Inhibition of ADP-ribosylation of histone by diadenosine 5′, 5‴-p1, p4-tetraphosphate☆
Abstract
Diadenosine 5′, 5‴-p1, p4-tetraphosphate (Ap4A) strongly inhibited ADP-ribosylation reaction of histone by purified bovine thymus poly(ADP-ribose) polymerase. This compound showed a relatively weak inhibitory effect on Mg2+-dependent, enzyme-bound poly(ADP-ribose) synthesis. Among various adenine nucleotides tested, including several diadenosine nucleotides with varying phosphate chain length, Ap4A was the most effective inhibitor of the histone-modification reaction. Ap5A and Ap6A showed slightly lower inhibitory effect than Ap4A. Kinetic analysis of the inhibitor (Ap4A) with varying concentration of substrate (NAD+) revealed that this compound is a “mixed type inhibitor”, with a Ki value of 5.1 μM.
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Cited by (18)
Isothermal titration calorimetry reveals a zinc ion as an atomic switch in the diadenosine polyphosphates
2002, Journal of Biological ChemistryDiadenosine polyphosphates (diadenosine 5′,5‴-P1,Pn-polyphosphate (ApnA)) are 5′-5‴-phosphate-bridged dinucleosides that have been proposed to act as signaling molecules in a variety of biological systems. Isothermal titration calorimetry was used to measure the affinities of a variety of metal cations for ATP, diadenosine 5′,5‴-P1,P3-triphosphate (Ap3A), diadenosine 5′,5‴-P1,P4-tetraphosphate (Ap4A), and diadenosine 5′,5‴-P1,P5-pentaphosphate (Ap5A). The binding of Mg2+, Ca2+, and Mn2+ to ATP is shown to take place with the β,γ-phosphates (primary site) and be endothermic in character. The binding of Ni2+, Cd2+, and Zn2+ to ATP is found to take place at both the primary site and at a secondary site identified as N-7 of the adenine ring. Binding to this second site is exothermic in character. Generally, the binding of metal cations to diadenosine polyphosphates involves a similar primary site to ATP. No exothermic binding events are identified. Critically, the binding of Zn2+ to diadenosine polyphosphates proves to be exceptional. This appears to involve a very high affinity association involving the N-7 atoms of both adenine rings in each ApnA, as well as the more usual endothermic association with the phosphate chain. The high affinity association is also endothermic in character. A combination of NMR and CD evidence is provided in support of the calorimetry data demonstrating chemical shift changes and base stacking disruptions entirely consistent with N-7 bridging interactions. N-7 bridging interactions are entirely reversible, as demonstrated by EDTA titration. Considering the effects of Zn2+ on a wide variety of dinucleoside polyphosphate-metabolizing enzymes, we examine the possibility of Zn2+ acting as an atomic switch to control the biological function of the diadenosine polyphosphates.
Is Ap<inf>4</inf>A involved in DNA repair processes?
1988, Experimental Cell ResearchHepatoma tissue culture (HTC) cells were incubated in the presence of the alkylating agent N-methyl-N′-nitro-N-nitrosoguanidine (MNNG) to study the variations in the bisnucleosides polyphosphates (Ap4X) pool size. A transient but sensitive accumulation of these compounds is observed; if 3-aminobenzamide (3AB) which is a potent inhibitor of the ADP-ribosyltransferase (ADPRT) is added after the MNNG treatment, a more pronounced and persistent accumulation of Ap4X can be seen. A moderate heat-shock (30 min at 43 °C) results also in a small accumulation of Ap4X but the shape of the accumulation curve is quite different and the increase of the Ap4X pool is not sensitive to the presence of 3AB. However, both MNNG treatment and hyperthermia cause a marked inhibition of protein synthesis. On the other hand, the ADPRT activity is enhanced in the presence of MNNG whereas hyperthermia has little or a slightly inhibitory effect on this activity. These results suggest that MNNG treatment triggers an Ap4X accumulation in eukaryotic cells different from that observed after heat-shock and it seems likely that these compounds are involved in the DNA excision repair system in which the ADPRT enzyme is also implicated.
Diadenosine tetraphosphate as a signal molecule linked with the functional state of rat liver
1985, GastroenterologyDiadenosine 5′,5″-P1,P4-tetraphosphate (Ap4A), which has been suggested to be a signal nucleotide involved in positive growth regulation, was determined in rat liver under various conditions using the luciferin/luciferase system. The Ap4A level was 920 pmol/g of wet liver in normal adult rats, but was high at early stages of growth, the maximum value being 1460 pmol/g in 3-wk-old rats. An increase in Ap4A levels preceded deoxyribonucleic acid synthesis in liver after hepatectomy. Ap4A levels were increased 5.5-fold (5125 pmol/g) 24 h after hepatectomy. The range of such increases was closely correlated with the mass of liver removed. Cholestasis produced by bile duct ligation and intraperitoneal injection of D-galactosamine remarkably lowered the Ap4A level to 20% and 42%, respectively; however, the adenosine triphosphate level was only slightly decreased to 65% and 78%, respectively. Fourteen days after bile duct ligation, Ap4A levels fell to <220 pmol/g, and the rats could no longer survive partial hepatectomy. These results suggest that hepatic Ap4A levels correlate positively with regenerative activity.
Formaldehyde-induced mutagenesis: A novel mechanism for its action
1985, Mutation Research/Reviews in Genetic ToxicologyA novel and unique mechanism for formaldehyde-induced mutagenesis is described which is mediated by the formation of an N6-substituted adenine ribonucleoside analogue, N6-hydroxymethyl adenosine, after an in vitro reaction of formaldehyde with adenosine. This type of ribonucleoside analogue (the deoxyribose derivative is ineffective) exhibits a powerful and remarkable germ-cell-stage-specific mutagenic effect in male Drosophila larvae, apparently by interfering with DNA repair.
Circumstantial evidence is presented which indicates that the analogue most probably acts by its utilisation in the synthesis of diadenosine tetraphosphate (Ap4A) to form an antimetabolite(s) of Ap4A which subsequently interferes with Ap4A-mediated intracellular events, amongst which an effect on DNA repair would appear to be its mutagenic mechanism of action.
Determination of diadenosine 5′,5‴,-P<sup>1</sup>,P<sup>4</sup>-tetraphosphate levels in cultured mammalian cells
1984, Analytical BiochemistryA simple method for measuring the cellular content of diadenosine 5′,5‴-P1,P4-tetraphosphate (Ap4A) in cultured mammalian cells is described. Ap4A was rapidly extracted by dissolving cell monolayers using 0.1 n NaOH. It was separated from the bulk of cellular components in a single step by selective adsorption to a highly specific boronate affinity resin. Subpicomole amounts were quantified by a luciferin-luciferase bioluminescence assay performed in the presence of alkaline phosphatase and venom phosphodiesterase. The selectivity of this assay for Ap4A in cultured mouse cells was established by high-performance liquid chromatography. This method allows the routine measurement of subpicomole amounts of Ap4A derived from a single dish of cells.
Do stalled replication forks synthesize a specific alarmone?
1983, Journal of Theoretical BiologyPotential causes of premature arrest of a replication fork in vivo include an encounter with a chemical lesion in the DNA, inhibition of one of the essential enzymes of the fork, and spontaneous failure of the fork due to its finite degree of processivity. I suggest that a premature arrest of either a eukaryotic or prokaryotic replication fork induces it to enter a different state in which the fork synthesizes a specific signal nucleotide (“alarmone”). One function of the postulated new alarmone would be to increase the probability of re-initiation of DNA replication, either in cis (at an origin proximal to a site of the fork arrest) or in trans (at many different origins). An additional, mechanistically related function of the postulated alarmone could be to increase the probability of re-assembly of an arrested fork beyond an otherwise impassable DNA lesion. In case of multiple fork arrests, an alarmone-mediated increase in the probability of replicon reinitiation (disproportionate DNA replication) would result in gene amplification at many different loci, thereby increasing the probability of cell's survival in a cytotoxic medium. Other likely functions of a fork-produced alarmone may include stimulation of DNA repair pathways including excision repair. I review the experimental evidence which although indirect, is consistent with the idea of a fork-produced alarmone and specifically with the possibility that the postulated alarmone is diadenosine 5', 5′′′P1, P4-tetraphosphate (Ap4A) or a closely related adenylated nucleotide. The proposed hypothesis leads to specific, testable predictions; it also provides a unifying explanation for several hitherto unconnected observations on DNA replication, repair and amplification.
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This work was supported in part by a Grant in aid for Cancer Reaction (501036) and a research grant (558078) from the Minitry of Education, Science and Culture of Japan.