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K Frenkel

Publications and source records attributed to K Frenkel.

70 records · Page 4Linked to original sources
PubMed1986

Copper ions and hydrogen peroxide form hypochlorite from NaCl thereby mimicking myeloperoxidase.

Sea urchins have elaborated multiple defenses to assure monospermic fertilization. In this work, we have concentrated on a study of the mechanism(s) by which hydrogen peroxide (H2O2) prevents polyspermy in Arbacia punctulata. We found that it is not H2O2 but probably hypochlorous acid/hypochlorite (HOCl/OCl-) derived from H2O2 that is toxic to the supernumerary sperm. The spermicidal activity of H2O2 is potentiated by at least one order of magnitude by cupric ions (Cu2+). This increased toxicity is not due to the formation of hydroxyl radicals (.OH) because .OH scavengers did not counteract the activity of Cu2+. Moreover, substitution of Cu2+ by ferrous ions (Fe2+), which are known to cause formation of .OH from H2O2, had no effect on fertilization even at 10(2)-10(3) times higher concentrations. In contrast, 3-amino-1,2,4-triazole (AT), and HOCl/OCl- scavenger, totally reversed the toxic effects of Cu2+. Furthermore, we found that HOCl/OCl- is generated in solutions of H2O2 and Cu2+ in the presence of 0.5 M NaCl and that its accumulation is abolished by AT. Thus it is possible that the antifertility properties of copper are due to its ability to mediate formation of HOCl/OCl-. HOCl/OCl- generated by Cu2+ from H2O2 and Cl-, a low concentration of exogenously added HOCl/OCl-, or increased concentrations of H2O2 has similar inhibitory effects on the fertilization process in sea urchins. Therefore, we suggest that polyspermy is prevented by the action of a myeloperoxidase that affects the formation of HOCl/OCl- from the Cl- present in sea water through reaction with H2O2 generated by the newly fertilized egg.

Amitrole↗
PubMed1986

Radiation-like modification of bases in DNA exposed to tumor promoter-activated polymorphonuclear leukocytes.

Oxygen species generated by human polymorphonuclear leukocytes (PMNs) activated by 12-O-tetradecanoylphorbol-13-acetate (TPA) caused the formation of 5-hydroxymethyl-2'-deoxyuridine (HMdUrd), and (+) and (-) diastereoisomers of cis-thymidine glycol (dTG) in DNA that was exposed to them. There were 9 HMdUrds and 31 dTGs formed per 1 X 10(6) thymidine residues. When Fe(II)/ethylenediaminetetraacetic acid was added to TPA-activated PMNs at 0, 10, 15, and 20 min after TPA, HMdUrd formation increased 5-, 13-, 30-, and 35-fold. Although dTG was initially formed in larger amounts than HMdUrd, it eventually decreased but was still 5-, 6-, 5.5-, and 3-5-fold, respectively, higher than in the absence of iron. From 65 to 1800 times more HMdUrd was formed in DNA when autologous plasma was present during incubation of DNA with TPA-activated PMNs than in its absence. The levels of dTG also varied from about the same as HMdUrd to the nondetectable. Reconstituted human serum transferrin used instead of plasma or Fe(II) also supported the formation of HMdUrd and dTG. When DNA was treated with Fe(II)-reduced H2O2 in the absence of PMNs and TPA, both derivatives were formed. However, the same treatment of marker dTG of dTG-containing polydeoxyadenylic-thymidylic acid caused the decomposition of dTG. Thus, the reduction of hydrogen peroxide by Fe(II) complexed to either ethylenediaminetetraacetic acid or amino acids amy be responsible for the formation of HMdUrd and dTG and for subsequent decomposition of dTG in DNA exposed to the TPA-activated PMNs.

DNA↗
PubMed1985

Quantitative determination of the 5-(hydroxymethyl)uracil moiety in the DNA of gamma-irradiated cells.

5-(Hydroxymethyl)uracil (HMUra) is a chemically stable derivative of thymine formed through the action of ionizing radiation which we previously identified in the DNA of gamma-irradiated HeLa cells [Teebor, G. W., Frenkel, K., & Goldstein, M. S. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 318-321]. In this report, we determine whether HMUra can be used as a marker of exposure of DNA to ionizing radiation. Dose-response curves for its formation in [3H]thymidine-labeled DNA were constructed by exposing the DNA to increasing amounts of gamma-radiation and measuring the HMUra content. DNA was irradiated both in solution and in intact cells. HMUra was identified as the 2'-deoxyribonucleoside 5-(hydroxymethyl)-2'-deoxyuridine (HMdU) by subjecting the irradiated DNA to enzymatic digestion and analyzing the mixture of 2'-deoxyribonucleosides by high-pressure liquid chromatography. The identity of the radiogenically formed HMdU was confirmed by acetylation and the structure of the acetyl derivative obtained by mass and nuclear magnetic resonance spectroscopies. At two different DNA concentrations in solution, the same number of thymidine moieties were converted to HMdU, indicating that within this range of concentration the formation of HMdU was mediated through the indirect action of ionizing radiation. Equal amounts of HMdU were formed in single- and double-stranded DNA at each radiation dose, indicating that DNA conformation did not affect HMdU formation. Surprisingly, the G value (number of HMdU molecules formed/100 eV) was higher in irradiated cellular DNA than in DNA irradiated in solution.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylation↗
PubMed1984

Ionizing radiation and tritium transmutation both cause formation of 5-hydroxymethyl-2'-deoxyuridine in cellular DNA.

HeLa cells grown in the presence of [methyl-3H]thymidine contained large amounts of 5-hydroxymethyl-2'-deoxyuridine (HMdU) in their DNA. When the cells were grown in [6-3H]thymidine and their DNA was labeled to the same specific activity, no HMdU was present. When such [6-3H]thymidine-labeled cells were exposed to increasing amounts of gamma-radiation, small but increasing amounts of HMdU were formed in their DNA. This indicates that HMdU can be formed in DNA by two distinct mechanisms. The first is the result of the transmutation of 3H to 3He (beta decay) in the methyl group of thymidine, leading to formation of a carbocation. This short-lived ion reacts with hydroxide ions of water, yielding the hydroxymethyl group. HMdU that is formed by this mechanism is formed at the rate of beta decay of 3H. It appears only in [methyl-3H]thymidine residues and is present in the DNA of both nonirradiated and gamma-irradiated cells. The second mechanism is the result of the radiolysis of water caused by ionizing radiation. The resultant radical species, particularly hydroxyl radicals, may react with many sites on DNA. When the methyl group of thymine is attacked by hydroxyl radicals, the hydroxymethyl group is formed. The formation of HMdU by this mechanism was detected only when [6-3H]thymidine-labeled cells were used, since transmutation of 3H in position 6 of thymine cannot yield HMdU.

DNA↗
PubMed1983

Free oxygen radicals: necessary contributors to tumor promotion and cocarcinogenesis.

The two stage model of carcinogenesis postulates that agents which lead to tumor formation are either initiating or promoting agents. Initiating agents induce chemical modifications of DNA. Promoting agents have been assumed to induce cell proliferation and thereby establish a milieu in which the initiated cells can express their irreversibly altered genotype. Promoters can induce both the migration of leukocytes to the skin and stimulate their respiratory activity leading to the formation of active O2 species which cause lipid peroxidation and thereby change membrane properties. However, it was recently found that the O2 species emanating from promoter-activated leukocytes also cause formation of strand breaks both in the DNA of the leukocytes themselves and in the DNA of cells cocultivated with such leukocytes. The contribution of activated O2 species to tumor promotion has been confirmed by the finding that antioxidants and copper containing superoxide dismutase (SOD)-mimetic compounds, protease inhibitors and retinoids (which block free radical formation) inhibit promotion. Furthermore, cocarcinogens such as gossypol and pyrogallol may also cause conversion of O2 to the superoxide anion radical. The heritable change caused by initiators may render the cell more vulnerable to promoter-mediated DNA damage. Experimental investigation of these hypotheses may elucidate our understanding of the contribution of promoter-mediated DNA damage in tumorigenesis and identify preventive agents for carcinogenesis.

Animals↗
PubMed1982

Identification of radiation-induced thymine derivatives in DNA.

A methodology for the separation of radiation-induced thymine derivatives in DNA using high pressure liquid chromatography is presented. DNA was subjected to enzymatic hydrolysis yielding 2'-deoxyribonucleosides and the hydrolysate cochromatographed with marker compounds. Confirmation of the presence of derivatives was accomplished by chromatography on Sephadex LH-20 and microderivatization. The method separates free bases from nucleosides allowing for identification of spontaneously released bases or those released through the action of repair enzymes. The results indicate that most of the thymine derivatives formed in irradiated cellular DNA were the same as those found in DNA irradiated in solution. However, the major cellular derivative was not present in the latter. This derivative was identified as 5-hydroxymethyl-2'-deoxyuridine (HMdU). HMdU has previously been shown to be cytotoxic to cells in culture and caused diarrhea and bone marrow failure when administered to mice. Thus, the presence of this radiation-induced thymine derivative in cellular DNA correlates with the known effects of ionizing radiation on cells and animals.

DNA↗
PubMed1981

Identification of the cis-thymine glycol moiety in chemically oxidized and gamma-irradiated deoxyribonucleic acid by high-pressure liquid chromatography analysis.

5,6-Dihydroxy-5,6-dihydrothymine (thymine glycol) is formed in DNA by chemical oxidants and ionizing radiation. We describe the separation of thymine glycol, 5,6-dihydroxy-5,6-dihydrothymidine (thymidine glycol), thymine, and thymidine by high-pressure liquid chromatography (HPLC). Enzymatic hydrolysates of chemically oxidized or gamma-irradiated single-stranded DNA were cochromatographed with 14C-containing marker compounds. In chemically oxidized DNA, thymidine glycol was the major derivative formed. In addition, there were four rapidly eluting thymine-derived components. In irradiated DNA, thymidine glycol constituted about 5% of the modified thymines, and the rapidly eluting fractions were proportionately increased. DNA isolated from gamma-irradiated and nonirradiated HeLa cells grown in the presence of [3H]thymidine was subjected to enzymatic hydrolysis and HPLC analysis. In control DNA, 0.3% of the thymines were modified. Thirty-six kilorads of gamma radiation caused a 30% increase in thymine damage. Thus, most of the base damage was due to internal beta radiation from incorporated [3H]thymidine. The chromatographic patterns of irradiated and nonirradiated samples were qualitatively the same, but the yields of some products increased 2-fold, while others remained unchanged. A comparison of the HPLC profiles of hydrolysates of in vitro oxidized and irradiated DNA with those of the cellular DNA revealed one fast eluting peak to be absent in cellular DNA, suggesting that it was formed only in single-stranded DNA. In cellular DNA, the major modified thymine was a more hydrophobic derivative not formed by in vitro radiation nor chemical oxidation. As in in vitro irradiated DNA, thymidine glycol constituted 5% of the modified thymines. The presence of cis-thymidine glycol in hydrolysates was confirmed by chromatography on Sephadex LH-20 using water and borate as eluants.

Chromatography, High Pressure Liquid↗
PubMed1981

Identification of novel 7,12-dimethylbenz[a]anthracene adducts in cellular ribonucleic acid.

The interaction of guanosine with 7,12-dimethylbenz[a]anthracene (DMBA) 5,6-oxide under alkaline conditions resulted in the formation of six derivatives. These six compounds were cochromatographed with nucleosides obtained by hydrolysis of RNA isolated from rat liver cells treated with [3H]DMBA. The cochromatography showed that three of these adducts were formed in cellular RNA. The three products constituted less than 5% of the total nucleoside-DMBA adducts as shown by chromatography on Sephadex LH-20 and high-pressure liquid chromatography. In one of them, the 2'-hydroxy group of the ribose moiety of guanosine was linked to the C-5, and in the second, to the C-6 position of the DMBA 5,6-oxide residue. In the third derivative, the C-8 position of guanosine was linked to the C-5 of the DMBA 5,6-oxide moiety. These results show, for the first time, modifications of the ribose moiety and of the guanine residue at the C-8 position in the cellular RNA by a metabolite of a polycyclic hydrocarbon.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1981

Identification of the cis-thymine glycol moiety in oxidized deoxyribonucleic acid.

5,6-Dihydroxy-5,6-dihydrothymine (thymine glycol) is formed in DNA by reaction with oxidizing agents and as a result of ionizing and near-ultraviolet radiation. We describe a rapid purification of cis-5,6-dihydroxy-5,6-dihydrothymine and cis-5,6-dihydroxy-5,6-dihydrothymidine (cis-thymidine glycol) and their use as markers in identifying the thymine glycol moiety in oxidized DNA. Both glycols were prepared by oxidation of [14C]thymine and -thymidine with KMnO4 followed by purification on Sephadex LH-20 (LH-20). [3H]DNA was oxidized with KMnO4 and the thymidine glycol in DNA identified by enzymatic digestion of the DNA followed by cochromatography of the digest with marker [14C]thymidine glycol on LH-20. The cis conformation of the glycol was confirmed by the change in the elution pattern when borate rather than water was used as eluent. Alkaline hydrolysis of a mixture of [14C]thymine glycol and oxidized [3H]DNA followed by trichloroacetic acid precipitation and LH-20 chromatographic analysis of the neutralized supernatant yielded a complex pattern of radioactive degradation products with coincidence of one 14C marker- and one [3H]-DNA-derived peak. All applied radioactivity was recovered. This methodology should be useful in determining thymine glycol content of irradiated DNA and in elucidating the mechanism by which these altered residues are removed from cellular DNA by repair enzymes.

Chemical Phenomena↗
PubMed1978

Conformation of dinucleoside monophosphates modified with benzo[a]pyrene-7,8-dihydrodiol 9,10-oxide as measured by circular dichroism.

The conformational properties of GpU modified with the reactive derivative of benzo[a]pyrene, (+/-)-7beta,8alpha-dihydroxy-9alpha,10alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene, has been investigated utilizing circular dichroism spectroscopy. Binding of this carcinogen to the N2 of G residues in GpU resulted in the formation of four compounds (I to IV) representing two pairs of diastereoisomers. The molar ellipticity values of the modified dimers were approximately twofold higher than those of the modified guanosine monomers. These values were decreased appreciably when the spectra of the dimers were obtained at 80 degrees C or in methanol rather than at 25 degrees C in water, suggesting that under the latter conditions there is a stacking interaction between the carcinogen and the neighboring uridine residue. Based on these results, a conformation is proposed for modified GpU. It includes insertion of the benzo[a]pyrene moiety, by rotation of the modified guanine residue about its glycoside bond, coplanar to the neighboring uridine and perpendicular to the phosphodiester backbone.

Benzopyrenes↗
PubMedDate not supplied

Decomposition of nucleoside hydroperoxide by metals and metalloproteins.

5-Hydroperoxymethyl-2'-deoxyuridine (HPMdU) is formed in DNA by ionizing radiation. Although relatively stable, HPMdU eventually decomposes to two products 5-hydroxymethyl-2'-deoxyuridine (HMdU) and 5-formyl-2'-deoxyuridine (FdU). We show that a number of transition metal ions and metalloproteins accelerate this process. Of the metal ions tested, Sn(II) and Fe(II) were the most active, with the former producing exclusively HMdU, and the latter, a mixture of both. Cu(I), Cu(II), Co(II), and Ni(II) induced a predominant generation of FdU, with copper ions being more effective than Co and Ni. FdU was also preferentially formed in the presence of the iron-containing proteins transferrin and ferritin, whereas HMdU was the major product in the presence of apotransferrin as well as in the presence of ceruloplasmin, a copper-containing protein.

Chemical Phenomena↗