Base-specific and enantioselective studies for the DNA binding of iron(II) mixed-ligand complexes containing 1,10-phenanthroline and dipyrido[3,2-a:2',3'-c]phenazine

Base specificity and enantioselectivity for the DNA binding of [Fe(phen)2(dppz)]2+ (phen=1,10-phenanthroline and dppz=dipyrido[3,2-a:2',3'-c]phenazine) have been studied by determining the equilibrium binding constant (Kb) of the iron(II) complex to calf thymus DNA (ct-DNA), poly[(dA-dT)2], poly[(dG-dC)2] and poly[(dI-dC)2] using spectrophotometric titration and by monitoring the CD spectral profile of the iron(II) complex in the presence and absence of different types of DNA using circular dichroism (CD) spectroscopy, respectively. It has been shown that [Fe(phen)2(dppz)]2+ prefers to intercalate into the A-T and I-C sequences of poly[(dA-dT)2] and poly[(dI-dC)2] rather than into the G-C sequences of poly[(dG-dC)2] or into the base pairs of ct-DNA. In contrast to previous reports, it is a surprising observation that the enantioselectivity of the DNA binding for [Fe(phen)2(dppz)]2+ is base-dependent in nature. The Delta-enantiomer of [Fe(phen)2(dppz)]2+ is preferentially intercalated into the base pairs of poly[(dG-dC)2] or ct-DNA as indicated by its CD spectral profiles. On the other hand, the Lambda-enantiomer of [Fe(phen)2(dppz)]2+ is favorably intercalated into poly[(dA-dT)2] or poly[(dI-dC)2] as suggested by the opposite CD spectral profile. This preferential binding of Lambda-[Fe(phen)2(dppz)]2+)for the A-T sequence may be attributed to the fact that the binding site for the A-T sequence is relatively facile and thus the steric effect caused by the ancillary (non-intercalated) phen ligands is alleviated. The degree of enantioselectivity represented by inversion constants (Kinv) decreases as the salt concentration in the solution increases, indicating that electrostatic interaction is also operating in the ct-DNA-binding events of the iron (II) complex.     

Base-specific Photocleavage of DNA Induced by Pazelliptine Sensitization: Study of the Mechanism by Time-resolved Absorption and Fluorescence

The intercalating antitumoral drug pazelliptine (PZE) is able to photosensitize the formation of single- and double-strand breaks in supercoiled plasmid DNA and selective photocleavage at guanine residues is observed. In order to understand the mechanisms of DNA cleavage mediated by the photoexcited drug, singlet and triplet excited-state processes in PZE complexed with poly(dA-dT)-poly(dA-dT), poly(dG-dC)-poly(dG-dC) and calf thymus DNA have been investigated by means of single photon counting fluorescence decay and transient absorption techniques. For each complex, three different binding sites have been identified, due to the existence of different geometric structures of the drug in the ground state. For one type of binding site, a proton transfer reaction occurs in the singlet excited state whatever the nucleic acid environment. In contrast, the relaxation dynamics for the other two sites are found to depend widely upon the type of polynucleotide in which the drug has been intercalated. From the results of this study, we suggest that the photodynamic action of PZE does not originate from excitation of the drug in the environment of G-C base pairs but is initiated from its triplet state that reacts by electron transfer with the adenine bases. The specificity of cleavage could be the result of subsequent reactions leading to guanine oxidation.     

Spectroscopic study of the interaction of pazelliptine with nucleic acids

The antitumor drug pazelliptine (PZE) binds to natural and synthetic DNA sequences at 100 mM NaCl, pH 7.0, as deduced from the absorption and fluorescence data. Scatchard plots constructed from the results obtained with poly(dG-dC)-poly(dG-dC) give binding constants of base pairs in the range (2–6) × 10⁵ M⁻¹. The modifications in the absorption and fluorescence spectra observed when PZE binds to various polynucleotides, namely poly(dA-dT)-poly(dA-dT), poly(dA)-poly(dT), poly(dG-dC)-poly(dG-dC) and calf thymus DNA. reveal a change in the protonation state of the drug upon binding, increasing the apparent pK_a of its 9-N nitrogen atom. The PZE excited state properties serve as a sensitive probe to distinguish between homo and hetero A-T sites as well as between AT and GC sites. Fluorescence studies reveal that energy transfer occurs from polynucleotide bases to the bound PZE chromophore, a result consistent with an intercalative mode of binding of the drug to DNA. The emission is enhanced when PZE is bound to A-T base pairs ( 30% increase of φ_F) whereas it is quenched in the vicinity of G-C base pairs ( 90% decrease of φ_F). Furthermore, the fluorescence spectrum obtained with calf thymus DNA is hardly distinguishable from that obtained with poly(dG-dC)-polu(dG-dC), suggesting a binding of PZE to G-C rich regions.     

Circular dichroism spectroscopic studies reveal pH dependent binding of curcumin in the minor groove of natural and synthetic nucleic acids

For the first time, an interaction between the non-toxic, cancer chemopreventive agent curcumin and both natural and synthetic DNA duplexes has been demonstrated by using circular dichroism (CD) and absorption spectroscopy techniques. Upon addition of curcumin to calf thymus DNA, poly(dG-dC).poly(dG-dC) and poly(dA-dT).poly(dA-dT) solutions, an intense positive induced CD band centered around 460-470 nm was observed depending on the actual pH and Na+ ion concentration of the medium; no CD signal was obtained, however, with single stranded poly(dC). Interaction of curcumin with calf thymus DNA was observed already at pH 6.5 in contrast with poly(dG-dC).poly(dG-dC) which induces no extrinsic Cotton effect above a pH value of 5. The protonated, Hoogsteen base-paired structure of poly(dG-dC).poly(dG-dC) is necessary for curcumin binding while the alternating AT-rich polymer formed complexes with curcumin only at certain Na+ concentrations. Evaluation of the spectral data and molecular modeling calculations suggested that curcumin, this dietary polyphenolic compound binds in the minor groove of the double helix. The mechanism of the induced CD activity, the effects of the pH and Na+ ions on the ligand binding and conformation of the double helix are discussed in detail. As well as being an essentially new phenolic minor groove binder agent curcumin is also a promising molecular probe to study biologically important, pH and cation induced conformational polymorphisms of nucleic acids.     

Calorimetric and thermal analysis studies on the binding of phenothiazinium dye thionine with DNA polynucleotides

Binding of the phenothaizinium dye thionine with four sequence specific deoxyribopolynucleotides, poly(dG-dC).poly(dG-dC), poly(dG).poly(dC), poly(dA-dT).poly(dA-dT), and poly(dA).poly(dT) has been investigated by means of thermal helix melting, isothermal titration calorimetry, and differential scanning calorimetry experiments. The binding affinity values evaluated from isothermal titration calorimetry suggests that thionine exhibits the highest binding affinity to poly(dG-dC).poly(dG-dC). The binding to poly(dG-dC).poly(dG-dC), poly(dA-dT).poly(dA-dT), and poly(dG).poly(dC) is exothermic and favoured by negative enthalpy changes while binding to poly(dA).poly(dT) is endothermic and anomalous. The values of heat capacity changes of the interaction are negative and in the range (?0.4 to ?0.5) kJ · K^?1 · mol^?1. The binding is characterized by strong stabilization of the polynucleotides against thermal strand separation. The binding affinity values derived from thermal melting data are in excellent agreement with those obtained from isothermal titration calorimetry data. Insights into the energetic aspects and guanine–cytosine selectivity of the DNA interaction of thionine have been obtained from these studies.     

Thermodynamic profiles of the DNA binding of benzophenanthridines sanguinarine and ethidium: A comparative study with sequence specific polynucleotides

Energetics of the binding of two known classical DNA intercalating molecules, ethidium and sanguinarine with four sequence specific polynucleotides, poly(dG-dC).poly(dG-dC), poly(dG).poly(dC), poly(dA-dT).poly(dA-dT), and poly(dA).poly(dT) have been compared under identical conditions. The binding of both the molecules was characterized by strong stabilization of the polynucleotides against thermal strand separation in optical melting as well as differential scanning calorimetry studies. Isothermal titration calorimetry results revealed that the binding of both sanguinarine and ethidium to poly(dG-dC).poly(dG-dC), poly(dA-dT).poly(dA-dT), and poly(dG).poly(dC) was exothermic and favoured by negative enthalpy changes. On the other hand, the binding of both molecules to poly(dA).poly(dT) was endothermic and entropy driven. The binding affinity values obtained from isothermal titration calorimetry data was in close proximity to that derived from thermal melting data. The heat capacity changes obtained from temperature dependence of the enthalpy change gave negative values in the range (?0.4 to 1.25) kJ · mol^?1 · K^?1 for the binding of ethidium and sanguinarine to these polynucleotides. The variations in the values indicate important differences in the formation of the complexes. New insights into the energetics and specificity aspects of interaction of these molecules to DNA have emerged from these studies.     

Sequence-dependent interactions of cationic naphthalimides and polynucleotides

The binding interactions of three naphthalimide derivatives with heteropoly nucleic acids have been evaluated using fluorescence, absorption and circular dichroism spectroscopies. Mono- and bifunctionalized naphthalimides exhibit sequence-dependent variations in their affinity toward DNA. The heteropoly nucleic acids, [Poly(dA-dT)]2 and [Poly(dG-dC)]2, as well as calf thymus (CT) DNA, were used to understand the factors that govern binding strength and selectivity. Sequence selectivity was addressed by determining the binding constants as a function of polynucleotide composition according to the noncooperative McGhee-von Hippel binding model. Binding affinities toward [poly(dA-dT)](2) were the largest for spermine-substituted naphthalimides (Kb = 2-6 x 10(6) M(-1)). The association constants for complex formation between the cationic naphthalimides and [poly(dG-dC)]2 or CT DNA (58% A-T content) were 2-500 times smaller, depending on the naphthalimide-polynucleotide pair. The binding modes were also assessed using a combination of induced circular dichroism and salt effects to determine whether the naphthalimides associate with DNA through intercalative, electrostatic or groove-binding. The results show that the monofunctionalized spermine and pyridinium-substituted naphthalimides associate with DNA through electrostatic interactions. In contrast, intercalative interactions are predominant in the complex formed between the bifunctionalized spermine compound and all of the polynucleotides.     

MONOADDITION OF DICTAMNINE TO SYNTHETIC DOUBLE-STRANDED POLYDEOXYRIBONUCLEOTIDES IN UVA AND THE EFFECT OF PHOTOMODIFIED DNA ON TEMPLATE ACTIVITY

Abstract— The photoreactivity of dictamnine, a furoquinoline alkaloid, towards different synthetic DNAs has been studied. The ratio of the photobinding of [₃H]-dictamnine to poly(dA-dT) poly(dA-dT): poly(dG-dC) poly(dG-dC): poly(dA-dU) poly(dA-dU): poly(dA) poly(dT), in relation to that of calf thymus DNA, is 18:1:0.5:0.3. Prior treatment of calf thymus DNA with dictamnine in light inhibits the subsequent incorporation of 8-methoxypsoralen (8-MOP). These results suggest that the sites in DNA for the photobinding of dictamnine are probably identical with those for monoad-ducts of 8-MOP. Furthermore, the template activity of photomodified DNA in the RNA polymerase reaction is considerably inhibited for poly(dA-dT)poly(dA-dT), to a lesser extent for calf thymus DNA, but almost not affected for the linear copolymer, poly(dA)-poly(dT).     

EFFICIENCY OF PHOTOAFFINITY LABELING DNA HOMOPOLYMERS AND COPOLYMERS WITH ETHIDIUM MONOAZIDE *

Abstract— Photoaffinity labeling of synthetic DN As with ethidium monoazide was studied to determine if the efficiency of adduct formation was related to DNA sequence. Equilibrium drug binding to DNA homopolymers and copolymers was quanitified by phase partition techniques. The amount of drug bound to a deoxypolymer at equilibrium was then compared to the fraction of ethidium analog covalently-linked following photoactivation at the same drug/DNA input ratio. There were significant sequence-related differences in the ability of the photoaffinity probe to label DNA covalently. The efficiency of covalent-adduct formation decreased in the order poly(dG-dC)^. poly(dG-dC)> poly-(dG). poly(dC)poly(dA-dT). poly(dA-dT)poly(dA). poly(dT). Ethidium monoazide was about 2-fold more efficient in labeling deoxyhomopolymers and deoxycopolymers composed of G-C pairs than the A-T base counterparts. In low ionic buffers (0.015 M Na⁺), the efficiency of photoactivation decreased with increasing ethidium monoazide concentrations. However. the base sequence effect was observed over a 40-fold range of drug concentrations. Therefore, the amount of ethidium monoazide bound to a DNA site after irradiation does not appear to represent the true affinity of the drug for that site.     

Long-range assemblies on poly(dG-dC)2 and poly(dA-dT)2: phosphonium cationic porphyrins and the importance of the charge

The present paper describes synthesis and spectroscopic properties of novel cationic meso-tetraphenylporphyrins bearing two (trans) (P2) or three (P3) triphenylphosphonium substituents. The porphyrin aggregation in aqueous solutions is discussed in detail. Porphyrin binding to and self-organization onto long-range assemblies on poly(dA-dT)2 or poly(dG-dC)2 were probed by combination of absorption, fluorescence, circular dichroism (CD), transient and resonance light-scattering (RLS) techniques. The higher hydrophobicity of P2 is manifested by more extensive self-organization. Induced CD and intensive RLS indicate binding to the chiral environment on the nucleic acids exterior and exciton coupling between adjacent porphyrin moieties. The CD spectra of P2 on poly(dG-dC), and poly(dA-dT)2 suggest that the binding geometry is essentially independent of the base sequence. The fluorescence lifetime of about 4 ns was attributed to the long-range assembly. In the case of P3 the distinctly different CD spectra induced by GC or AT base-pair regions reveal that the number of the substituents determines how closely the porphyrin can approach the specific electronic environment on the nucleic acid exterior. The fluorescence lifetime of the P3 assembly is about 2 ns.     

Luminescent Ir(III) complex exclusively made of polypyridine ligands capable of intercalating into calf-thymus DNA

Efficient intercalation of a luminescent Ir(III) complex exclusively made of polypyridine ligands in natural and synthetic biopolymers is reported for the first time. The emission of the complex is largely enhanced in the presence of [poly(dA-dT)(2)] and strongly quenched in the presence of [poly(dG-dC)(2)]. By comparing the emission decays in DNA and in synthetic polynucleotides, it is proposed that the emission quenching of the title compound by guanine residues in DNA is no longer effective over a distance of four dA-dT base pairs.     

ACMA (9-amino-6-chloro-2-methoxy acridine) forms three complexes in the presence of DNA

The interaction of ACMA (9-amino-6-chloro-2-methoxy acridine) (D) with DNA (P) has been studied by absorbance, fluorescence, circular dichroism, spectrophotometry, viscometry and unwinding electrophoresis. A T-jump kinetic study has also been undertaken. The experimental data show that, totally unlike other drugs, ACMA is able to form with DNA three complexes (PD(I), PD(II), PD(III)) that differ from each other by the characteristics and extent of the binding process. The main features of PD(I) fulfil the classical intercalation pattern and the formation/dissociation kinetics have been elucidated by T-jump techniques. PD(II) and PD(III) are also intercalated species but, in addition to the dye units lodged between base pairs, they also bear dye molecules externally bound, more in PD(III) relative to PD(II). A reaction mechanism is put forward here. Comparison between absorbance, fluorescence and kinetic experiments has enabled us to determine the binding constants of the three complexes, namely (6.5 ± 1.1) × 10(4) M(-1) (PD(I)), (5.5 ± 1.5) × 10(4) M(-1) (PD(II)) and (5.7 ± 0.03) × 10(4) M(-1) (PD(III)). The Comet assay reveals that the ACMA binding to DNA brings about genotoxic properties. The mutagenic potential studied by the Ames test reveals that ACMA can produce frameshift and transversion/transition mutations. ACMA also is able to produce base-pair substitution in the presence of S9 mix. Moreover, the MTT assays have revealed cytotoxicity. The biological effects observed have been rationalized in light of these features.     

Synthesis and DNA binding of novel water-soluble cationic methylcobalt porphyrins

Organocobalt derivatives of tetracationic water-soluble porphyrins are difficult to prepare via the typical reductive alkylation of the Co(II)(por) (porH(2) = porphyrin ligand). None have been reported. The problem may arise because the porphyrin core is made relatively electron poor by the positively charged peripheral groups. We have circumvented this problem by using the [Co(III)(NH(3))(5)CH(3)](2+) reagent, which inserts the Co(III)-CH(3) moiety directly into porH(2) in water under basic conditions. The method afforded two new [CH(3)Co(por)](4+) derivatives, [CH(3)CoTMpyP(4)](4+) and [CH(3)CoTMAP](4+), where [TMpyP(4)](4+) and [TMAP](4+) are the coordinated, NH-deprotonated forms of meso-tetrakis(N-methyl-4-pyridiniumyl)porphyrin and meso-tetrakis(N,N,N-trimethylaniliniumyl)porphyrin, respectively. The binding of the two new [CH(3)Co(por)](4+) cations to DNA and to the synthetic DNA polymers [poly(dA-dT)](2) and [poly(dG-dC)](2) was studied. Using published criteria by which changes in DNA viscosity and in the visible and CD spectra in the Soret region can be used to assess DNA binding, we conclude that both are outside binders. A large hypochromicity of the Soret bands of the [CH(3)Co(por)](4+) cations observed upon outside binding to DNA may indicate a high degree of self-stacking. The visible absorption and CD spectra of the [CH(3)Co(por)](4+) cations in the presence of 1:1 mixtures of [poly(dA-dT)](2) and [poly(dG-dC)](2) are nearly identical to those with [poly(dA-dT)](2) alone and are very different from those of [poly(dG-dC)](2) alone. Thus, both cations show a high preference for outside binding at AT-rich over GC-rich DNA sites. Upon binding of each of the [CH(3)Co(por)](4+) cations to all of the DNA polymers, the Soret bands exhibit blue shifts, whereas the Soret bands of the corresponding [(H(2)O)(2)Co(por)](5+) cations exhibit red shifts. The blue shifts strongly suggest that the [CH(3)Co(por)](4+) cations, particularly [CH(3)CoTMAP](4+), become five-coordinate forms to some extent on DNA binding; this result is the first good evidence for the presence at equilibrium of five-coordinate CH(3)Co(III)(N(4)) forms in water.     

Synthesis and photophysical evaluation of a pyridinium 4-amino-1,8-naphthalimide derivative that upon intercalation displays preference for AT-rich double-stranded DNA

The synthesis, characterisation and solid state crystal structure of a cationic 4-amino-1,8-naphthalimide derivative (1) are described. The photophysical properties of 1 are shown to vary with the solvent polarity and H-bonding ability. The fluorescence of 1 is enhanced and blue-shifted in its 1:1 complex with 5'-adenosine-monophosphate while it is partially quenched and red-shifted in its complex with 5'-guanosine-monophosphate. Linear and circular dichroism measurements show that 1 binds to double-stranded DNA by intercalation. Comparative UV-visible and fluorescence studies with double stranded synthetic polynucleotides poly(dA-dT)(2) and poly(dG-dC)(2) show that 1 binds much more strongly to the AT polymer; 1 also has a strong preference for A-T rich sequences in natural DNA. Thermal denaturation measurements also reveal a much greater stabilisation of the double-stranded poly(dA-dT)(2) than of natural DNA.     

Photooxidation of guanine by a ruthenium dipyridophenazine complex intercalated in a double-stranded polynucleotide monitored directly by picosecond visible and infrared transient absorption spectroscopy

Transient species formed by photoexcitation (400 nm) of [Ru(dppz)(tap)2]2+ (1) (dppz = dipyrido[3,2-a:2',3'-c]phenazine; tap=1,4,5,8-tetraazaphenanthrene) in aqueous solution and when intercalated into a double-stranded synthetic polynucleotide, [poly(dG-dC)]2, have been observed on a picosecond timescale by both visible transient absorption (allowing monitoring of the metal complex intermediates) and transient infrared (IR) absorption spectroscopy (allowing direct study of the DNA nucleobases). By contrast with its behavior when free in aqueous solution, excitation of 1 when bound to [poly(dG-dC)]2 causes a strong increase in absorbance at 515 nm due to formation of the reduced complex [Ru(dppz)(tap)2]+ (rate constant=(2.0+/-0.2) x 10(9) s(-1)). The subsequent reformation of 1 proceeds with a rate constant of (1.1+/-0.2) x 10(8) s(-1). When the process is carried out in D2O, the rates of formation and removal of [Ru(dppz)(tap)2]+ are reduced (rate constants (1.5+/-0.3) x 10(9) and (0.7+/-0.2) x 10(8) s(-1) respectively) consistent with proton-coupled electron transfer processes. Picosecond transient IR measurements in the 1540-1720 cm(-1) region in D2O solution confirm that the reduction of 1 intercalated into [poly(dG-dC)]2 is accompanied by bleaching of IR ground-state bands of guanine (1690 cm(-1)) and cytosine (1656 cm(-1)), each with similar rate constants.     

A PORPHYRIN-DNA EXCIPLEX: RESONANCE RAMAN SPECTRA OF ELECTRONICALLY EXCITED Cu(TMpy-P4) BOUND TO POLY(dA-dT)-POLY(dA-dT)

The resonance Raman spectra of water-soluble porphyrins, Cu(TMpy-P4) and Ni(TMpy-P4), and their mixtures with DNA, Poly(dG-dC).Poly(dG-dC), and Poly(dA-dT).Poly(dA-dT) were measured using 426 nm pulsed laser excitation (and 556 nm for some applications). At high laser power, the solution of Cu(TMpy-P4) mixed with DNA or Poly(dA-dT).Poly(dA-dT) exhibits new bands at 1550 and 1349 cm-1 that are not observed for Cu(TMpy-P4) alone or for Cu(TMpy-P4) mixed with Poly(dG-dC).Poly(dG-dC). These extra bands do not appear when the resonance Raman spectra are measured by a cw laser or by a pulsed laser with low power. Similar mixtures of M(TMpy-P4) (where M = Ni, Zn, Co, Mn, and H2) with these nucleic acids exhibit no such bands even by high power pulsed laser excitation. We attribute the new resonance Raman bands to an electronically excited Cu(TMpy-P4), stabilized by forming an exciplex with the A-T site of the nucleic acid. The minimum lifetime value of such an exciplex was estimated to be on the order of 10 ps.     

The flash-quench technique in protein-DNA electron transfer: reduction of the guanine radical by ferrocytochrome c

Electron transfer from a protein to oxidatively damaged DNA, specifically from ferrocytochrome c to the guanine radical, was examined using the flash-quench technique. Ru(phen)2dppz2+ (dppz = dipyridophenazine) was employed as the photosensitive intercalator, and ferricytochrome c (Fe3+ cyt c), as the oxidative quencher. Using transient absorption and time-resolved luminescence spectroscopies, we examined the electron-transfer reactions following photoexcitation of the ruthenium complex in the presence of poly(dA-dT) or poly(dG-dC). The luminescence-quenching titrations of excited Ru(phen)2dppz2+ by Fe3+ cyt c are nearly identical for the two DNA polymers. However, the spectral characteristics of the long-lived transient produced by the quenching depend strongly upon the DNA. For poly(dA-dT), the transient has a spectrum consistent with formation of a [Ru(phen)2dppz3+, Fe2+ cyt c] intermediate, indicating that the system regenerates itself via electron transfer from the protein to the Ru(III) metallointercalator for this polymer. For poly(dG-dC), however, the transient has the characteristics expected for an intermediate of Fe2+ cyt c and the neutral guanine radical. The characteristics of the transient formed with the GC polymer are consistent with rapid oxidation of guanine by the Ru(III) complex, followed by slow electron transfer from Fe2+ cyt c to the guanine radical. These experiments show that electron holes on DNA can be repaired by protein and demonstrate how the flash-quench technique can be used generally in studying electron transfer from proteins to guanine radicals in duplex DNA.     

Understanding the factors controlling the photo-oxidation of natural DNA by enantiomerically pure intercalating ruthenium polypyridyl complexes through TA/TRIR studies with polydeoxynucleotides and mixed sequence oligodeoxynucleotides

Ruthenium polypyridyl complexes which can sensitise the photo-oxidation of nucleic acids and other biological molecules show potential for photo-therapeutic applications. In this article a combination of transient visible absorption (TrA) and time-resolved infra-red (TRIR) spectroscopy are used to compare the photo-oxidation of guanine by the enantiomers of [Ru(TAP)₂(dppz)]²⁺ in both polymeric {poly(dG-dC), poly(dA-dT) and natural DNA} and small mixed-sequence duplex-forming oligodeoxynucleotides. The products of electron transfer are readily monitored by the appearance of a characteristic TRIR band centred at ca. 1700 cm⁻¹ for the guanine radical cation and a band centered at ca. 515 nm in the TrA for the reduced ruthenium complex. It is found that efficient electron transfer requires that the complex be intercalated at a G-C base-pair containing site. Significantly, changes in the nucleobase vibrations of the TRIR spectra induced by the bound excited state before electron transfer takes place are used to identify preferred intercalation sites in mixed-sequence oligodeoxynucleotides and natural DNA. Interestingly, with natural DNA, while it is found that quenching is inefficient in the picosecond range, a slower electron transfer process occurs, which is not found with the mixed-sequence duplex-forming oligodeoxynucleotides studied.

Efficient electron transfer requires the complex to be intercalated at a G-C base-pair. Identification of preferred intercalation sites is achieved by TRIR monitoring of the nucleobase vibrations before electron transfer.     

三维有序金掺杂纳米TiO_2修饰电极用于抗肿瘤药物与DNA相互作用的研究

利用模板法在氧化铟锡(ITO)电极表面制备了三维有序多孔结构的金掺杂纳米Ti O2薄膜修饰电极(3DOM GTD/ITO),并在此修饰电极上成功固定小牛胸腺DNA(ct DNA),从而构建了一种新型的DNA生物传感器(DNA/3DOM GTD/ITO),并通过透射电镜(TEM)、扫描电镜(SEM)对修饰电极的表面形貌进行表征。采用电化学交流阻抗(EIS)法研究了ct DNA在3DOM GTD/ITO修饰电极表面的固定情况,结果表明,ct DNA已被成功地固定在3DOM GTD/ITO修饰电极表面。采用循环伏安法、微分脉冲伏安法等电化学方法研究了抗肿瘤药物槲皮素(Qu)在3DOM GTD/ITO修饰电极表面的电化学性质及与ct DNA的相互作用。结果表明,Qu在3DOM GTD/ITO修饰电极表面有1对准可逆的氧化还原峰,其氧化还原反应为2电子和2质子的转移过程。Qu可与固定在修饰电极上的ct DNA发生较强的结合作用,其结合常数(K)为3.61×106L/mol。循环伏安实验、紫外-可见吸收光谱、分子荧光光谱、圆二色性光谱均表明Qu与ct DNA之间的相互作用模式为嵌插作用。Qu与ct DNA的碱基结合具有序列选择性,对Qu与聚(d G-d C)及聚(d A-d T)的结合常数进行计算,得到结合常数比K(d G-d C)/K(d A-d T)=3.5,表明Qu与ct DNA发生嵌插作用时更倾向于结合在GC富集区域。     

硫堇与寡核苷酸结合序列选择性的光谱研究

用UV-Vis吸收光谱、荧光光谱、圆二色谱以及核磁共振光谱等手段研究了硫堇(TH)与两个不同序列寡核苷酸的作用。TH与寡核苷酸作用后的吸收光谱和荧光光谱产生了明显的减色红移和荧光猝灭效应。分别计算了TH与[oligo d(GC)]2和[oligo d(AT)]2作用的荧光猝灭常数和结合常数,结果表明TH与GC序列的结合能力比与AT序列更强。通过TH与[oligo d(GC)]2作用后双螺旋链构象变化以及TH质子的1HNMR谱峰明显变宽,进一步说明TH与寡核苷酸结合的序列选择性。