 4 -OH-dCMP), crystallo- graphic studies were undertaken. Structures of three mouse TS (mTS) complexes with the inhibitor were solved, based on crystals formed by the enzyme protein in the presence of either only N 4 -OH-dCMP (crystal A, belonging to the space group C 1 2 1, with two monomers in asymmetric unit (ASU), measured to 1.75 A resolution) or both N 4 -OH-dCMP and N 5,10 - methylenetetrahydrofolate (mTHF) (crystals B and C, both belonging to the space group C 2 2 21, each with a single monomer in ASU, measured to resolution of 1.35 A and 1.17 A , respectively). Whereas crystal A-based structure revealed the mTS-N 4 -OH-dCMP binary complex, as expected, crystals B- and C-based structures showed the enzyme to be involved in a ternary complex with N 4 -OH-dCMP and noncovalently bound dihydrofolate (DHF), instead of expected mTHF, sug- gesting the inhibition to be a consequence of an abortive enzyme-catalyzed reaction, involving a transfer of the one- carbon group to a hitherto unknown site and oxidation of THF to DHF. Moreover, both C(5) and C(6) inhibitor atoms showed sp 3 hybridization, suggesting C(5) reduction, with no apparent indication of C(5) proton release. In accord- ance with our previous results, in all subunits of these structures the inhibitor molecule was identified as the anti rotamer of imino tautomer, forming, similar to deoxyuridine monophosphate, two hydrogen bonds with a conservative asparagine (mouse Asn220) side chain.
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mechanism of thymidylate synthase inhibition by n4 hydroxy n4 hydroxy 5 fluoro dCMP in view of the structure and conformation of n4 hydroxy n4 hydroxy 5 fluoro cytosine calculated by the ab initio quantum mechanical methods
Advances in Experimental Medicine and Biology, 1993Co-Authors: Ludwik Adamowicz, Andrzej Leś, Wojciech RodeAbstract:Thymidylate synthase (EC 2.1.1.45) slow-binding inhibition by N4-hydroxy-dCMP (oh4dCMP) was shown to depend on conformation of the exocyclic N4-OH group, with the anti rotamer, relative to the ring N(3), indicated as an active species. 1 Potentiation of inhibition by the 5-fluoro substituent, observed for N4-hydroxy-5-fluoro-dCMP (oh4f5CMP), was explained in terms of hydrogen bonding between the N4-OH and C(5)-F groups, influencing an assumed syn-anti equilibrium by stabilization of the anti rotamer.1 In order to test the latter hypothesis two cytosine analogues, N4-hydroxy-cytosine (oh4C) and N4-hydroxy-5-fluorocytosine (oh4f5C), were theoretically studied, and their molecular structures determined, by ab initio quantum mechanical methods.
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mechanism of inhibition of mammalian tumor and other thymidylate synthases by n4 hydroxy dCMP n4 hydroxy 5 fluoro dCMP and related analogues
Biochemistry, 1990Co-Authors: Wojciech Rode, Zbigniew Zielinski, Jolanta M Dzik, Tadeusz Kulikowski, Maria Bretner, Borys Kierdaszuk, Joanna Ciesla, David ShugarAbstract:N{sup 4}-Hydroxy-dCMP (N{sup 4}-OH-dCMP), N{sup 4}-methoxy-dCMP (N{sup 4}-OMe-dCMP), and their 5-fluoro congeners were all slow-binding inhibitors of Ehrlich carcinoma thymidylate synthase (TS), competitive with respect to dUMP, and had differing kinetic constants describing interactions with the two TS binding sites. N{sup 4}-OH-dCMP was not a substrate and its inactivation of TS was methylenetetrahydrofolate-dependent, hence mechanism-based. K{sub i} values for N{sup 4}-OH-dCMP and its 5-fluoro analogue were in the range 10{sup {minus}7}-10{sup {minus}8} M, 2-3 orders of magnitude higher for the corresponding N{sup 4}-OMe analogues. The 5-methyl analogue of N{sup 4}-OHdCMP was 10{sup 4}-fold less potent, pointing to the anti rotamer of the imino form of exocyclic N{sup 4}-OH, relative to the ring N(3), as the active species. This is consistent with weaker slow-binding inhibition of the altered enzyme from 5-FdUrd-resistant, relative to parent, L1210 cells by both FdUMP and N{sup 4}-OH-dCMP, suggesting interaction of both N{sup 4}-OH and C(5)-F groups with the same region of the active center. Kinetic studies with purified enzyme from five sources, viz., Ehrlich carcinoma, L1210 parental, and 5-FdUrd-resistant cells, regenerating rat liver, and the tapeworm Hymenolepis diminuta, demonstrated that addition of a 5-fluoro substituent to N{sup 4}-OH-dCMP increased its affinity from 2- to 20-fold formore » the enzyme from different sources. With the Ehrlich and tapeworm enzymes, N{sup 4}-OH-FdCMP and FdUMP were almost equally effective inhibitors.« less
Weerachai Nasomphan - One of the best experts on this subject based on the ideXlab platform.
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selective fluorescence sensing of deoxycytidine 5 monophosphate dCMP employing a bis diphenylphosphate diimine ligand
Journal of Fluorescence, 2011Co-Authors: Weerachai Nasomphan, Pramuan Tangboriboonrat, Srung SmanmooAbstract:A new bis(diphenylphosphate)diimine ligand (BP1) was prepared and evaluated for its ability for selective detection of deoxycytidine 5′-monophosphate (dCMP). BP1 exhibited off-type fluorescence in the presence of dCMP. The fluorescence of BP1 was significantly quenched upon the addition of 2.5 × 10−4 M dCMP and the detection limit was 1.25 × 10−5 M in MeCN-H2O (1:1, v/v). The binding ratio between BP1 and dCMP was determined to be 1:1 with the binding constant of 3.98 ± 0.60 × 10−3 M−1.
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Selective Fluorescence Sensing of Deoxycytidine 5′-Monophosphate (dCMP) Employing a Bis(diphenylphosphate)diimine Ligand
Journal of Fluorescence, 2010Co-Authors: Weerachai Nasomphan, Pramuan Tangboriboonrat, Srung SmanmooAbstract:A new bis(diphenylphosphate)diimine ligand (BP1) was prepared and evaluated for its ability for selective detection of deoxycytidine 5′-monophosphate (dCMP). BP1 exhibited off-type fluorescence in the presence of dCMP. The fluorescence of BP1 was significantly quenched upon the addition of 2.5 × 10−4 M dCMP and the detection limit was 1.25 × 10−5 M in MeCN-H2O (1:1, v/v). The binding ratio between BP1 and dCMP was determined to be 1:1 with the binding constant of 3.98 ± 0.60 × 10−3 M−1.
Bernhard Lippert - One of the best experts on this subject based on the ideXlab platform.
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cis diammineplatinum ii forms a macrochelate with 2 deoxycytidine 5 monophosphate dCMP 2 reactivity and acid base properties of cis pt nh 3 2 dCMP
Journal of Biological Inorganic Chemistry, 1998Co-Authors: Gerda Oswald, Ingo Rombeck, Bin Song, Helmut Sigel, Bernhard LippertAbstract:The synthesis of cis-Pt(NH3)2(dCMP) is reported and by various physico-chemical methods it is demonstrated that it is a macrochelate in which Pt(II) is bound simultaneously to the N3 site of cytosine in dCMP2– and to a phosphate-oxygen atom. According to the NOESY spectra (cross-peaks between cytosine H6 and H2′ and H3′) the cytosine ring adopts an anti orientation. Highly unusual is the significant (1 ppm) downfield shift of the sugar proton H5″ in the 1H-NMR spectrum and the sensitivity of the cytosine H6 resonance on the protonation state of the phosphate group. Based on these three features a geometry for the macrochelate is proposed. The compound is a major product of the reaction of cis-[Pt(NH3)2(H2O)2]2+ with dCMP2– at neutral pH, but it even forms at pH 5. By applying pD-dependent NMR spectroscopy (1H, 31P) and potentiometric pH titration, it is demonstrated that the Pt-coordinated phosphate group can be protonated (pKa/1=3.21±0.10 and 3.31±0.05, respectively), and 1H- and 31P-NMR spectra also indicate deprotonation (pKa/2=13.35±0.25) of the exocyclic amino group of the cytosine moiety. The metal ion binding affinity of cis-Pt(NH3)2(dCMP) is very small, as shown for Cu2+ (log K<0.6). The cis-Pt(NH3)2(dCMP) complex reacts with nucleosides and nucleotides (L′) by losing its chelate structure and forming mixed ligand complexes, cis-Pt(NH3)2(dCMP)(L′); this means that the phosphate group is released from the coordination sphere of Pt(II), indicating that the Pt(II)-O(phosphate) bond is not very strong.
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Cis -diammineplatinum(II) forms a macrochelate with 2′-deoxycytidine 5′-monophosphate (dCMP 2– )! Reactivity and acid-base properties of cis -Pt(NH 3 ) 2 (dCMP)
Journal of Biological Inorganic Chemistry, 1998Co-Authors: Gerda Oswald, Ingo Rombeck, Bin Song, Helmut Sigel, Bernhard LippertAbstract:The synthesis of cis-Pt(NH3)2(dCMP) is reported and by various physico-chemical methods it is demonstrated that it is a macrochelate in which Pt(II) is bound simultaneously to the N3 site of cytosine in dCMP2– and to a phosphate-oxygen atom. According to the NOESY spectra (cross-peaks between cytosine H6 and H2′ and H3′) the cytosine ring adopts an anti orientation. Highly unusual is the significant (1 ppm) downfield shift of the sugar proton H5″ in the 1H-NMR spectrum and the sensitivity of the cytosine H6 resonance on the protonation state of the phosphate group. Based on these three features a geometry for the macrochelate is proposed. The compound is a major product of the reaction of cis-[Pt(NH3)2(H2O)2]2+ with dCMP2– at neutral pH, but it even forms at pH 5. By applying pD-dependent NMR spectroscopy (1H, 31P) and potentiometric pH titration, it is demonstrated that the Pt-coordinated phosphate group can be protonated (pKa/1=3.21±0.10 and 3.31±0.05, respectively), and 1H- and 31P-NMR spectra also indicate deprotonation (pKa/2=13.35±0.25) of the exocyclic amino group of the cytosine moiety. The metal ion binding affinity of cis-Pt(NH3)2(dCMP) is very small, as shown for Cu2+ (log K