Cleavage Of Supercoiled DNA Research Articles

Synthesis, DNA recognition and cleavage studies of novel tetrapeptide complexes, Cu(II)/Zn(II)-Ala-Pro-Ala-Pro

New tetrapeptide complexes Cu(II)·Ala-Pro-Ala-Pro (1) and Zn(II)·Ala-Pro-Ala-Pro (2) were synthesized from the reaction of tetrapeptide, Ala-Pro-Ala-Pro and CuCl2/ZnCl2 and were thoroughly characterized by elemental analysis, IR,1H and 13C NMR (in case of 2), ESI-MS, UV and molar conductance measurements. The solution stability study was carried out employing UV–vis absorption titrations over a broad range of pH which suggested the stability of the complexes in solution. In vitro interaction of complexes 1 and 2 with CT-DNA was studied employing UV–vis, fluorescence, circular dichroic and viscometry studies. To throw insight into molecular binding event at the target site, UV–vis titrations of 1 and 2 with mononucleotides of interest viz.; 5′-GMP and 5′-TMP were carried out. Cleavage activity of the complexes with pBR322 plasmid DNA was evaluated by agarose gel electrophoresis and, the electrophoresis pattern demonstrated that both the complexes 1 and 2 are efficient cleavage agents. Further, the Cu(II) complex displayed efficient oxidative cleavage of supercoiled DNA while various reactive oxygen species are responsible for the cleavage in Zn(II) complex.

Facile synthesis of a polyether-tethered dimeric berberine as a highly effective DNA-cleaving agent in the presence of Cu(ii) ion

A polyether-tethered dimeric berberine was synthesized and found to be capable of efficiently converting pBR322 DNA into open circular and linear forms in the presence of Cu(II) ions under physiological conditions, most probably via an oxidative mechanism. Michaelis–Menten analysis afforded ca. 3.5 × 109-fold rate acceleration over uncatalyzed cleavage of supercoiled DNA. The probable mode of action is also briefly discussed.

Synthesis, Crystal Structures and DNA-Cleaving Activities of [Cemp]<sub>2</sub>[MCl<sub>4</sub>] (Cemp=<i>N</i>-Carbethoxymethyl-1,10-phenanthrolinium, M=Cu<sup>II</sup>, Zn<sup>II</sup>, Co<sup>II</sup>, Ni<sup>II</sup> and Mn<sup>II</sup>)

N-Carbethoxymethyl-1,10-phenanthrolinium bromide (CempBr) and its five ionic metal complexes, [Cemp]2[MCl4] where M = Cu(II) (1), Zn(II) (2), Co(II) (3), Ni(II) (4) and Mn(II) (5) were synthesized and fully characterized. Complexes 1-5 have similar structures, and consist of isolated [Cemp](+) cations and [MCl4](2-) anions in which there are no obvious interactions between the oxygen or nitrogen donor atoms in [Cemp](+) and the metal center in [MCl4](2-). Agarose gel electrophoresis studies on the cleavage of plasmid pBR322 DNA by complexes 1-5 indicated that complex 1 was capable of efficiently cleaving DNA under physiological conditions, most probably via an oxidative mechanism. Kinetic assay of complex 1 afforded the maximal catalytic rate constant kmax of 0.55 h(-1) and Michaelis constant KM of 47.6 µM, respectively, which gives about 1.5×10(7)-fold rate acceleration over uncatalyzed cleavage of supercoiled DNA. Ethidium bromide displacement experiments indicated that complex 1 had a binding affinity of (1.58±1.12)×10(6) M(-1) toward calf-thymus DNA, 20-100-fold higher than those shown by CempBr and complexes 2-5. The high cleaving efficacy of complex 1 is thought to be due to the efficient catalysis of the copper(II)-coordinated center and the efficient binding of the quaternized 1,10-phenanthroline cation to DNA.

Studies on the nuclease activity and interactions of the mixed‐polypyridyl [Ni2(1,3‐tpbd)(diimine)2(H2O)2]4+ complexes with thioredoxin reductase

Three new nickel(II) complexes formulated as [Ni2(1,3‐tpbd)(diimine)2(H2O)2]4+ [1,3‐tpbd = N,N,N′,N′‐tetrakis(2‐pyridylmethyl)benzene‐1,3‐diamine, where diimine is an N,N‐donor heterocyclic base like 1,10‐phenanthroline (phen),2,2′‐bipyridine (bpy), 4,5‐diazafluoren‐9‐one (dafo)], have been synthesized and structurally characterized by X‐ray crystallography: [Ni2(1,3‐tpbd)(phen)2(H2O)2]4+ (1), [Ni2(1,3‐tpbd)(bpy)2(H2O)2]4+(2) and [Ni2(1,3‐tpbd)(dafo)2(H2O)2]4+ (3). Single‐crystal diffraction reveals that the metal atoms in the complexes are all in a distorted octahedral geometry and in a trans arrangement around 1,3‐tpbd ligand. The interactions of the three complexes with calf thymus DNA (CT‐DNA) have been investigated by UV absorption, fluorescence spectroscopy, circular dichroism and viscosity. The apparent binding constant (Kapp) values are calculated to be 1.91 × 105 m−1 for 1, 1.18 × 105 m−1 for 2, and 1.35 × 105 m−1 for 3, following the order 1 > 3 > 2. The higher DNA binding affinity of 1 is due to the involvement in partial insertion of the phen ring between the DNA base pairs. A decrease in relative viscosities of DNA upon binding to 1–3 is consistent with the DNA binding affinities. These complexes efficiently display oxidative cleavage of supercoiled DNA in the presence of H2O2 (250 µ m), with 3 exhibiting the highest nuclease activity. The rate constants for the conversion of supercoiled to nicked DNA are 5.28 × 10−5 s−1 (for 1), 6.67 × 10−5 s−1 (for 2) and 1.39 × 10−4 s−1 (for 3), also indicating that complex 3 shows higher catalytic activity than 1 and 2. Here the nuclease activity is not readily correlated to binding affinity. The inhibitory effect of complexes 1–3 on thioredoxin reductase has also been examined. The IC50 values are calculated to be 26.54 ± 0.57, 31.03 ± 3.33 and 8.69 ± 2.54 µ m, respectively, showing a more marked inhibitory effect on thioredoxin reductase by complex 3 than the other two complexes. Copyright © 2012 John Wiley & Sons, Ltd.

Two new dicopper(II) complexes with oxamido-bridged ligand: Synthesis, crystal structures, DNA binding/cleavage and BSA binding activity

New oxamido-bridged copper(II) complexes, [Cu2(oxbp)(H2O)2(NCS)2] (1) and [Cu2(oxbp)(μ-DMSO)2(NCS)2][Cu2(oxbp)(DMSO)2(NCS)2] (2) (H2oxbp=N,N′-bis(2-(diethylamino)ethyl)oxalamide), were synthesized and characterized by single-crystal X-ray diffraction, elemental analysis, IR, and electronic spectra. X-ray analysis revealed that complex 1 consists of neutral binuclear [Cu2(oxbp)(H2O)2(NCS)2] units which forms a two-dimensional network through intermolecular hydrogen bonds and complex 2 is constructed by neutral [Cu2(oxbp)(μ-DMSO)2(NCS)2] (2a) and [Cu2(oxbp)(DMSO)2(NCS)2] (2b) entities which alternately distribute to form a two-dimensional network by means of quasi μ-DMSO bridge and intermolecular hydrogen bonds. In both 1 and 2, bicopper centers are linked by the “trans-form” oxamido bridges with the distances of 5.272Å for 1 and av. 5.296Å for 2, respectively. The interaction of Cu(II) complexes with DNA was investigated by UV-visible, fluorescence emission spectrometry and agarose gel electrophoresis. The apparent binding constant (Kapp) values of 3.16×105M−1 for 1 and 4.9×105M−1 for 2 suggest moderate intercalative binding modes between the complexes and DNA. Complex 1 displayed efficient oxidative cleavage of supercoiled DNA, which might indicate that the underlying mechanism involves singlet oxygen (1O2) as reactive oxygen species. Complex 2 is characteristic of the involvement of a singlet oxygen-like entity and hydrogen peroxide in the cleavage process. In addition, our present work showed, by fluorescence spectrometry of BSA with complexes, both 1 and 2 bind to BSA with a medium affinity through a static mode which is tentatively assigned binding to Trp134 in BSA.

DNA binding, nuclease activity and cytotoxicity studies of Cu(ii) complexes of tridentate ligands

Cu(II) complexes of three tridentate ligands, L(1), L(2) and L(3), [L(1), N-((1H-imidazole-2-yl)methyl)-2-(pyridine-2-yl)ethanamine; L(2), N-((1-methyl-1H-imidazole-2-yl)methyl)-2-(pyridine-2-yl)ethanamine; L(3), 2-(pyridine-2-yl)-N-((pyridine-2-yl)methyl)ethanamine] respectively, were synthesized and characterized. The single crystal X-ray structure of complex 1 reveals the pseudo octahedral coordination geometry around the copper center. Absorption and fluorescence experimental evidence show good DNA binding propensity (in the order of 10(5) M(-1)) of the complexes. Thermal denaturation and circular dichroism (CD) analyses reveal minor structural changes of calf thymus (CT) DNA in presence of complexes and groove and/or surface binding of the complexes to CT-DNA. Kinetic DNA cleavage assay shows pseudo-first-order kinetic reaction between the complex and supercoiled (SC) DNA. In addition, mechanistic SC DNA cleavage results show higher DNA cleavage activity in presence of reducing agent, due to the presence of hydroxyl radicals. In vitro cytotoxicity assay of the complexes demonstrate that the complexes have low toxicity for different cancer cell lines and IC(50) values were between 37 and 156 μM.

Effects of Sanionia uncinata extracts in protecting against and inducing DNA cleavage by reactive oxygen species

When mosses are exposed to increased quantities of ultraviolet (UV) radiation, they produce more secondary metabolites. Antarctica moss Sanionia uncinata (Hedw.) Loeske has presented high carotenoid contents in response to an increase in UVB radiation. This moss has been recommended as a potential source of antioxidants. In the present work, the protective and enhancing effects of aqueous (AE) and hydroalcoholic (HE) extracts of S. uncinata on the cleavage of supercoiled DNA were evaluated through topological modifications, quantified by densitometry after agarose gel electrophoresis. Total phenolic contents reached 5.89 mg/g. Our data demonstrated that the extract does not induce DNA cleavage. Furthermore, both extracts showed antioxidant activity that protected the DNA against cleavage induced by (i) O2•−, 89% (AE) and 94% (HE) (P < 0.05), and (ii) .OH, 17% (AE) and 18% (HE). However, the extracts intensified cleavage induced by Fenton-like reactions: (i) Cu2+/H2O2, 94% (AE) and 100% (HE) (P < 0.05), and (ii) SnCl2, 62% (AE) and 56% (HE). DNA damages seem to follow different ways: (i) in the presence of Fenton-like reactions could be via reactive oxygen species generation and (ii) with HE/Cu2+ could have also been triggered by other mechanisms.

DNA binding, oxidative DNA cleavage, cytotoxicity, and apoptosis-inducing activity of copper(II) complexes with 1,4-tpbd ( N,N,N′,N′-tetrakis(2-yridylmethyl)benzene-1,4-diamine) ligand

Three new binuclear copper(II) complexes have been synthesized and structurally characterized by X-ray crystallography, [Cu 2(1,4-tpbd)(dafo) 2(MeOH) 2](ClO 4) 4·2.5H 2O (1), [Cu 2(1,4-tpbd) (DMSO) 2(ClO 4) 2](OH) 2·6H 2O (2) and [Cu 2(1,4-tpbd)(OAC) 2(ClO 4) 2]·5H 2O (3) (1,4-tpbd = N, N, N′, N′-tetrakis(2-pyridylmethyl)benzene-1,4-diamine). Complex 1 to 3 shows similar binuclear structure and each Cu atom adopts five-coordinated square-pyramidal geometry. The interactions of the three complexes with CT-DNA (Calf-thymus DNA) have been investigated by UV absorption, fluorescence spectroscopy, circular dichroism spectroscopy and viscosity. Furthermore, the three complexes display oxidative cleavage of supercoiled DNA in the presence of external agents. Complex 3 shows higher DNA affinity and nuclease activity may be attributed to its cis structural configuration and labile acetate and perchlorate anions. The cleavage mechanisms between the complexes and plasmid DNA are likely to involve singlet oxygen or singlet oxygen-like entity as reactive oxygen species. In addition, in vitro cytotoxicity studies on the Hela cell line show that the IC 50 values of complexes 1– 3 are 14.75, 13.67 and 16.58 μM, respectively. The apoptosis-inducing activity was also assessed by AO/EB (Acridine Orange/Ethidium bromide) staining assay, indicating they have the potential to act as effective metal-based anticancer drugs.

Study on potential antitumor mechanism of a novel Schiff Base copper(II) complex: Synthesis, crystal structure, DNA binding, cytotoxicity and apoptosis induction activity

A new cytotoxic copper(II) complex with Schiff base ligand [CuII(5-Cl-pap)(OAc)(H2O)]·2H2O (1) (5-Cl-pap=N-2-pyridiylmethylidene-2-hydroxy-5-chloro-phenylamine), was synthesized and structurally characterized by X-ray diffraction. Single-crystal analysis revealed that the copper atom shows a 4+1 pyramidal coordination, a water oxygen appears in the apical position, and three of the basal positions are occupied by the NNO tridentate ligand and the fourth by an acetate oxygen. The interaction of Schiff base copper(II) complex 1 with DNA was investigated by UV–visible spectra, fluorescence spectra and agarose gel electrophoresis. The apparent binding constant (Kapp) value of 6.40×105M−1 for 1 with DNA suggests moderate intercalative binding mode. This copper(II) complex displayed efficient oxidative cleavage of supercoiled DNA, which might indicate that the underlying mechanism involve hydroxyl radical, singlet oxygen-like species, and hydrogen peroxide as reactive oxygen species. In addition, our present work showed the antitumor effect of 1 on cell cycle and apoptosis. Flow cytometric analysis revealed that HeLa cells were arrested in the S phase after treatment with 1. Fluorescence microscopic observation indicated that complex 1 can induce apoptosis of HeLa cells, whose process was mediated by intrinsic mitochondrial apoptotic pathway owing to the activation of caspase-9 and caspase-3.

Anaerobic DNA cleavage in red light by dicopper(II) complexes on disulphide bond activation

Binuclear complexes [Cu(µ-RSSR)]2 (1) and [M2(µ-PDS)(H2O)]2 (M = Cu(II), 2; Fe(II), 3), where H2RSSR is a reduced Schiff base derived from 2-(thioethyl)salicylaldimine having a disulphide moiety and H2PDS is derived from dimerization of D-penicillamine, have been prepared, structurally characterized, and their photo-induced DNA cleavage activity studied. The crystal structure of 1 shows the complex as a discrete binuclear species with each metal in a CuN2O2 square-planar geometry (Cu…Cu, 6·420 Å). The tetradentate RSSR2− acts as a bridging ligand. The sulphur atoms in the disulphide unit do not interact with the metal ions. Complexes 1–3 do not show any DNA cleavage activity in darkness. The copper(II) complexes exhibit chemical nuclease activity in the presence of 3-mercaptopropionic acid. Cleavage of supercoiled DNA has been observed in UV-A light of 365 nm for 1 and red light of 647·1 nm for both 1 and 2 in air. Mechanistic data reveal the involvement of the disulphide unit as photosensitizer generating hydroxyl radicals (•OH) as the reactive species. Photo-induced DNA cleavage in red light seems to involve sulphide radicals in a type-I process and hydroxyl radicals. The dicopper(II) complexes show significant anaerobic photo-induced DNA cleavage activity in red light under argon following type-I pathway without involving any reactive oxygen species.

Impact of the Aliphatic Dicarboxylate Chain of Novel Dinuclear Palladium(II) Complexes: Synthesis and Biological Evaluation

AbstractFive dinuclear palladium(II) complexes with HOOC(CH2)nCOOH (n=2–6) and 2,2′‐bipyridine ligands were synthesigned by a reaction with K2PdCl4. To prepare such complexes with different aliphatic dicarboxylate chain lengths was in an attempt to correlate this length factor, which influences the biological activity of the complexes, with fluorescence spectra, DNA cleavage and cytotoxic activity. The results indicate that the complexes bind to fish sperm DNA (FS‐DNA) in an intercalative mode via fluorescence spectra, and the five complexes show different cleavage of supercoiled DNA, and then a cytotoxicity assay of these dinuclear palladium(II) complexes on human tumor cell lines was performed. In most of the cell lines, complex 5 (n=6) and 4 (n=5) showed much higher cytotoxicity than cis‐platin. On the other hand, complex 3 (n=4) was found to be moderately active, and complex 1 (n=2), complex 2 (n=3) were found only marginally cytotoxic. Implications of these findings were discussed from a structure‐activity relationship.

Cyclization kinetics and biological evaluation of an anticancer 1,2-dialkynylimidazole

1,2-Dialkynylimidazoles have been reported to undergo thermal cyclization/rearrangement to diradical and carbene intermediates. Optimization of the synthesis of the 1,2-dialkynylimidazole has provided sufficient material for kinetic and biological studies. The 1,2-dialkynylimidazole is cytotoxic against a wide range of cancer cells and induces apoptosis in A549 cells. Experimentally-determined kinetics of the thermolysis of (E(a) = 30.0 kcal mol(-1)) are in excellent agreement with DFT calculations of the cyclization/rearrangement to diradical and cyclopentapyrazine carbene intermediates (E(a) = 29.7 kcal mol(-1)). Commensurate with the relatively high barrier for cyclization of , no evidence for cleavage of supercoiled DNA under physiological conditions was found; however, under aqueous conditions at 70 degrees C formed a covalent adduct with a model peptide. These studies indicate that if cyclization of is involved in its anticancer activity, the cyclization must be facilitated, perhaps through initial protein binding, which could lead to covalent protein modification.

DNA cleavage activities and mechanism of a novel heterodinuclear macrocyclic complex

Abstract A new unsymmetrical bis-furan pendant-armed macrocyclic heterodinuclear Cu(II)Zn(II) complex, [CuZnL(OAc)](ClO4)·3MeCN·H2O(1) (H2L was derived from the condensation between 1,6-bis(2-furyl)-2,5-bis(2-hydroxy-3-formyl-5-bromobenzyl)-2, 5-diazahexane and 1,3-diaminopropane), has been synthesized and characterized by EDS (energy dispersive spectroscopy) and X-ray crystallography determination. The interaction between the complex and calf thymus (CT) DNA was investigated by UV–vis absorption, viscosity experiment, fluorescent and CD spectroscopy. The mechanism of the cleavage of supercoiled DNA (pBR 322DNA) process was also studied. It is proposed that DNA cleavage promoted by the complex occurrence via oxidative mechanism.

Synthesis and DNA‐Cleavage Activities of Dinuclear Copper(II) Complexes of Urea‐Bridged Macrocyclic Polyamines

Two dinuclear macrocyclic polyamine copper(II) (Cu(II)) complexes, which have two cyclen units linked by urea, were synthesized as DNA-cleavage agents. The structures of these new dinuclear complexes were identified by HR-ESI-MS and IR analyses. The catalytic activities of DNA cleavage of these dinuclear Cu(II) complexes were subsequently studied. The results show that 6a was the better catalyst in the DNA-cleavage process than 6b. The effects of reaction time and concentration of complexes were also investigated. The results indicate, that the Cu(II) complexes could catalyze the cleavage of supercoiled DNA (pUC 19 plasmid DNA; Form I) under physiological conditions to produce selectively nicked DNA (Form II; no Form III was produced) with high yields (nearly 100%) in short time in the absence of reductant or oxidant.

Synthesis, X‐ray Crystal Structures, DNA Binding and Nuclease Activities of Two Novel 1,2,4‐Triazole‐Based CuII Complexes

AbstractTwo novel 1,2,4‐triazole‐based copper(II) complexes have been synthesized and structurally characterized by X‐ray crystallography: [Cu2LCl3]·0.5[CuCl4]·EtOH (1) and [Cu2LBr3]·Br·H2O (2) (L = 3,5‐bis{[bis(2‐methoxyethyl)amino]methyl}‐4H‐1,2,4‐triazol‐4‐amine). Single‐crystal diffraction reveals that the two Cu atoms have a distorted octahedral geometry for 2, but one Cu atom is in a distorted square‐pyramidal environment and the other one is in a distorted octahedral environment for 1 because of the different bridged halogen atoms. The interactions of the two complexes with calf thymus DNA (CT‐DNA) have been investigated by UV absorption, fluorescence spectroscopy, circular dichroism spectroscopy, viscosity measurements and cyclic voltammetry. The apparent binding constant (Kapp) values for complexes 1 and 2 are 2.98 × 106 and 4.05 × 106 M–1. Furthermore, both the compounds display efficient oxidative cleavage of supercoiled DNA in the presence of external agents. The rate constants for the conversions of supercoiled to nicked DNA are 4.38 × 10–4 s–1 (for 1) and 5.26 × 10–4 s–1 (for 2). (© Wiley‐VCH Verlag GmbH &amp; Co. KGaA, 69451 Weinheim, Germany, 2010)

Biochemical and mutagenic analysis of I-CreII reveals distinct but important roles for both the H-N-H and GIY-YIG motifs

Homing endonucleases typically contain one of four conserved catalytic motifs, and other elements that confer tight DNA binding. I-CreII, which catalyzes homing of the Cr.psbA4 intron, is unusual in containing two potential catalytic motifs, H-N-H and GIY-YIG. Previously, we showed that cleavage by I-CreII leaves ends (2-nt 3′ overhangs) that are characteristic of GIY-YIG endonucleases, yet it has a relaxed metal requirement like H-N-H enzymes. Here we show that I-CreII can bind DNA without an added metal ion, and that it binds as a monomer, akin to GIY-YIG enzymes. Moreover, cleavage of supercoiled DNA, and estimates of strand-specific cleavage rates, suggest that I-CreII uses a sequential cleavage mechanism. Alanine substitution of a number of residues in the GIY-YIG motif, however, did not block cleavage activity, although DNA binding was substantially reduced in several variants. Substitution of conserved histidines in the H-N-H motif resulted in variants that did not promote DNA cleavage, but retained high-affinity DNA binding—thus identifying it as the catalytic motif. Unlike the non-specific H-N-H colicins, however; substitution of the conserved asparagine substantially reduced DNA binding (though not the ability to promote cleavage). These results indicate that, in I-CreII, two catalytic motifs have evolved to play important roles in specific DNA binding. The data also indicate that only the H-N-H motif has retained catalytic ability.

Hydrolytic cleavage of DNA by urea-bridged macrocyclic polyamines

Novel bis-cyclen derivative A with urea, mono-cyclen derivative B with urea and amino acid urea C were synthesized as DNA cleavage agents. The structures of these new compounds were identified by MS and 1H NMR spectroscopy. The catalytic activities on DNA cleavage of these compounds were subsequently studied, and results show that A is a much better catalyst in DNA cleavage process than that of B and C. The effects of reaction time and catalyst concentration were also investigated. The results indicate that A can catalyze the cleavage of supercoiled DNA (pUC 19 plasmid DNA) to nicked DNA under physiological conditions with high yields (nearly 100%) via a hydrolytic mechanism.

Synthesis and DNA-cleaving activity of a series of substituted arenediazonium ions

We investigated the reactions of substituted aryl radicals and aryl cations derived from arenediazonium ions and their ability to cause cleavage of supercoiled DNA and their tendency toward free radical or cation formation in the presence and absent of copper (I) chloride. It was found that the substituted arenediazonium salts can cleave supercoiled DNA to the open circular form II DNA and linear form III DNA. Results of methodical studies indicate that both carbon-centered radicals and aryl cations participate in the cleavage pathways.

Arm effects of mononuclear armed cyclen copper complexes on DNA cleavage

Novel cyclen copper(II) complexes appending different side arms were synthesized as DNA cleavage agents. Both the intermediates and mononuclear complexes were characterized by 1H NMR, ESI-HRMS, Elemental analyses and IR, and their catalytic activities for DNA cleavage and DNA binding abilities were investigated. The results indicate that the copper(II) complexes could catalyze the cleavage of supercoiled DNA (pUC 19 plasmid DNA) under physiological conditions to produce nicked DNA with high yields (nearly 100%) via an oxidative mechanism in the absence of exogenous agents; The copper complex bearing an 9-anthryl group gave superior DNA interactions to those bearing phenyl or methyl groups.

Synthesis, DNA‐Binding, Cleavage, and Cytotoxic Activity of New 1,7‐Dioxa‐4,10‐diazacyclododecane Artificial Receptors Containing Bisguanidinoethyl or Diaminoethyl Double Side Arms

Novel 1,7-dioxa-4,10-diazacyclododecane artificial receptors with two pendant aminoethyl (3) or guanidinoethyl (4) side arms have been synthesized. Spectroscopy, including fluorescence and CD spectroscopy, of the interactions of 3, 4, and their copper(II) complexes with calf thymus DNA indicated that the DNA binding affinity of these compounds follows the order Cu(2+)-4>Cu(2+)-3>4>3, and the binding constants of Cu(2+)-3 are Cu(2+)-4 are 7.2x10(4) and 8.7x10(4) M(-1), respectively. Assessment by agarose gel electrophoresis of the plasmid pUC 19 DNA cleavage activity in the presence of the receptors showed that the complexes Cu(2+)-3 and Cu(2+)-4 exhibit powerful supercoiled DNA cleavage efficiency. Kinetic data of DNA cleavage promoted by Cu(2+)-3 and Cu(2+)-4 under physiological conditions fit to a saturation kinetic profile with kmax values of 0.865 and 0.596 h(-1), respectively, which give about 10(8)-fold rate acceleration over uncatalyzed supercoiled DNA. This acceleration is due to efficient cooperative catalysis of the copper(II) center and the functional (diamino or bisguanidinium) groups. In-vitro cytotoxic activities toward murine melanoma B16 cells and human leukemia HL-60 cells were also examined: Cu(2+)-4 shows the highest activity with IC(50) values of 1.62x10(-4) and 1.19x10(-5) M, respectively.