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1.
Rapid turnover of endogenous endonuclease activity in thymocytes: effects of inhibitors of macromolecular synthesis 总被引:4,自引:0,他引:4
Previous work has shown that inhibitors of protein or mRNA synthesis block endonuclease activation in thymocytes undergoing programmed cell death. In the present study we used isolated nuclei to investigate the effects of cycloheximide and actinomycin D, inhibitors of protein and mRNA synthesis, respectively, on endogenous endonuclease activity in thymocytes. We observed a rapid loss of Ca2(+)-dependent endonuclease activity in nuclei isolated from thymocytes treated with these inhibitors. In contrast, pretreatment of cells with antipain and leupeptin, inhibitors of proteases, prevented the depletion of endonuclease activity in the nuclei, suggesting that proteolysis was involved. The effects of cycloheximide and actinomycin D were mimicked by incubating thymocytes with treatments known to exert their effects via activation of protein kinase C. Our results suggest that endonuclease activity in thymocyte nuclei undergoes rapid, spontaneous turnover. Agents interfering with macromolecular synthesis may therefore block DNA fragmentation in thymocytes by depleting nuclei of endogenous endonuclease activity. 相似文献
2.
We have detected Ca2+, Mg2+-dependent endonuclease activity in spleen cells of normal, Friend erythroleukemic, and phenylhydrazine-treated mice. When nuclei were isolated and incubated in the presence of Ca2+ and Mg2+ ions, the activity resulted in the production of 3'-OH termini in the cellular DNA and the release of chromatin due to internucleosomal DNA fragmentation. This enzyme activity was chromatin-bound and could be extracted from chromatin in an active form in 0.35 M KCl. The majority of endonuclease activity from erythroleukemic spleens was present in nuclei of precursor erythroid cells of low buoyant density (1.025-1.05 g/ml). Uninfected normal splenic tissue contained an endonuclease activity which was almost entirely confined to a B-lymphocyte population of high buoyant density (greater than 1.07 g/ml). Erythroid cell-enriched spleens from phenylhydrazine-treated mice exhibited a distribution of endonuclease activity in cells at low and high densities reflecting a mixture of erythroid and lymphoid cells. Cloned erythroleukemic cell lines propagated in vitro lacked cells of low density and showed no detectable endonuclease activity. However, nuclei from these cell lines were susceptible to exogenously added endonuclease extracted from erythroleukemic spleen cells. These same cell lines propagated as subcutaneous tumors contained endonuclease activity and a morphologically-similar low-density cell population which accounted for the endonuclease activity in these tumors. Nuclei from cloned lymphoid cell lines, representing different B-lymphocyte phenotypes, showed differences in the presence of endonuclease activity. Among the cell lines tested, only those expressing late B-cell markers showed detectable endonuclease activity. 相似文献
3.
Bae SH Kim DW Kim J Kim JH Kim DH Kim HD Kang HY Seo YS 《The Journal of biological chemistry》2002,277(29):26632-26641
Saccharomyces cerevisiae Dna2 possesses both helicase and endonuclease activities. Its endonuclease activity is essential and well suited to remove RNA-DNA primers of Okazaki fragments. In contrast, its helicase activity, although required for optimal growth, is not essential when the rate of cell growth is reduced. These findings suggest that DNA unwinding activity of Dna2 plays an auxiliary role in Okazaki fragment processing. To address this issue, we examined whether the Dna2 helicase activity influenced its intrinsic endonuclease activity using two mutant proteins, Dna2D657A and Dna2K1080E, which contain only helicase or endonuclease activity, respectively. Experiments performed with a mixture of Dna2D657A and Dna2K1080E enzymes revealed that cleavage of a single-stranded DNA by endonuclease activity of Dna2 occurs while the enzyme translocates along the substrate. In addition, DNA unwinding activity efficiently removed the secondary structure formed in the flap structure, which was further aided by replication protein A. Our results suggest that the Dna2 unwinding activity plays a role in facilitating the removal of the flap DNA by its intrinsic endonuclease activity. 相似文献
4.
Differential effects of Ca2+ and Mg2+ on endonuclease activation in isolated promyelocytic HL-60 cell nuclei 总被引:1,自引:0,他引:1
Apoptosisorprogrammedcelldeath(PCD)isaprocessofcelldeletionwhichoccursinresponsetoanumberofcytotoxicandphysiologicallyrelevantstimuli.Thisprocessischaracterizedbyseveralearlymorphologicalterationsincludingplasmaandnuclearmembraneblebbing.Endogenousendo… 相似文献
5.
beta-globin mRNA bearing a nonsense codon is degraded in the cytoplasm of erythroid cells by endonuclease cleavage, preferentially at UG dinucleotides. An endonuclease activity in polysomes of MEL cells cleaved beta-globin and albumin mRNA in vitro at many of the same sites as PMR1, an mRNA endonuclease purified from Xenopus liver. Stable transfection of MEL cells expressing normal human beta-globin mRNA with a plasmid vector expressing the catalytically active form of PMR1 reduced the half-life of beta-globin mRNA from 12 to 1-2 h without altering GAPDH mRNA decay. The reduced stability of beta-globin mRNA in these cells was accompanied by an increase in the production of mRNA decay products corresponding to those seen in the degradation of nonsense-containing beta-globin mRNA. Therefore, beta-globin mRNA is cleaved in vivo by an endonuclease with properties similar to PMR1. Inhibiting translation with cycloheximide stabilized nonsense-containing beta-globin mRNA, resulting in a fivefold increase in its steady-state level. Taken together, our results indicate that the surveillance of nonsense-containing beta-globin mRNA in erythroid cells is a cytoplasmic process that functions on translating mRNA, and endonucleolytic cleavage constitutes one step in the process of beta-globin mRNA decay. 相似文献
6.
The endogenous endonuclease activity of chromatin in isolated rat liver nuclei in the presence of Mn2+, Mg2+ and Ca2+ + Mg2+ was studied. The existence of a Mn2+-dependent endonuclease activity not coupled with the Ca2+, Mg2+-dependent endonuclease was demonstrated, which was weaker than the former one in isolated cell nuclei but higher than in the preparation of Ca2+, Mg2+-dependent nuclease obtained by gel filtration through Toyopearl HW 60F. The Mn2+-dependent splitting of chromatin predominantly occurs at linker DNA of distal parts of chromatin loops. A split-off of purified DNA was more universal than in the presence of Ca2+, Mg2+-dependent endonuclease; the hydrolysis rate of native and denaturated DNA appeared to be the same. 相似文献
7.
Lee KH Kim DW Bae SH Kim JA Ryu GH Kwon YN Kim KA Koo HS Seo YS 《Nucleic acids research》2000,28(15):2873-2881
Dna2 is a multifunctional enzyme in yeast that possesses endonuclease activity well suited to remove RNA–DNA primers of Okazaki fragments, raising the question of whether endonuclease activity is essential for in vivo Dna2 function. Systematic site-directed mutations of amino acid residues in Saccharomyces cerevisiae DNA2 conserved in the central region of many eukaryotic DNA2 homologs allowed us to identify mutant dna2 alleles that were divided into three groups based on the viability of the mutant cells: (i) viable; (ii) inviable only when expression was repressed; (iii) inviable. Biochemical analyses of recombinant mutant Dna2 proteins isolated from the latter two groups revealed that they possessed normal ATPase/helicase activity, but were impaired in their endonuclease activity. Cells expressing mutant Dna2 enzymes partially impaired in endonuclease activity were viable, but were unable to grow when expression of their mutant Dna2 enzymes was further reduced. Their growth was restored when the mutant Dna2 proteins decreased in nuclease activity were induced to overexpress. In contrast, mutant Dna2 proteins lacking endonuclease activity did not allow cells to grow under any conditions tested. These in vivo and in vitro results demonstrate that the endonuclease activity of Dna2 is essential for Okazaki fragment processing. 相似文献
8.
DNase Induced After Infection of KB Cells by Herpes Simplex Virus Type 1 or Type 2 II. Characterization of an Associated Endonuclease Activity
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Purified preparations of the "exonuclease" specified by herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) possess an endonuclease activity. The exonuclease and endonuclease activities copurify and cosediment in a sucrose density gradient. Endonuclease activity is only observed in the presence of a divalent cation, and Mg(2+) or Mn(2+) is equally effective as a cofactor with an optimal concentration of 2 mM. A slight amount of endonuclease activity is observed in the presence of Ca(2+), whereas no activity occurs in the presence of Zn(2+). In the presence of Mg(2+), Ca(2+) and Zn(2+) are inhibitory. Comparison of exonuclease and endonuclease activity in the presence of various divalent cations revealed that, at concentrations of Mn(2+) greater than 1 mM, only endonuclease activity occurs whereas endonuclease and exonuclease activity occur at all concentrations of Mg(2+). The endonuclease was affected by putrescine and spermidine to the same extent as the exonuclease activity, but in marked contrast the endonuclease was inhibited by a 10-fold-lower concentration of spermine compared to the exonuclease. The activity specified by HSV-1 and HSV-2 has very similar properties. HSV-1 and HSV-2 endonuclease cleave covalently closed circular DNA to yield, firstly, nicked circles and then linear DNA which is subsequently hydrolyzed to small oligonucleotides. Cleavage does not appear to be base sequence specific. Conversion of nicked circles to linear DNA and subsequent degradation of linear DNA occurs more rapidly in the presence of Mg(2+) than Mn(2+) presumably by virtue of the presence of the exonuclease activity. Nonsuperhelical covalently closed circular duplex DNA is cleaved by the endonucleases at a rate 60 times slower than the rate observed on the supercoiled form. These data indicate that the HSV-1 and HSV-2 endonuclease preferentially recognize single-stranded DNA regions. 相似文献
9.
Adeno-associated virus type 2 Rep endonuclease activity is necessary for both viral DNA replication and site-specific integration of the viral genome into human chromosome 19. The biochemical activities required for site-specific endonuclease activity (namely specific DNA binding and transesterification activity) have been mapped to the amino-terminal domain of the AAV2 Rep protein. The amino-terminal 208 amino acids are alone sufficient for site-specific endonuclease activity, and nicking by this domain is metal-dependent. To identify this metal-binding site, we have employed a cysteine mutagenesis approach that targets conserved acidic amino acids. By using this technique, we provide functional biochemical data supporting a role for glutamate 83 in the coordination of metal ions in the context of Rep endonuclease activity. In addition, our biochemical data suggest that glutamate 164, although not involved in the coordination of metal ions, is closely associated with the active site. Thus, in lieu of a crystal structure for the AAV type 2 amino-terminal domain, our data corroborate the recently published structural studies of the AAV type 5 endonuclease and suggest that although the two enzymes are not highly conserved with respect to the AAV family, their active sites are highly conserved. 相似文献
10.
Bcl2 not only prolongs cell survival but also suppresses the repair of abasic (AP) sites of DNA lesions. Apurinic/apyrimidinic endonuclease 1 (APE1) plays a central role in the repair of AP sites via the base excision repair pathway. Here we found that Bcl2 down-regulates APE1 endonuclease activity in association with inhibition of AP site repair. Exposure of cells to nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone results in accumulation of Bcl2 in the nucleus and interaction with APE1, which requires all of the BH domains of Bcl2. Deletion of any of the BH domains from Bcl2 abrogates the ability of Bcl2 to interact with APE1 as well as the inhibitory effects of Bcl2 on APE1 activity and AP site repair. Overexpression of Bcl2 in cells reduces formation of the APE1.XRCC1 complex, and purified Bcl2 protein directly disrupts the APE1.XRCC1 complex with suppression of APE1 endonuclease activity in vitro. Importantly, specific knockdown of endogenous Bcl2 by RNA interference enhances APE1 endonuclease activity with accelerated AP site repair. Thus, Bcl2 inhibition of AP site repair may occur in a novel mechanism by down-regulating APE1 endonuclease activity, which may promote genetic instability and tumorigenesis. 相似文献
11.
Tumour necrosis factor alpha (TNF-alpha) at 20 ng/ml induced apoptosis in human hepatoma cells in vitro. The effect of TNF-alpha-induced apoptosis was exacerbated by the hypoxanthine-xanthine oxidase (HX/XO) system and cycloheximide (CHX), but alleviated by superoxide dismutase (SOD), suggesting that TNF-alpha-induced apoptosis may be due to oxidative stress, and independent of protein synthesis. TNF-alpha elevated free Ca(2+)concentration, triggered lipid peroxidation and decreased the expression of bcl-2 protein. The findings suggest that TNF-alpha-induced apoptosis may be involved in stimulating Ca(2+)-dependent endonuclease activity and increasing membrane lipid peroxidation. Bcl-2 may play a pivotal role in serving as a Ca(2+)regulator or antioxidant, preventing lipid peroxidation in the process. 相似文献
12.
In autodigestion assays, endonucleaw activity in non-apoptotic HL-60 promydocytic leukemia cell nuclei cleaved the chromatin of he autologous cells to an oligonucleosomal length pattern. Both EGTA and EDTA inhibited the activation of endonuclease activity in isolated HL-60 cell nuclei. The inhibition by EDTA could be reversed by exogenous Ca2+. but not by exogenous Mg2+. In Ca2+/Mg2+-free nuclei digation buffer, addition of Ca2→ (1-10 mmol/L) induced endonuclease activity in the isolated nuclei, while addition of Mg2+ had no effect. In the presence of Ca2+(0.1 mmol/L), endonuclease activity was enhanced by exogenous Mg2+ (0.1-10mmol/L). These results suggest that the endonuclease responsible for internucleosomal DNA fragmentation in HL-60 cells during apoptosis is activated by Ca2+ and further modulated by Mg2+ in the presence of ca2+. 相似文献
13.
Apoptosis: mode of cell death induced in T cell leukemia lines by dexamethasone and other agents 总被引:10,自引:0,他引:10
Glucocorticoids can mediate the destruction of thymocytes and T cell-derived leukemia cells through a mechanism known as apoptosis. The characteristic feature of apoptosis is fragmentation of DNA at internucleosomal linkers through the activity of a specific endonuclease. In this study, an attempt was made to compare dexamethasone-induced apoptosis in two T cell-derived human leukemia lines (CEM-C1 and CEM-C7) to the cell killing brought about by selected cytotoxic agents. In the CEM-C7 cell line (dexamethasone-sensitive), apoptosis was induced not only by dexamethasone but by actinomycin D, cycloheximide, and 25-OH cholesterol. In the CEM-C1 cell line (dexamethasone-resistant) cycloheximide, 25-OH cholesterol, or cell starvation could induce apoptosis. It appears that in leukemic cells apoptosis may be induced by a variety of unrelated toxic agents and is not limited to glucocorticoids. 相似文献
14.
Protection from tumor necrosis factor-mediated cytolysis by overexpression of plasminogen activator inhibitor type-2 总被引:14,自引:0,他引:14
Pretreatment of HT-1080 fibrosarcoma cells with tumor necrosis factor (TNF) induced resistance to the cytolytic activity of this cytokine in combination with cycloheximide. This resistance correlated with the synthesis of plasminogen activator inhibitor type-2 (PAI-2). HT-1080 cells were transfected with a PAI-2 expression vector in both sense and antisense orientation. The resistance to TNF-mediated cytolysis of transfected cell clones was correlated with the level of PAI-2 expression. Cells expressing antisense PAI-2 RNA showed reduced expression of PAI-2 and increased sensitivity to TNF-mediated cytolysis. Cells expressing constitutively PAI-2 were treated with TNF and cycloheximide to select cells with increased resistance to cytolysis and enhanced PAI-2 expression. PAI-2 gradually disappeared during a treatment with TNF and cycloheximide. This finding suggested that PAI-2 formed a complex with a target proteinase, which could be involved in mediating the cytolytic activity of TNF. 相似文献
15.
Double-stranded RNA causes synthesis of 2',5'-oligo(A) and degradation of messenger RNA in interferon-treated cells 总被引:5,自引:0,他引:5
Interferon-treated HeLa cells were incubated with [3H]uridine to label mRNA and were then exposed to the double-stranded RNA poly(inosinic acid).poly(cytidylic acid) (In.Cn). The incubation with In.Cn greatly enhanced the decay of mRNA. When the cells were incubated in this way in the presence of cycloheximide, which blocks ribosome movement along mRNA, extensive polysome degradation was detected in interferon-treated cells. Products of degradation of mRNA were recovered from monosomes which were presumably formed as a result of endonucleolytic breaks of mRNA. This endonucleolytic activity was correlated with the formation of 2',5'-oligo(A) by an enzyme induced by interferon and activated by double-stranded RNA; the 2',5'-oligo(A) was previously shown to activate an endonuclease in cell extracts. The 2',5'-oligo(A) levels in cells were measured by a competition-binding assay. Details of the procedure used are described, including synthesis of highly radioactive (2'-5')pppA3[32P]cytidine 3',5'-diphosphate, separation of 2',5'-oligo(A) binding from degrading activities, and specificity of the assay. 相似文献
16.
Spectra of thymocyte nuclear DNAases of control and irradiated (4 Gy) rats have been investigated. Using the method of SDS-electrophoresis of nuclear proteins in DNA-polyacrylamide gel (PAAG) the authors managed to discover a number of polypeptides of 35, 32, 17.7, 17.2, and 16.4 kDA molecular mass possessing a DNAase activity. The enzyme of 35 kDA is only active in the presence of Ca2+ and Mg2+ ions. Nucleases of 32, 17.7, 17.2, and 16.4 kDA are active in the presence of Ca2+ ions and inactive in the presence of Mg2+ ions or in the absence of divalent cations. A simultaneous addition of Ca2+ and Mg2+ ions to the incubation medium causes a synergistic effect with respect to the manifestation of these DNAase activities. Nucleases of 32, 17.7, 17.2, and 16.4 kDa only emerge after the preliminary removal of histones by ion exchange chromatography on a column with CM-sephadex C-50. The enzymic activity of 32 kDA protein increases 60 min after irradiation and drops to the control value in 4 h. At the same time, the postirradiation increase in DNAase activity of a low-molecular weight enzyme group remains invariable throughout the entire period of observation (1-4 h). The preinjection of cycloheximide (CHI) prevents the postirradiation degradation of chromatin and, simultaneously, makes the enzymic activity, corresponding to 35 kDA protein, disappear at the electrophoregrams. The experiments with CHI permit to identify the given enzymic fraction as Ca/Mg-dependent endonuclease. This indicates the participation of normally pre-existing Ca/Mg-dependent endonuclease in implementing the process of chromatin enzymic degradation in the irradiated thymocytes. 相似文献
17.
Purification and properties of 2-aminoadipate: 2-oxoglutarate aminotransferase from bovine kidney.
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Escherichia coli endonuclease IV hydrolyses the C(3')-O-P bond 5' to a 3'-terminal base-free deoxyribose. It also hydrolyses the C(3')-O-P bond 5' to a 3'-terminal base-free 2',3'-unsaturated sugar produced by nicking 3' to an AP (apurinic or apyrimidinic) site by beta-elimination; this explains why the unproductive end produced by beta-elimination is converted by the enzyme into a 3'-OH end able to prime DNA synthesis. The action of E. coli endonuclease IV on an internal AP site is more complex: in a first step the C(3')-O-P bond 5' to the AP site is hydrolysed, but in a second step the 5'-terminal base-free deoxyribose 5'-phosphate is lost. This loss is due to a spontaneous beta-elimination reaction in which the enzyme plays no role. The extreme lability of the C(3')-O-P bond 3' to a 5'-terminal AP site contrasts with the relative stability of the same bond 3' to an internal AP site; in the absence of beta-elimination catalysts, at 37 degrees C the half-life of the former is about 2 h and that of the latter 200 h. The extreme lability of a 5'-terminal AP site means that, after nicking 5' to an AP site with an AP endonuclease, in principle no 5'----3' exonuclease is needed to excise the AP site: it falls off spontaneously. We have repaired DNA containing AP sites with an AP endonuclease (E. coli endonuclease IV or the chromatin AP endonuclease from rat liver), a DNA polymerase devoid of 5'----3' exonuclease activity (Klenow polymerase or rat liver DNA polymerase beta) and a DNA ligase. Catalysts of beta-elimination, such as spermine, can drastically shorten the already brief half-life of a 5'-terminal AP site; it is what very probably happens in the chromatin of eukaryotic cells. E. coli endonuclease IV also probably participates in the repair of strand breaks produced by ionizing radiations: as E. coli endonuclease VI/exonuclease III, it is a 3'-phosphoglycollatase and also a 3'-phosphatase. The 3'-phosphatase activity of E. coli endonuclease VI/exonuclease III and E. coli endonuclease IV can also be useful when the AP site has been excised by a beta delta-elimination reaction. 相似文献
18.
The RAD2 family of nucleases includes human XPG (Class I), FEN1 (Class II), and HEX1/hEXO1 (Class III) products gene. These proteins exhibit a blend of substrate specific exo- and endonuclease activities and contribute to repair, recombination, and/or replication. To date, the substrate preferences of the EXO1-like Class III proteins have not been thoroughly defined. We report here that the RAD2 domain of human exonuclease 1 (HEX1-N2) exhibits both a robust 5' to 3' exonuclease activity on single- and double-stranded DNA substrates as well as a flap structure-specific endonuclease activity but does not show specific endonuclease activity at 10-base pair bubble-like structures, G:T mismatches, or uracil residues. Both the 5' to 3' exonuclease and flap endonuclease activities require a divalent metal cofactor, with Mg(2+) being the preferred metal ion. HEX1-N2 is approximately 3-fold less active in Mn(2+)-containing buffers and exhibits <5% activity in the presence of Co(2+), Zn(2+), or Ca(2+). The optimal pH range for the nuclease activities of HEX1-N2 is 7.2-8.2. The specific activity of its 5' to 3' exonuclease function is 2.5-7-fold higher on blunt end and 5'-recessed double-stranded DNA substrates compared with duplex 5'-overhang or single-stranded DNAs. The flap endonuclease activity of HEX1-N2 is similar to that of human flap endonuclease-1, both in terms of turnover efficiency (k(cat)) and site of incision, and is as efficient (k(cat)/K(m)) as its exonuclease function. The nuclease activities of HEX1-N2 described here indicate functions for the EXO1-like proteins in replication, repair, and/or recombination that may overlap with human flap endonuclease-1. 相似文献
19.
Activation of an Mg2+-dependent DNA endonuclease of avian myeloblastosis virus alpha beta DNA polymerase by in vitro proteolytic cleavage. 总被引:14,自引:12,他引:2
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Partial chymotryptic digestion of purified avian myeloblastosis virus alpha beta DNA polymerase resulted in the activation of a Mg2+-dependent DNA endonuclease activity. Incubation of the polymerase-protease mixture in the presence of super-coiled DNA and Mg2+ permitted detection of the cleaved polymerase fragment possessing DNA nicking activity. Protease digestion conditions were established permitting selective cleavage of beta to alpha, which contained DNA polymerase and RNase H activity and to a family of polypeptides ranging in size from 30,000 to 34,000 daltons. These latter beta-unique fragments were purified by polyuridylate-Sepharose 4B chromatography and were shown to contain both DNA binding and DNA endonuclease activities. We have demonstrated that this group of polymerase fragments derived by chymotryptic digestion of alpha beta DNA polymerase is similar to the in vivo-isolated avian myeloblastosis virus p32pol in size, sequence, and DNA endonuclease activity. 相似文献
20.
A novel endonuclease from adult hen liver nuclei has been purified to a homogeneous state through salt extraction, ammonium sulfate fractionation, gel filtration, acetone fractionation, and successive chromatography of 1) hydroxyapatite and DNA Sepharose and 2) hydroxyapatite and isoelectric focusing. The endonuclease has a pH optimum at 9.0 and requires Mg2+ for activity. The enzyme hydrolyzes more rapidly in the order of polynucleotide: denatured DNA = rRNA greater than poly(dA) = poly(dT) greater than poly(dC) = poly(dG) greater than native DNA. This endonuclease degrades denatured DNA about 20 times more rapidly than does the native DNA. The products contain 5'-phosphoryl and 3'-hydroxyl termini and all four deoxynucleotides are present while dGMP is predominant. The enzyme cleaves the circular duplex PM2 DNA, endonucleotically, via single strand scission. The isoelectric point is 10.2 +/- 0.2 and the molecular weight is 43,000 +/- 2,000, determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration. Pyridoxal 5'-phosphate and 2,3-butanedione inhibit the catalytic activity, respectively. The inhibition of DNA binding activity was also seen with former, but not with the latter. Purified Mg2+-dependent alkaline endonuclease was used to investigate the nature of poly(ADP-ribose) inhibition of the enzyme. In contrast to the Ca2+/Mg2+-dependent endonuclease (Yoshihara, K., Tanigawa, Y., Burzio, L., and Koide, S. S. (1975) Proc. Natl. Acad. Sci. U. S. A. 72, 289-293), ADP-ribosylation of the endonuclease protein was not observed. When 100 ng of the poly(ADP-ribose) having four to five ADP-ribose units per molecule were added to the nuclease assay system (total volume of 0.2 ml) 14% inhibition was observed, and increase in the chain length increased the inhibition. When 100 ng of poly(ADP-ribose) consisting of 20 or more units of the ADP-ribose per mol were added, the inhibition was over 95%. The possible role of the poly(ADP-ribose)-sensitive endonuclease is discussed. 相似文献