Interaction of the antineoplastic agent mitoxantrone with double-stranded nucleic acids

The binding of mitoxantrone with double-helical nucleic acids was investigated by the methods of isothermal microcalorimetry, circular dichroism and absorption at the ionic strength mu = 0.11 and 0.011 M NaCl at temperature region of 30 divided by 60 degrees C. The investigation shows, that at mu = 0.11 M NaCl mitoxantrone interacts with double-helical nucleic acids in one way only. For such conditions using spectrophotometric titration data Scatchard plots for the binding of mitoxantrone with double-helical nucleic acids were constructed. The calculations show that the saturation stoichiometry is one mitoxantrone molecule per 2 divided by 3 base pairs DNA and 6 divided by 8 base pairs RNA. The dependence of binding constant from GC-content is observed. It is shown that the binding enthalpy of mitoxantrone with DNA and RNA increases linearly and reaches -(3.0 +/- 0.5) kkal per 1 mol mitoxantrone. It is shown that a binding mitoxantrone with double-helical nucleic acids, besides the intercalation of rings, a determinate contribution in the binding is given also by electrostatic interaction of side chains mitoxantrone with nucleic acids.     

DNA interaction studies of an antiviral drug, ribavirin, using different instrumental methods

Interaction of ribavirin with CT-DNA was investigated by emission, absorption, circular dichroism, and viscosity studies to determine the binding mode and binding constant of this drug with DNA. The calculated binding constant, K(b), obtained from UV-vis absorption studies was 4.6 × 10(3) M(-1). In fluorimetric studies, the enthalpy (ΔH<0) and entropy (ΔS>0) of the reaction between ribavirin and CT-DNA showed a hydrophobic interaction. In addition, in the circular dichroism spectrum, the drug induces a B → A structural transition of CT-DNA. These results demonstrate that ribavirin interacts with CT-DNA via the groove binding mode. It was observed that the drug has ability to cleave supercoiled plasmid DNA.     

6-Nitro-L-tryptophan: a novel spectroscopic probe of trp aporepressor and human serum albumin

The binding of 6-nitro-L-tryptophan to trp aporepressor and human serum albumin has been examined by visible difference spectroscopy and circular dichroism. 6-Nitro-L-tryptophan, prepared by nitration of L-tryptophan with nitric acid in glacial acetic acid, exhibits a visible and near-uv absorption spectrum with lambda max at about 330 nm (epsilon = 7 X 10(3) M-1 cm-1) and a shoulder near 380 nm in H2O. In the presence of trp aporepressor, the visible absorption intensity is sharply diminished. Visible difference spectral titration data give KD = 1.27 X 10(-4) M and n = 0.95 per subunit at 25 degrees C. While 6-nitro-L-tryptophan exhibits no significant circular dichroism between 300 and 500 nm, the complex with trp aporepressor exhibits strong circular dichroism signals, with a negative maximum at 386 nm (delta epsilon = -7.5 M-1 cm-1) and a positive maximum at 310 nm (delta epsilon = +6 M-1 cm-1). Circular dichroism titration data give KD = 1.69 X 10(-4) M and n = 0.90 per subunit at 25 degrees C. The KD values determined spectroscopically are in excellent agreement with that determined by equilibrium dialysis, KD = 1.5 X 10(-4) M at 25 degrees C. In the presence of human serum albumin, the spectrum of 6-nitro-L-tryptophan exhibits a blue shift and an increase in absorption intensity; similar changes are observed in solvents of low dielectric contrast such as 80% aqueous dioxane. Visible difference spectral titration data give KD = 8.0 X 10(-5) M and n = 0.95 for human serum albumin. The complex of 6-nitro-L-tryptophan with human serum albumin exhibits a strong positive circular dichroism maximum at 380 nm (delta epsilon = +9.8 M-1 cm-1) with a shoulder at 310-320 nm. Circular dichroism titration data give KD = 6.4 X 10(-5) M and n = 0.83, in good agreement with the visible difference spectral results. Taken together, our results demonstrate the utility of 6-nitro-L-tryptophan as a spectroscopic probe for tryptophan-binding proteins.     

Synthesis, characterization, DNA-binding and spectral properties of complexes [Ru(L)4(dppz)]2+ (L=Im and MeIm)

Two new Ru(II) complexes [Ru(L)(4)(dppz)](2+) (L=imidazole (Im), 1-methylimidazole (MeIm); dppz=dipyrido[3,2-a:2',3'-c]phenazine), have been synthesized and characterized in detail by elemental analysis, (1)H NMR, Electrospray ionization mass spectrometry (ESI-MS) and UV-visible (UV-Vis) spectroscopic techniques. The interaction of these complexes with calf thymus DNA (CT-DNA) has been explored by using electronic absorption titration, competitive binding experiment, circular dichroism (CD), thermal denaturation and viscosity measurements. The experimental results show that: both the two complexes can bind to DNA in an intercalation mode; the DNA-binding affinity of complex [Ru(Im)(4)(dppz)](2+)1 (K(b)=2.5 x 10(6)M(-1)) is greater than that of complex [Ru(MeIm)(4)(dppz)](2+)2 (K(b)=1.1 x 10(6)M(-1)). Moreover, it is very interesting to find that the circular dichroic spectrum of DNA-complex 1 adduct, in which both bands centered at 277 nm and 236 nm are all negative, is very different from those of DNA-complex 2 adduct and other Ru(II) complexes binding to DNA in general intercalation mode. It may be due to the hydrogen-bonding effect or the contribution of induced CD signals of complex 1. Another interesting finding is that the hypochromism of the complexes is not linear relation to their DNA-binding affinities. In order to deeply study these experimental phenomena and trends, the density functional theory (DFT) and time-dependent DFT (TDDFT) computations were carried out, and on the basis of the DFT/TDDFT results and the frontier molecular orbital theory, the trend in DNA-binding affinities, the spectral properties as well as the interesting phenomena of larger extent of hypochromism but relatively smaller K(b) values for the title complexes have been reasonably explained.     

Specific binding and self-structure induction to poly(A) by the cytotoxic plant alkaloid sanguinarine

The cytotoxic plant alkaloid sanguinarine was found to bind preferentially and strongly to single stranded poly(A) with an association constant (K(a)) in the range 3.6-4.6 x 10(6) M(-1) in comparison to several nucleic acids. The binding induced unique self-structure formation in poly(A) that showed cooperative melting transition in circular dichroism, absorbance, and differential scanning calorimetry studies. The alkaloid binding was characterized to be intercalation as revealed from fluorescence quenching experiments and was predominantly enthalpy driven as revealed from isothermal titration calorimetry. Sanguinarine is the first and only natural product so far known to induce a self-structure formation in poly(A).     

Structural analysis of DNA interactions with biogenic polyamines and cobalt(III)hexamine studied by Fourier transform infrared and capillary electrophoresis

Biogenic polyamines, such as putrescine, spermidine, and spermine are small organic polycations involved in numerous diverse biological processes. These compounds play an important role in nucleic acid function due to their binding to DNA and RNA. It has been shown that biogenic polyamines cause DNA condensation and aggregation similar to that of inorganic cobalt(III)hexamine cation, which has the ability to induce DNA conformational changes. However, the nature of the polyamine.DNA binding at the molecular level is not clearly established and is the subject of much controversy. In the present study the effects of spermine, spermidine, putrescine, and cobalt(III)hexamine on the solution structure of calf-thymus DNA were investigated using affinity capillary electrophoresis, Fourier transform infrared, and circular dichroism spectroscopic methods. At low polycation concentrations, putrescine binds preferentially through the minor and major grooves of double strand DNA, whereas spermine, spermidine, and cobalt(III)hexamine bind to the major groove. At high polycation concentrations, putrescine interaction with the bases is weak, whereas strong base binding occurred for spermidine in the major and minor grooves of DNA duplex. However, major groove binding is preferred by spermine and cobalt(III)hexamine cations. Electrostatic attractions between polycation and the backbone phosphate group were also observed. No major alterations of B-DNA were observed for biogenic polyamines, whereas cobalt(III)hexamine induced a partial B --> A transition. DNA condensation was also observed for cobalt(III)hexamine cation, whereas organic polyamines induced duplex stabilization. The binding constants calculated for biogenic polyamines are K(Spm) = 2.3 x 10(5) M(-1), K(Spd) = 1.4 x 10(5) M(-1), and K(Put) = 1.02 x 10(5) M(-1). Two binding constants have been found for cobalt(III)hexamine with K(1) = 1.8 x 10(5) M(-1) and K(2) = 9.2 x 10(4) M(-1). The Hill coefficients indicate a positive cooperativity binding for biogenic polyamines and a negative cooperativity for cobalt(III)hexamine.     

Study of DNA interactions with steroidal and nonsteroidal estrogen-platinum (II)-based anticancer drugs

The interactions of the steroidal and nonsteroidal estrogen-platinum (Pt) (II)-based anticancer drugs 16beta-hydroxymethyl-16alpha-[8-(2-pyridin-2-yl-ethylamino)-3,6-dioxaoctyl]-1,3,5(10)-estratrien-3,17betadiol dichloroplatinum (II) (JPM-39), 4-[6-(2'-pyridylethylamino)-butyloxy)-phenyl]-7-methoxy-2,2-dimethyl-3-phenyl-chroman dichloroplatinum (II) (ATG-99), and 1-[(2-aminoethyl)amino]-9,10,10-tris(4-hydroxyphenyl)-9-decene dichloroplatinum (II) (GEB-28) with calf-thymus DNA in vitro using constant DNA concentration and various drug levels were studied. Fourier transform infrared (FTIR) and circular dichroism (CD) were studied with calf-thymus DNA in vitro using constant DNA concentration and various drug levels. FTIR, UV-visible, and CD spectroscopic methods were used to characterize the drug binding mode, the binding constant, and structural variations of DNA in aqueous solution. Spectroscopic evidence showed that the various Pt-based drugs bind indirectly to the major and minor grooves of DNA duplex with some degree of drug-phosphate interaction. The overall binding constants for JPM-39, GEB-28, and ATG-99 are K(JPM-39) = 4.2 (+/-0.75) x 10(3) M(-1), K(GEB-28) = 3.4 (+/-0.65) x 10(3) M(-1), and K(ATG-99) = 2.1 (+/-0.45) x 10(3) M(-1). DNA aggregation occurs at high drug concentration, while DNA remains in the B-family structure.     

Footprinting and circular dichroism studies on paromomycin binding to the packaging region of human immunodeficiency virus type-1

We have studied the interaction of the aminoglycoside drug, paromomycin, with a 171-mer from the packaging region of HIV-1 (psi-RNA), using quantitative footprinting and circular dichroism spectroscopy. The footprinting autoradiographic data were obtained by cutting end-labeled RNA with RNase I or RNase T1 in the presence of varying paromomycin concentrations. Scanning the autoradiograms produced footprinting plots showing cleavage intensities for specific sites on the psi-RNA as functions of drug concentration. Footprinting plots showing binding were analyzed using a two-state model to give apparent binding constants for specific sites of the psi-RNA. These plots show that the highest-affinity paromomycin binding site involves nucleotides near bulges in the main stem and SL-1, and other nucleotides in SL-4 of the psi-RNA. RNase I gives an apparent value of K for this drug site of approximately 1.7 x 10(5) M(-1) while RNase T1 reports a value of K of approximately 8 x 10(4) M(-1) (10 mM Tris HCl, pH 7). Footprinting shows that loading the highest affinity site with paromomycin causes structural changes in the single-stranded linker regions, between the stem-loops and main stem and the loops of SL-1 and SL-3. Drug-induced structural changes also affect the intensity of the 208 nm band in the circular dichroism spectrum of the psi-RNA. Fitting the changes in CD band intensity to a two-state model yielded a binding constant for the highest-affinity drug site of 6 x 10(6) M(-1). Thus, the binding constants from footprinting are lower than those obtained for the highest-affinity site from the circular dichroism spectrum, and lower than those earlier obtained using absorption spectroscopy (Sullivan, J. M.; Goodisman, J.; Dabrowiak, J. C., Bioorg. Med. Chem. Lett. 2002, 12, 615). The discrepancy may be due to competitive binding between drug and cleavage agent in the footprinting experiments, but other explanations are discussed. In addition to revealing sites of binding and regions of drug-induced structural change, footprinting showed that the loop regions of SL-1, SL-3 and SL-4 are exposed in the RNA, whereas the linker region between SL-1 and SL-2 is 'buried' and not accessible to cutting by RNase I or RNase T1.     

DNA interaction studies of a copper (II) complex containing an antiviral drug, valacyclovir: the effect of metal center on the mode of binding

The water-soluble complex, [Cu(Val)(2)(NO(3))(2)]; in which Val = valacyclovir, an antiviral drug, has been synthesized and characterized by elemental analysis, furier transfer-infrared, hydrogen nuclear magnetic resonance (H NMR), and UV-Vis techniques. The binding of this Cu (II) complex to calf thymus DNA was investigated using fluorimetry, spectrophotometry, circular dichroism, and viscosimetry. In fluorimetric studies, the enthalpy and entropy of the reaction between the complex and calf-thymus DNA (CT-DNA) showed that the reaction is endothermic (ΔH = 208.22 kJ mol(-1); ΔS = 851.35 J mol(-1)K(-1)). The complex showed the absorption hyperchromism in its ultra violet-visible (UV-Vis) spectrum with DNA. The calculated binding constant, K(b), obtained from UV-Vis absorption studies was 2 × 10(5) M(-1). Moreover, the complex induced detectable changes in the circular dichroism spectrum of CT-DNA, as well as changes in its viscosity. The results suggest that this copper (II) complex interacts with CT-DNA via a groove-binding mode.     

Interaction of antioxidant flavonoids with tRNA: intercalation or external binding and comparison with flavonoid-DNA adducts

Antioxidants are essential to good health. Flavonoids are powerful antioxidants, and prevent DNA damage. The antioxidative protections are related to their binding modes to a DNA duplex and complexation with free radicals in vivo. Recently we reported the interaction of flavonoids with DNA in vitro (Kanakis et al., J. Biomol. Struct. Dyn. 22, 719-724, 2005), where polyphenol different binding modes were discussed. The aim of this study was to examine the interaction of transfer RNA with quercetin (que), kaempferol (kae), and delphinidin (del) in aqueous solution at physiological conditions and to make a comparison with the corresponding pigment-DNA adducts. Constant tRNA concentration (6.25 mM) and various drug/RNA(phosphate) molar ratios of 1/48 to 1/8 were used. FTIR and UV-visible difference spectroscopic methods have been applied to determine the drug binding mode, the binding constants, and the effects of drug complexation on the stability and conformation of tRNA duplex. Both intercalative and external binding modes were observed. Structural analysis showed que, kae, and a del intercalate tRNA duplex with minor external binding to the major or minor groove and the backbone phosphate group with overall binding constants K (que) = 4.80 x 10(4) M(1), K (kae) = 4.65 x 10(4) M(1), and K (del) = 9.47 x 10(4) M(1). The stability of adduct formation is in the order of del > que > kae. A comparison with flavonoids-DNA adducts showed both intercalation and external bindings with the stability order K (que) = 7.25 x 10(4) M(1), K (kae) = 3.60 x 10(4) M(1), and K (del) = 1.66 x 10(4) M(1). Low flavonoid concentration induces helical stabilization, whereas high pigment content causes helix opening. A partial Bto A-DNA transition occurs at high drug concentration, while tRNA remains in the A-family structure.     

Acenaphtho[1,2-b]pyrrole derivatives as new family of intercalators: various DNA binding geometry and interesting antitumor capacity

A series of acenaphtho[1,2-b]pyrrole derivatives were synthesized and their intercalation geometries with DNA and antitumor activities were investigated in detail. From combination of SYBR Green-DNA melt curve, fluorescence titration, absorption titration, and circular dichroism (CD) studies, it was identified that to different extent, all the compounds behaved as DNA intercalators and transformed B form DNA to A-like conformation. The different intercalation modes for the compounds were revealed. The compounds containing a methylpiperazine substitution (series I) intercalated in a fashion that the long axis of the molecule paralleled to the base-pair long axis, while the alkylamine- substituted compounds (series II and III) located vertically to the long axis of DNA base pairs. Consequently, the DNA binding affinity of these compounds was obtained with the order of II>III>I, which attributed to the role of the substitution in binding geometry. Further, cell-based studies showed all the compounds exhibited outstanding antitumor activities against two human tumor cell lines with IC(50) ranging from 10(-7) to 10(-6)M. Interestingly, compound (1)a (a compound in series I), whose binding affinity was one of the lowest but altered DNA conformation most significantly, showed much lower IC(50) value than other compounds. Moreover, it could induce tumor cells apoptosis, while the compounds (2)a and (3)a (in series II and III, respectively) could only necrotize tumor cells. Their different mechanism of killing tumor cells might lie in their different DNA binding geometry. It could be concluded that the geometry of intercalator-DNA complex contributed much more to the antitumor property than binding affinity.     

Noncovalent DNA binding of bis(1,10-phenanthroline)copper(I) and related compounds

The noncovalent DNA binding of the bis(1,10-phenanthroline)copper(I) complex [(Phen)2CuI] was examined under anaerobic conditions by absorption and circular dichroism spectroscopy, and viscometry, as a function of phenanthroline concentration. Analyses according to the McGhee-von Hippel method indicated that binding exhibited both neighbor-exclusion and positive cooperativity effects, with a neighbor-exclusion parameter n approximately 2 and a cooperativity parameter omega approximately 4. The association constant for (Phen)2CuI binding decreased with increasing concentration of phenanthroline in excess over that required to stoichiometrically generate (Phen)2CuI, indicating that free phenanthroline was a weak competitive inhibitor of (Phen)2CuI binding. The maximal association constant for DNA binding of (Phen)2CuI in 0.2 M NaCl and 9.8% ethanol, extrapolated to zero concentration of excess phenanthroline, was 4.7 x 10(4) M-1 (DNA base pairs). The magnitude of the neighbor-exclusion parameter, the changes in spectral properties of (Phen)2CuI induced by DNA binding, and the increase in DNA solution viscosity upon (Phen)2CuI addition are consistent with a model for DNA binding by (Phen)2CuI involving partial intercalation of one phenanthroline ring of the complex between DNA base pairs in the minor groove as suggested previously [Veal & Rill (1989) Biochemistry 28, 3243-3250]. Viscosity measurements indicated that the mono(phenanthroline)copper(I) complex also binds to DNA by intercalation; however, no spectroscopic or viscometric evidence was found for DNA binding of free phenanthroline or the bis(2,9-dimethyl-1,10-phenanthroline)copper(I) complex. DNA binding of free phenanthroline may be cooperative and induced by prior binding of (Phen)2CuI.     

Biophysical studies of the c-MYC NHE III1 promoter: model quadruplex interactions with a cationic porphyrin

Regulation of the structural equilibrium of G-quadruplex-forming sequences located in the promoter regions of oncogenes by the binding of small molecules has shown potential as a new avenue for cancer chemotherapy. In this study, microcalorimetry (isothermal titration calorimetry and differential scanning calorimetry), electronic spectroscopy (ultraviolet-visible and circular dichroism), and molecular modeling were used to probe the complex interactions between a cationic porphryin mesotetra (N-methyl-4-pyridyl) porphine (TMPyP4) and the c-MYC PU 27-mer quadruplex. The stoichiometry at saturation is 4:1 mol of TMPyP4/c-MYC PU 27-mer G-quadruplex as determined by isothermal titration calorimetry, circular dichroism, and ultraviolet-visible spectroscopy. The four independent TMPyP4 binding sites fall into one of two modes. The two binding modes are different with respect to affinity, enthalpy change, and entropy change for formation of the 1:1 and 2:1, or 3:1 and 4:1 complexes. Binding of TMPyP4, at or near physiologic ionic strength ([K(+)] = 0.13 M), is described by a "two-independent-sites model." The two highest-affinity sites exhibit a K(1) of 1.6 x 10(7) M(-1) and the two lowest-affinity sites exhibit a K(2) of 4.2 x 10(5) M(-1). Dissection of the free-energy change into the enthalpy- and entropy-change contributions for the two modes is consistent with both "intercalative" and "exterior" binding mechanisms. An additional complexity is that there may be as many as six possible conformational quadruplex isomers based on the sequence. Differential scanning calorimetry experiments demonstrated two distinct melting events (T(m)1 = 74.7 degrees C and T(m)2 = 91.2 degrees C) resulting from a mixture of at least two conformers for the c-MYC PU 27-mer in solution.     

Interaction of DNA with actinocin amides containing cationoid centers in the amide groups

Complexes of DNA with actinocin derivatives containing ω-dialkylaminoalkyl groups in the1 and/or 9 positions of the chromophore were studied by spectrophotometric titration, circular dichroism, and viscometry. Induced circular dichroism (ICD) spectra of the DNA-ligand complexes were compared for the cases of the complexes of known structure established by other methods. It was shown that the presence of an isoelliptic point in the long-wavelength absorption band of the ICD spectra of the ligand under monomeric binding conditions could indicate intercalation of the actinocin chromophore into DNA. The separation of the cationoid center and the chromosphore upon elongation of the methylene chain increases the aggregability of the ligand pn the surface of the DNA double helix, which prevents the intercalation of the chromophore.     

Design and characterization of asparagine- and lysine-containing alanine-based helical peptides that bind selectively to A.T base pairs of oligonucleotides immobilized on a 27 mhz quartz crystal microbalance

We have systematically designed and synthesized six kinds of 16-17 mer alanine-based peptides containing four to six lysine (K) and one to four asparagine (N) residues to achieve the selective binding to A.T base pairs of DNA duplexes. The position and number of K and N residues were changed in the helical structure according to common features of the DNA-binding proteins, in which K and N residues are expected to interact electrostatically with phosphate groups and to interact with A.T base pairs by hydrogen bonding, respectively. The time courses of binding of these peptides to dA(30).dT(30) and dG(30).dC(30) duplexes immobilized on a 27 MHz quartz crystal microbalance (QCM) were studied in 10 mM phosphate buffer (pH 7.5) and 40 mM NaCl at 10 degrees C. The maximum binding amounts (Deltam(max)) on a nanogram scale and binding constants (K(a)) could be obtained from the frequency decrease (mass increase) of the oligonucleotide-immobilized QCM. The conformation changes of the peptides upon binding to DNAs were monitored by circular dichroism (CD) spectroscopy. The four properly arranged N residues in the six-cationic K peptide, K6N4(d), resulted in a 5-fold higher affinity for A.T base pairs (K(a) = 5.9 x 10(5) M(-1)) than for G.C base pairs (K(a) = 1.2 x 10(5) M(-1)), and alpha-helices were clearly promoted by the binding to A.T base pairs from CD spectral changes.     

DNA interaction studies of a platinum(II) complex containing L-histidine and 1,10-phenanthroline ligands

The aim of this study was developing coordination complexes that can be used as inorganic medicinal agents. The water soluble [Pt(phen)(His)]NO(3)·3H(2)O complex in which phen=1,10-phenantheroline and His=L-histidine was synthesized and characterized using physicochemical methods. Binding interaction of this complex with calf thymus (CT) DNA was investigated by emission, absorption, circular dichroism, and viscosity measurement techniques. Upon addition of CT-DNA, changes were observed in the characteristic ultraviolet-visible (UV-Vis) bands (hypochromism) of the complex. The complex binds to CT-DNA in an intercalative mode. The calculated binding constant, K(b), was 8 ± 0.2 × 10(4) M(-1). In addition, circular dichroism (CD) study showed that the phenanthroline ligand was inserted between the base pair stack of the double-helical structure of DNA. Also, the fluorescence spectral characteristics showed an increase in fluorescence intensity of the platinum complex in the presence of increasing amounts of DNA solution. The experimental results showed that the platinum complex binds to DNA via intercalative and hydrogen bonding mode.     

Molecular recognition between oligopeptides and nucleic acids. DNA sequence specificity and binding properties of an acridine-linked netropsin hybrid ligand

The binding to DNA of a mixed function ligand (NETGA) is described, in which a potential intercalating group, an acridine moiety, is incorporated at the carboxyl terminus of the minor groove binding oligopeptide netropsin skeleton. Scatchard analysis of absorption data provided evidence of two modes of binding to DNA with K1 = 9.1 x 10(5) M-1 at low r values (0.003-0.1), and a binding site size n = 10, indicative of binding of both moeities. At high binding ratios (greater than 0.1), K2 = 0.9 x 10(5) M-1 and n = 5 corresponding to external binding. Complementary strand MPE footprinting on a pBR322 restriction fragment showed NETGA binds to 5'-AAAT like netropsin. It causes enhanced cleavage by MPE, particularly at G-C rich sequences and remote from the preferred binding sites. Viscometry measurements provided evidence for biphasic modes of the two binding portions of NETGA. Fluorescence polarization and linear dichroism measurements were in accord with distinct modes of interaction of the acridine (intercalation) and oligopeptide (minor groove binding) portions of NETGA. LD measurements on NETGA indicate that the oligopeptide moiety (netropsin-like) has an orientation typical of minor groove binders, whereas the degree of intercalation of the acridine group is decreased by association of the oligopeptide moiety.     

Binding and photocleavage of cationic porphyrin-phenylpiperazine hybrids to DNA

The binding properties of cationic porphyrin-phenylpiperazine hybrids to calf thymus (CT) DNA were investigated by using absorption, fluorescence and circular dichroism (CD) spectra, and the apparent affinity binding constants (K(app)) of the porphyrins for CT DNA were determined by using a competition method with ethidium bromide (EB). Intercalation of porphyrin into CT DNA occurred when two phenylpiperazines were introduced at cis position onto the periphery of cationic porphyrin. The photocleavages of pBR322 plasmid DNA by the porphyrins were consistent with the values of K(app). With [porphyrin]/[DNA base pairs] ratio increased, the binding mode tended to be outside binding, and the cleavage abilities of the porphyrins varied. In the presence of sodium azide, a quencher of 1O2, the cleavage of DNA by the porphyrin of intercalation was less inhibited.     

Interaction of the antitumour alkaloid coralyne with duplex deoxyribonucleic acid structures: spectroscopic and viscometric studies.

The interaction of coralyne, an antitumour alkaloid with natural and synthetic duplex DNAs was investigated under conditions where the drug existed fully as a true monomer for the first time using spectrophotometric, spectrofluorimetric, circular dichroic and viscometric techniques. The absorption spectrum of coralyne monomer showed hypochromic and bathochromic effects on binding to duplex DNAs. This effect was used to determine the binding parameters of coralyne. The binding constants for four natural DNAs and four synthetic polynucleotides obtained from spectrophotometric titration, according to an excluded site model, using McGhee-von Hippel analysis, were all in the range of (0.38-9.8) x 10(5) M-1, and showed a relatively high specificity for the GC rich ML DNA and the alternating GC polynucleotide. The binding of coralyne decreased with increasing ionic strength, indicating that the binding affinity has a strong electrostatic component. Coralyne stabilized all the DNAs against thermal strand separation. The intense steady state fluorescence of coralyne was effectively quenched on binding to DNAs and the quantitative data on the Stern-Volmer quenching constant obtained was sequence dependent, being maximum with the GC rich DNA and alternating GC polymer. Circular dichriosm studies further evidenced for a strong perturbation of the B-conformation of DNAs consequent to coralyne binding with the concomitant development of extrinsic circular dichroic bands for the bound drug molecules suggesting their strong intercalated geometry in duplex DNAs. Further tests of intercalation using viscosity measurements on linear and covalently closed plasmid DNA conclusively proved the strong intercalation of coralyne in duplex DNA. Binding of the closely related natural alkaloid, berberine under these conditions showed considerably lower affinity to duplex DNAs in all experiments. Taken together, these results suggest that coralyne binds strongly to duplex DNAs by a mechanism of intercalation with specificity towards alternating GC duplex structure.