Deoxyguanylic Acid Research Articles

B- to Z-DNA transition probed by oligonucleotides containing methylphosphonates.

The simulation of the B--Z-DNA transition by using space-filling models of the dimer d(C-G) shows the possibility of hydrogen-bond formation between the N-2 amino group of the partially rotated guanine and one of the 5'-phosphate oxygens of deoxyguanylic acid. To probe the importance of this postulated interaction, analogs of the hexamer d(C-G)3 were synthesized. These analogs contained a methylphosphonate linkage, of distinct stereochemistry, which replaced the first 5'-phosphate linkage of deoxyguanosine. The CD spectra in high salt concentration showed that the hexamer containing a methylphosphonate linkage with the RP stereochemistry formed Z-DNA to the same extent as d(C-G)3, whereas the hexamer containing a methylphosphonate linkage with the SP stereochemistry did not form Z-DNA. These results are consistent with a mechanism in which an interaction between the N-2 amino group of guanine and the prochiral SP oxygen of deoxyguanosine 5'-phosphate kinetically controls the formation of Z-DNA. A water bridge between the N-2 amino group of guanine and the 3'-phosphate oxygen of deoxyguanylic acid has been implicated in the stabilization of Z-DNA. To probe the importance of this water bridge, two additional analogs of the hexamer d(C-G)3 were synthesized. These analogs contained a methylphosphonate linkage, of distinct stereochemistry, that replaced the first deoxyguanosine 3'-phosphate. The CD spectra showed that the hexamer containing a methylphosphonate linkage of the RP stereochemistry underwent the transition to Z-DNA to the same extent as d(C-G)3, whereas the hexamer containing a methylphosphonate linkage of the SP stereochemistry underwent the transition to Z-DNA to a 35% lesser extent. Thus the water bridge involving the prochiral SP oxygen provides modest stabilization energy for Z-DNA. These studies, therefore, suggest that the B--Z-DNA transition is regulated both thermodynamically and kinetically through hydrogen-bond interactions involving phosphate oxygens and the N-2 amino group of guanine.

Antibodies to guanylic acid: fractionation and specificities.

Guanylic-acid-specific antibodies were elicited in rabbits, using as immunogen pG linked through 5'-phosphate to thyroglobulin. Specificity and affinity of antibodies to nucleotides, nucleosides, DNA, and RNA were studied by their binding to radioactive ligands and competition experiments. Guanylic-acid-specific antibodies do not bind to deoxyguanylic acid and have an average association constant of 10(7) M-1 at 4 degrees C. Binding of the antibodies to 3H-RNA is G-specific. The antibodies do not bind to 32P-ssDNA or 32P-dsDNA. The pG-specific antibodies could be separated into different fractions by affinity chromatography. These fractions, though specific to pG, differ in their cross-reactivities to nucleosides and nucleotides.

Oxygen uptake in the radiolysis of aqueous solutions of nucleic acids and their constituents.

The 60Co-gamma-irradiation of organic compounds in oxygenated aqueous solution leads to consumption of oxygen. G (O2 uptake) have been measured for 27 compounds in N2O/O2 (4:1)- saturated solutions using an oxygen sensitive electrode. G (O2 uptake) (in brackets, measured at 20 degrees C and a dose rate of 0.4 Gy s-1) for isopropanol (3.0), sodium formate (3.0), D-glucose (3.2), 2-deoxy-D-ribose (3.0), t-butanol (4.2) and diethyl ether (4.5) were found to be in agreement with th expectation based on product yields and/or known mechanisms. High O2 uptake was observed with polyethyleneoxide (10.2), which increases with decreasing dose rate and/or increasing temperature (G(O2 uptake) = 40 at 0.04 Gy s-1 and 50 degrees C). These results are explained by assuming a chain reaction. The nucleotides 5'-thymidylic acid (4.4), 5'-deoxy-cytidylic acid (4.8), 5'-deoxyadenylic acid (1.9) and 5'-deoxyguanylic acid (1.6) show that the pyrimidine derivatives consume considerably more oxygen than the purine derivatives. Analogous results are obtained with the nucleobases and nucleosides. The pyrimidine-purine difference is even more pronounced in the corresponding polymer, poly U (21) and poly A (3.5). The large value of poly U shows that a significant contribution of a chain reaction is present. G(O2 uptake) for DNA are dose-rate, temperature and concentration-dependent. The O2 uptake for single-stranded DNA (6.8) and double-stranded DNA (4.6) is higher than for an equivalent mixture of nucleotides (3.2). These results indicate that in DNA also a short chain reaction takes place.

Cloning of rat alpha-fetoprotein 3'-terminal complementary deoxyribonucleic acid sequences and preparation of radioactively labeled hybridization probes from cloned deoxyribonucleic acid inserts.

Double-stranded complementary deoxyribonucleic acid (cDNA) was synthesized from rat yolk sac alpha-fetoprotein (AFP) mRNA, inserted into the PstI site of plasmid pBR322 by an oligo(deoxyguanylic acid).oligo(deoxycytidylic acid) joining technique, and cloned in Escherichia coli chi 1776. A plasmid containing an inserted AFP double-stranded cDNA with a contiguous poly(adenylic acid) [poly(A)] segment was identified and subsequently employed in a new method for preparing AFP-specific hybridization probe. Following an initial digestion of the AFP plasmid with HindIII to create an open, recessed 3' end, lambda exonuclease III was employed to remove the DNA strand opposite the coding strand of the cDNA insert. Oligo(thymidylic acid) was then annealed to the poly(A) segment and employed as primer for E. coli DNA polymerase I to synthesize a 32P-labeled cDNA copy of the AFP coding strand. The single-stranded cDNA product was easily isolated by sedimentation through isokinetic alkaline sucrose gradients. Hybridization with this AFP-specific cDNA probe showed that the yolk sac contained a 6-fold greater concentration of AFP mRNA than that of the fetal liver. AFP mRNA was also found in the normal adult liver, but at a much lower level than in the fetal liver. The concentrations of AFP mRNA in Morris hepatomas 7777 and 8994, however, were significantly elevated to a 2- to 3-fold higher concentration that in the fetal liver.

The in vivo formation and repair of DNA adducts from 1'-hydroxysafrole.

1'-Hydroxysafrole is a proximate carcinogenic metabolite of the naturally occurring hepatocarcinogen safrole. Comparison by high-performance liquid chromatography of the nucleoside adducts obtained from hepatic DNA of adult female mice treated with [2',3'-3H]1'-hydroxysafrole with those formed by reaction of deoxyribonucleosides with electrophilic derivatives of 1'-hydroxysafrole indicated that the four in vivo adducts studied were derived from an ester of 1'-hydroxysafrole. Three of the four adducts comigrated with products of the reaction of 1'-acetoxysafrole with deoxyguanosine, whereas the fourth adduct comigrated with the major reaction product of the ester with deoxyadenosine. Analysis of the three deoxyguanosine adducts indicated that all three involve substitution on the 2-amino group of guanine. A sample of ther major adduct prepared from deoxyguanylic acid has been characterized from its NMR spectrum as N2-(trans-isosafrol-3'-yl)-deoxyguanosine, and the deoxyadenosine adduct has been similarly characterized as N6-(trans-isosafrol-3'-yl)-deoxyadenosine. Repair replication was measured in cultured human T98G cells exposed to 1'-acetoxysafrole using the combined 5-bromodeoxyuridine density label and radioisotopic label method. At a concentration of 1 mM 1'-acetoxysafrole, the amount of repair synthesis approached maximum values only about 15% of those obtained after saturating doses of ultraviolet light. Repair patch size distribution was found to be similar in cells treated with ultraviolet light or 1'-acetoxysafrole as determined by the density of repair-labeled DNA relative to that of parental DNA.

79] Preparation of plasmids and tailing with oligodeoxyguanylic acid

Publisher Summary This chapter presents the preparation of plasmids and tailing with oligodeoxyguanylic acid. The plasmid DNA is prepared essentially as described. Covalently closed circular plasmid DNA is further purified by CsC1 density gradient centrifugation. The number of deoxyguanylic acid residues added to each 3' end of the linear DNA is determined by subtracting the number of residues added to the circular plasmid DNA from the number added to the linear plasmid DNA. The plasmid DNA containing homopolymer tails of oligo(dG) is purified by electrophoresis on a 0.7% agarose gel in order to remove small amounts of undigested circular plasmid DNA. The linear plasmid DNA is recovered from the gel by the KI-hydroxyapatite column chromatography method as described. The eluted DNA is extracted with n-butanol to remove ethidium bromide, dialyzed against a solution of 10 mM Tris • HCI, pH 8, and 1 mM Na 2 EDTA, and then precipitated with ethanol. Recovery with this purification step is 50-80%.

Single-stranded poly(deoxyguanylic acid) associates into double- and triple-stranded structures.

Circular plasmid deoxyribonucleic acid (DNA), pBR322, was digested with the restriction endonuclease PstI to give full-length double-stranded DNA molecules, terminated by two self-complementary single-stranded sequences: (formula: see text). The protruding 3' termini were extended with dG by using calf thymus terminal deoxynucleotidyl transferase and dGTP, to form single-stranded tails of oligo(dG). At a length of about dG15, such tails become resistant to single strand specific endonuclease S1, and also cease to function as substrate (initiator) for the terminal deoxynucleotidyl transferase. This altered reactivity arises from association of the oligo(dG) tails into double- and triple-stranded structures, resulting in linear, circular, and branched polymers of the monomeric linear plasmid DNA. All these polymeric structures of the plasmid DNA are stable at room temperature, can be observed in the electron microscope, and can be separated from each other by agarose gel electrophoresis. At 60 degrees C or in 50% formamide, most of the oligo(dG) self-association can be reversed (melted), and the plasmid DNA is again found as the original linear monomer.

Electrochemical behaviour of guanine and its derivatives. I. Fast cyclic voltammetry of guanosine and calf thymus DNA

Electrochemical behaviour of guanosine and calf thymus DNA was investigated at D.M.E. by fast cyclic voltammetry (5–100 V/s) with triangular wave in phosphate buffer containing ammonium formate. Concentrations of guanosine were in the range from 5×10−5 M to 1×10−3 M. Under these experimental conditions guanosine gives a well developed anodic peak in the vicinity of −0.2 V vs. S.C.E. It is necessary to polarize the electrode to potentials more negative than −1.8 V for the appearance of this peak. The influence of the guanosine concentration, scan rate, switching potential, pH of solution and nature of the background electrolyte on the properties of the anodic peak were studied. At concentrations lower than 2×10−4 M only one anodic peak appeared on the cyclic voltammogram which corresponded to a diffusion-controlled process. At higher concentrations of guanosine further anodic peaks appeared. The anodic peak (−0.2 V) was also observed on the cyclic voltammograms of guanine, deoxyguanosine, guanylic acid, deoxyguanylic acid and dinucleotide CpG. Besides the low-molecular weight substances the behaviour of polynucleotides was investigated: single-stranded (thermally denaturated) and double-helical (native) DNA. Denatured DNA yielded an anodic peak of properties similar to those of peaks of guanine derivatives. On the contrary, native DNA did not yield this peak. The results obtained with DNA agree with previous measurements carried out by means of oscillographic polarography at controlled a.c., and indicate that cyclic voltammetry may become an important tool in the study of the behaviour of guanine residues in a polynucleotide chain.

Preparation and specificity of endonuclease IV induced by bacteriophage T4.

Bacteriophage-T4-induced endonuclease IV suitable for DNA sequence analysis has been prepared by a modified and easily reproducible method. The specificity of T4-induced endonuclease IV has been investigated in order to verify whether this enzyme exhibits a single nucleotide recognition or a short sequence recognition. The 5'-terminal dinucleotides and 3'-terminal nucleotides of oligonucleotides released by T4-induced endonuclease IV from three single-stranded DNAs (from bacteriophages phiX174, fd, M 13) have been analysed. In different DNAs, 74-82% of the 5'-terminal dinucleotides end in 5'-deoxycytidylic acid; small but significant levels of several dinucleotides ending in 5'-deoxyadenylic acid, 5'-thymidylic acid and 5'-deoxyguanylic acid are also found. As far as 3'-terminal nucleotides are concerned all nucleotides are present with a large predominance of thymidylic acid. It is concluded that T4-induced endonuclease IV recognizes short nucleotide sequences like all other DNases investigated so far. The spectrum of such sequences is, however, very narrow.

Alkylation by propylene oxide: Lawley, P. D. & Jarman, M. (1972). Alkylation by propylene oxide of deoxyribonucleic acid, adenine, guanosine and deoxyguanylic acid. Biochem. J.126,893

Alkylation by propylene oxide: Lawley, P. D. & Jarman, M. (1972). Alkylation by propylene oxide of deoxyribonucleic acid, adenine, guanosine and deoxyguanylic acid. Biochem. J.126,893

Primer-template specificity of DNA polymerases bound to mitochondria and nuclei

Recently DNA polymerases of eukaryotes have been the subject of intensive investigation as a consequence of the importance of research on RNAdependent DNA polymerases in normal [ 1, 21 and in tumor cells and tissues [3-61. Attention has also been focused on the activity of DNA polymerases in regenerating tissues [7] and in developing systems [8-l 11. Margolis [ 121 has studied the distribution patterns of DNA-dependent DNA polymerase in different regions of brain of chick embryo, and one of us has shown [ 131 that mitochondria and nuclei of chick embryonic brain contain bound enzymes which catalyze the polymerization of dGMP when poly dG:poly dC or poly dG:poly rC are provided as primer templates. In the present paper we report the characterization of partially purified poly dT:poly rA-dependent DNA polymerase (bound to nuclei) and oligo dT:poly rAdependent DNA polymerase (bound to mitochondria), both of them polymerizing thymidylic acid. Moreover, we have studied poly dG:poly dC and poly dG:poly rC-dependent activities polymerizing deoxyguanylic acid, in both mitochondria and nuclei.

2617. Alkylation by propylene oxide: Lawley, P. D. & Jarman, M. (1972). Alkylation by propylene oxide of deoxyribonucleic acid, adenine, guanosine and deoxyguanylic acid. Biochem. J.126, 893

2617. Alkylation by propylene oxide: Lawley, P. D. & Jarman, M. (1972). Alkylation by propylene oxide of deoxyribonucleic acid, adenine, guanosine and deoxyguanylic acid. Biochem. J.126, 893

Alkylation by propylene oxide of deoxyribonucleic acid, adenine, guanosine and deoxyguanylic acid.

1. Propylene oxide reacts with DNA in aqueous buffer solution at about neutral pH to yield two principal products, identified as 7-(2-hydroxypropyl)guanine and 3-(2-hydroxypropyl)adenine, which hydrolyse out of the alkylated DNA at neutral pH values at 37 degrees C. 2. These products were obtained in quantity by reactions between propylene oxide and guanosine or adenine respectively. 3. The reactions between propylene oxide and adenine in acetic acid were parallel to those between dimethyl sulphate and adenine in neutral aqueous solution; the alkylated positions in adenine in order of decreasing reactivity were N-3, N-1 and N-9. A method for separating these alkyladenines is described. 4. Deoxyguanylic acid sodium salt was alkylated at N-7 by propylene oxide in neutral aqueous solution. 5. The nature of the side chain in the principal alkylation products was established by mass spectrometry, and the nature of the products is consistent with their formation by the bimolecular reaction mechanism.

Nuclear magnetic resonance spectral studies on actinomycin D. Preliminary observations on the effect of complex formation with 5'-deoxyguanylic acid.

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTNuclear magnetic resonance spectral studies on actinomycin D. Preliminary observations on the effect of complex formation with 5'-deoxyguanylic acidByron H. Arison and Karst HoogsteenCite this: Biochemistry 1970, 9, 20, 3976–3983Publication Date (Print):September 29, 1970Publication History Published online1 May 2002Published inissue 29 September 1970https://doi.org/10.1021/bi00822a016RIGHTS & PERMISSIONSArticle Views61Altmetric-Citations56LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (752 KB) Get e-Alerts Get e-Alerts

Reactions between Singlet Oxygen and the Constituents of Nucleic Acids: Importance of Reactions in Photodynamic Processes

Bases, nucleosides, nucleotides, and polynucleotides were exposed to chemically generated singlet oxygen to determine whether the species oxidized paralleled those oxidized in photodynamic reactions. In neutral or basic aqueous solution guanine, guanosine, deoxyguanosine, guanylic acid, deoxyguanylic acid, thymine, and uracil reacted with singlet oxygen. Since these compounds are oxidized in photodynamic processes, this study provides further evidence that singlet oxygen is the active intermediate in the photodynamic oxidation of nucleic acid constituents. Dienophilic attack by singlet oxygen is considered to be a plausible mechanism in these reactions.

The Photosensitized Reaction of Deoxyguanosine in the Presence of Methylene Blue

Abstract The kinetics of the photosensitized decomposition of deoxyguanosine with methylene blue was studied by means of spectrophotometric measurements. The reaction rate increased with an increase in the concentration of the reactant or of oxygen up to a saturation level for each case at a given concentration of the dye. In a buffer solution of a lower phosphate concentration, a larger rate was observed. On the basis of these dependencies of the rate, the reaction scheme was discussed, and the ratios of some rate constants were estimated based on Schenck’s mechanism. A preliminary NMR measurement of the protons of deoxyguanylic acid in D2O suggested that the basepart of the nucleotide was specifically attacked by this photosensitized reaction.

Studies on the binding of actinomycin D to DNA and DNA model polymers

The ability of 17 different DNA's to bind actinomycin D was studied by a variety of techniques including equilibrium dialysis, in vitro transcription and analytical buoyant density centrifugation. The major conclusions are as follows: (1) The presence of deoxyguanylic acid in a DNA is not necessary for complex formation. Poly dI binds approximately one-quarter as much AM as DNA's which contain 50% G + C; the equilibrium constant for the poly dI-AM complex is as large as that observed for 50% G + C DNA's. (2) The presence of deoxyguanylic acid in a DNA is not sufficient for complex formation. Poly d(A-T-C) · poly d(G-A-T), which contains 33% G + C, binds little or no AM as judged by five different techniques. (3) A marked nucleotide sequence preference exists for the binding reaction. When comparing sequence isomeric DNA's, the poly d(pur n -pyr m ) · poly d(pur m -pyr n ) isomer binds more AM and binds AM more tightly than does the poly d pur · poly d pyr isomer. (4) The guanine-containing DNA's tested, with the exception of poly d(A-T-C) · poly d(G-A-T), bind AM and are: Micrococcus luteus DNA, salmon sperm DNA, poly d(G-C) · poly d(G-C), poly dG · poly dC, poly d(T-G) · poly d(C-A), poly d(T-C) · poly d(G-A), poly d(T-T-G) · poly d(C-A-A), poly d(T-T-C) · poly d(G-A-A) and poly d(T-A-C) · poly d(G-T-A). (5) The DNA's which are devoid of deoxyguanine, with the exception of poly dI, do not bind AM and are: poly d(A-T) · poly d(A-T), poly dA · poly dT, poly dI · poly dC and poly d(I-C) · poly d(I-C). The results are discussed in relation to two models for the AM-DNA complex, the hydrogen-bonded, “outside-binding” model of Hamilton, Fuller & Reich (1963) and the intercalation model of Müller & Crothers (1968). The data are not consistent with the hydrogen-bonded model.

In Vitro Antiviral Activity of Mycophenolic Acid and Its Reversal by Guanine-Type Compounds

With the agar diffusion test and BS-C-1 cells, mycophenolic acid was found to give a straight-line dose-response activity in inhibiting the cytopathic effects of vaccinia, herpes simplex, and measles viruses. Plaque tests have shown 100% reduction of virus plaques by mycophenolic acid over drug ranges of 10 to 50 μg/ml and virus input as high as 6,000 plaque-forming units (PFU) per flask. Back titration studies with measles virus inhibited by mycophenolic acid have indicated that extracellular virus titers were reduced by approximately 3 logs 10 and total virus was reduced by 1 log 10 . The agar diffusion test system lends itself readily to drug reversal studies. Mycophenolic acid incorporated into agar at 10 μg/ml gave 100% protection to virus-infected cells. Filter paper discs impregnated with selected chemical agents at concentrations of 1,000 μg/ml (20 μg per filter paper disc) were placed on the agar surface. Reversal of the antiviral activity of mycophenolic acid was indicated by virus breakthrough in those cells in close proximity to the filter paper disc. Chemicals showing the best reversal of the antiviral activity of mycophenolic acid were guanine, guanosine, guanylic acid, deoxyguanylic acid, and 2,6-diaminopurine. The reversal of antiviral activity was confirmed by titrations of virus produced with various amounts of both mycophenolic acid and guanine present and by isotope tracer methods with uptakes of labeled uridine, guanine, leucine, and thymidine in treated and nontreated, infected and noninfected cells as parameters. All antiviral effects of mycophenolic acid at 10 μg/ml could be reversed to the range shown by untreated controls by the addition of 10 μg/ml of those chemicals exhibiting reversal activity.

In Vitro Antiviral Activity of Mycophenolic Acid and Its Reversal by Guanine-Type Compounds

With the agar diffusion test and BS-C-1 cells, mycophenolic acid was found to give a straight-line dose-response activity in inhibiting the cytopathic effects of vaccinia, herpes simplex, and measles viruses. Plaque tests have shown 100% reduction of virus plaques by mycophenolic acid over drug ranges of 10 to 50 mug/ml and virus input as high as 6,000 plaque-forming units (PFU) per flask. Back titration studies with measles virus inhibited by mycophenolic acid have indicated that extracellular virus titers were reduced by approximately 3 logs(10) and total virus was reduced by 1 log(10). The agar diffusion test system lends itself readily to drug reversal studies. Mycophenolic acid incorporated into agar at 10 mug/ml gave 100% protection to virus-infected cells. Filter paper discs impregnated with selected chemical agents at concentrations of 1,000 mug/ml (20 mug per filter paper disc) were placed on the agar surface. Reversal of the antiviral activity of mycophenolic acid was indicated by virus breakthrough in those cells in close proximity to the filter paper disc. Chemicals showing the best reversal of the antiviral activity of mycophenolic acid were guanine, guanosine, guanylic acid, deoxyguanylic acid, and 2,6-diaminopurine. The reversal of antiviral activity was confirmed by titrations of virus produced with various amounts of both mycophenolic acid and guanine present and by isotope tracer methods with uptakes of labeled uridine, guanine, leucine, and thymidine in treated and nontreated, infected and noninfected cells as parameters. All antiviral effects of mycophenolic acid at 10 mug/ml could be reversed to the range shown by untreated controls by the addition of 10 mug/ml of those chemicals exhibiting reversal activity.

Inhibition of deoxycytidylic and deoxyguanylic acid kinase activity by adenosine triphosphate and deoxyadenosine triphosphate in pneumococci

Inhibition of deoxycytidylic and deoxyguanylic acid kinase activity by adenosine triphosphate and deoxyadenosine triphosphate in pneumococci