Unmediated by DNA electron transfer in redox-labeled DNA duplexes end-tethered to gold electrodes

Electron transfer (ET) between gold electrodes and redox-labeled DNA duplexes, immobilized onto the electrodes through the alkanethiol linker at the 3'-end and having internal either methylene blue (MB) or anthraquinone (AQ) redox labels, was shown to depend on the redox label charge and the way the redox label is linked to DNA. For loosely packed DNA monolayers, the conjugation of the positively charged MB to DNA through the long and flexible alkane linker provided ET whose kinetics was formally governed by the diffusion of the redox label to the negatively charged electrode surface. For the uncharged AQ label no ET signal was detected. The conjugation of AQ to DNA through the short and more conductive acetylene linker did not provide the anticipated DNA-mediated ET to the AQ-moiety: ET appeared to be low-efficient if any in the studied system, for which no intercalation of AQ within the DNA duplex occurred. The ET communication between the electrode and AQ, built in DNA through the acetylene linker, was achieved only when Ru(NH(3))(6)(3+) molecules were electrostatically attached to the DNA duplex, thus forming the electronic wire. These results are of particular importance both for the fundamental understanding of the interfacial behavior of the redox labeled DNA on electrodes and for the design of biosensors exploiting a variation of ET properties of DNA in the course of hybridization.     

Fluorescent silver nanoclusters in hybridized DNA duplexes for the turn-on detection of Hg2+ ions

Using the Hg(2+)-mediated T-T formation to strengthen the DNA duplexes and influence the configuration of fluorescent Ag NCs-forming sequences, a turn-on fluorescence detection method for Hg(2+) has been established.     

Charge transfer excited-state dynamics in DNA duplexes substituted with an ethynylpyrenyldeoxyuridine electron source and a fluorodeoxyuridine electron trap

Studies of six 5-(pyren-1-yl-ethynyl)-2'-deoxyuridine (U(PY))-substituted DNA duplexes in this work test and support the conclusions reported by Gaballah et al. (J. Phys. Chem. B 2005, 109, 5927-5934) based on investigations of 5-(2-pyren-1-yl-ethylenyl)-2'-deoxyuridine (U(PE))-substituted DNA hairpins. As expected because of the rigid ethynyl linker in U(PY) (compared to the flexible ethylenyl linker in U(PE)), U(PY)-substituted duplexes do not show enhanced charge transfer (CT) emission quantum yields for duplexes with 5-fluorodeoxyuridine (U(F)) electron traps near U(PY) compared to duplexes without traps. Furthermore, the average CT lifetime and emission quantum yield of U(PY)-substituted duplexes is independent of the U(F) trap location. These new results strongly suggest that the excess electron in the PY*+/dU*- CT state of U(PY) is restrained from hopping to nearby U(F) traps due to attraction to PY*+.     

Site-selective photoinduced electron transfer from alcoholic solvents to the chromophore facilitated by hydrogen bonding: a new fluorescence quenching mechanism

Solute-solvent intermolecular photoinduced electron transfer (ET) reaction was proposed to account for the drastic fluorescence quenching behaviors of oxazine 750 (OX750) chromophore in protic alcoholic solvents. According to our theoretical calculations for the hydrogen-bonded OX750-(alcohol)(n) complexes using the time-dependent density functional theory (TDDFT) method, we demonstrated that the ET reaction takes place from the alcoholic solvents to the chromophore and the intermolecular ET passing through the site-specific intermolecular hydrogen bonds exhibits an unambiguous site selectivity. In our motivated experiments of femtosecond time-resolved stimulated emission pumping fluorescence depletion spectroscopy (FS TR SEP FD), it could be noted that the ultrafast ET reaction takes place as fast as 200 fs. This ultrafast intermolecular photoinduced ET is much faster than the diffusive solvation process, and even significantly faster than the intramolecular vibrational redistribution (IVR) process of the OX750 chromophore. Therefore, the ultrafast intermolecular ET should be coupled with the hydrogen-bonding dynamics occurring in the sub-picosecond time domain. We theoretically demonstrated for the first time that the selected hydrogen bonds are transiently strengthened in the excited states for facilitating the ultrafast solute-solvent intermolecular ET reaction.     

Dynamics of electron transport by elastic bending of short DNA duplexes. Experimental study and quantitative modeling of the cyclic voltammetric behavior of 3'-ferrocenyl DNA end-grafted on gold

The dynamics of electron transport within a molecular monolayer of 3'-ferrocenylated-(dT)(20) strands, 5'-thiol end-grafted onto gold electrode surfaces via a short C2-alkyl linker, is analyzed using cyclic voltammetry as the excitation/measurement technique. It is shown that the single-stranded DNA layer behaves as a diffusionless system, due to the high flexibility of the ss-DNA chain. Upon hybridization by the fully complementary (dA)(20) target, the DNA-modified gold electrode displays a highly unusual voltammetric behavior, the faradaic signal even ultimately switching off at a high enough potential scan rate. This remarkable extinction phenomenon is qualitatively and quantitatively justified by the model of elastic bending diffusion developed in the present work which describes the motion of the DNA-borne ferrocene moiety as resulting from the elastic bending of the duplex DNA toward and away from the electrode surface. Its use allows us to demonstrate that the dynamics of electron transport within the hybridized DNA layer is solely controlled by the intrinsic bending elasticity of ds-DNA. Fast scan rate cyclic voltammetry of end-grafted, redox-labeled DNA layers is shown to be an extremely efficient method to probe the bending dynamics of short-DNA fragments in the submillisecond time range. The persistence length of the end-anchored ds-DNA, a parameter quantifying the flexibility of the nanometer-long duplex, can then be straightforwardly and accurately determined from the voltammetry data.     

Direct energy transfer from conjugated polymer to DNA intercalated dye: label-free fluorescent DNA detection

The development of methods for DNA detection is of importance in disease diagnosis, gene-targeted drug discovery and molecular biology field. In this paper, we synthesize a new cationic water-soluble CP containing fluorene moiety and flexible ethylenic moiety in the backbone (PFV) for label-free DNA detection. The conformational freedom of PFV provides stronger interactions with double-stranded DNA (dsDNA) and optimizes the orientation of transition moments between PFV and ethidium bromide (EB) intercalated in dsDNA. The efficient FRET from PFV (donor) to EB (acceptor) intercalated in dsDNA is observed and the emission of EB is amplified by the good light-harvesting ability of conjugated polymers. The interactions between PFV and DNA can also be probed by measuring the FRET ratio between PFV and EB intercalated in DNA. In comparison to other DNA detection assays based on FRET and conjugated polymers, synthesis of dye-labeled DNA probe is avoided in our method, which significantly reduces the cost and the synthetic complexity. The PFV/dsDNA/EB system provides promising applications on DNA detection with a simply, fast and label-free manner.     

Intermolecular vs. intramolecular photoinduced electron transfer from nucleotides in DNA to acridinium ion derivatives in relation with DNA cleavage

Photoirradiation of various 10-methylacridinium ions (AcrR⁺, R = H, ⁱPr, and Ph) intercalated in DNA results in ultrafast intramolecular electron transfer, followed by rapid back electron transfer between AcrR⁺ and nucleotides in DNA. The electron-transfer dynamics in DNA were monitored by femtosecond time-resolved transient absorption spectroscopy. Both acridinyl radical and nucleotide radical cations, formed in the photoinduced electron transfer in DNA, were successfully detected in an aqueous solution. These transient absorption spectra were assigned by the comparison with those of DNA nucleotide radical cations, which were obtained by the intermolecular electron-transfer oxidation of nucleotides with the electron-transfer state of 9-mesityl-10-methylacridinium ion (Acr–Mes⁺) produced upon photoexcitation of Acr⁺–Mes. Photoinduced cleavage of DNA with various acridinium ions (AcrR⁺, R = H, ⁱPr, Ph, and Mes) has also been examined by agarose gel electrophoresis, which indicates that the rapid intramolecular back electron transfer between acridinyl radical and nucleotide radical cation in DNA suppresses the DNA cleavage as compared with the intermolecular electron-transfer oxidation of nucleotides with Acr–Mes⁺.     

Carbon paste electrode for voltammetric detection of a specific DNA sequence from potentially aflatoxigenic Aspergillus species

An electrochemical biosensor for specific detection of DNA sequences from Aspergillus, a potentially aflatoxigenic fungus is reported. The detection technique was optimized using short DNA sequences, then applied for a 92-mer from the Aspergillus gene aflR that codes a biochemical pathway of aflatoxin B1 production.Voltammetric detection of the specific Aspergillus DNA sequence is based on hybridization of adsorbed target DNA with a biotinylated probe and subsequent binding with streptavidin alkaline phosphatase conjugate. Such modified electrode surface of carbon paste electrode (CPE) is incubated in a buffer with an electrochemically inactive substrate (1-naphthyl phosphate). Alkaline phosphatase converts 1-naphthyl phosphate into 1-naphthol, which is determined by the selected voltammetric technique. The optimized procedure provided good differentiation between complementary and noncomplementary probes and also between potentially aflatoxigenic and non-aflatoxigenic fungi of the genus Aspergillus.     

A voltammetric study of direct electron transfer to cytochrome c using a very large assembly of carbon microelectrodes

A very large assembly of more than 8000 carbon fibre microdisk electrodes was used to study direct electron transfer to cytochrome c. Near steady-state cyclic voltammograms were observed, which exhibited excellent signal-to-noise ratios despite the low concentrations of cytochrome c employed (1-50 microM). The high resolution of the voltammograms allowed the formal potential of the native form of cytochrome c to be determined over a range of solution pH.     

Interprotein electron transfer in a confined space: uncoupling protein dynamics from electron transfer by sol-gel encapsulation

In this paper, we describe the first observations of photoinitiated interprotein electron transfer (ET) within sol-gels. We have encapsulated three protein-protein complexes, specifically selected because they represent a full range of affinities, are sensitive to different types of dynamic processes, and thus are expected to respond differently to sol-gel encapsulation. The three systems are (i) the [Zn, Fe(3+)L] mixed-metal hemoglobin hybrids, where the alpha(1)-Zn and beta(2)-Fe subunits correspond to a "predocked" protein-protein complex with a crystallographically defined interface (Natan, M. J.; Baxter, W. W.; Kuila, D.; Gingrich, D. J.; Martin, G. S.; Hoffman, B. M. Adv. Chem. Ser. 1991, 228 (Electron-Transfer Inorg., Org., Biol. Syst.), 201-213), (ii) the Zn-cytochrome c peroxidase complex with cytochrome c, [ZnCcP, Fe(3+)Cc], having an intermediate affinity between its partners (Nocek, J. M.; Zhou, J. S.; De Forest, S.; Priyadarshy, S.; Beratan, D. N.; Onuchic, J. N.; Hoffman, B. M. Chem. Rev. 1996, 96, 2459-2489), and (iii) the [Zn-deuteromyoglobin, ferricytochrome b(5)] complex, [ZnDMb, Fe(3+)b(5)], which is loosely bound and highly dynamic (Liang, Z.-X.; Nocek, J.; Huang, K.; Hayes, R. T.; Kurnikov, I. V.; Beratan, D. N.; Hoffman, B. M. J. Am. Chem. Soc. 2002, 124, 6849-6859. Intersubunit ET within the hybrid does not involve second-order processes or subunit rearrangements, and thus is influenced only by perturbations of high-frequency motions coupled to ET. For the latter two complexes, sol-gel encapsulation eliminates second-order processes: protein partners encapsulated as a complex must stay together throughout a photoinitiated ET cycle, while proteins encapsulated alone cannot acquire a partner. It further modulates intracomplex motions of the two partners.     

NMR studies of solvent-assisted proton transfer in a biologically relevant Schiff base: toward a distinction of geometric and equilibrium H-bond isotope effects

The tautomeric equilibrium in a Schiff base, N-(3,5-dibromosalicylidene)-methylamine 1, a model for the hydrogen bonded structure of the cofactor pyridoxal-5'-phosphate PLP which is located in the active site of the enzyme, was measured by means of 1H and 15N NMR and deuterium isotope effects on 15N chemical shifts at variable temperature and in different organic solvents. The position of the equilibrium was estimated using the one-bond 1J(OHN) and vicinal 3J(H(alpha)CNH) scalar coupling constants. Additionally, DFT calculations of a series of Schiff bases, N-(R1-salicylidene)-alkyl(R2)amines, were performed to obtain the hydrogen bond geometries. The latter made it possible to investigate a broad range of equilibrium positions. The increase of the polarity of the aprotic solvent shifts the proton in the intramolecular OHN hydrogen bond closer to the nitrogen. The addition of methanol and of hexafluoro-2-propanol to 1 in aprotic solvents models the PLP-water interaction in the enzymatic active site. The alcohols, which vary in acidity and change the polarity around the hydrogen bond, also stabilize the equilibrium, so that the proton is shifted to the nitrogen.     

Photo-induced intermolecular electron transfer from electron donating solvents to Coumarin dyes in bile salt aggregates: role of diffusion in electron transfer reaction

The photo-induced electron transfer between Coumarin dyes and aromatic amines has been investigated using steady state and time-resolved fluorescence quenching studies. We have observed a Marcus type inversion in the electron transfer rate in correlation of quenching constant to the free energy change occurred during reaction. To justify the "inverted region" obtained in the correlation of quenching constant versus free energy curve, we have performed anisotropy measurement and estimated the several diffusional parameters. The translational diffusion coefficients exhibit a similar picture like electron transfer rate constant when it is plotted against free energy. Thus we argued that the diffusion has played an important role in the electron transfer kinetics.     

Repair of guanyl radicals in plasmid DNA by electron transfer is coupled to proton transfer

By using gamma-irradiation in the presence of thiocyanate ions, we have generated guanyl radicals in plasmid DNA. These can be detected by using an Escherichia coli base excision repair endonuclease to convert their stable end products to strand breaks. The yield of enzyme-sensitive sites is strongly attenuated by the presence of micromolar concentrations of one of a series of singly substituted phenols, and it is possible to derive bimolecular rate constants for the reduction of DNA guanyl radicals by these phenols. More strongly reducing phenols were found to react more rapidly. This electron-transfer reaction also involves a proton transfer. By comparing the expected energetics of the reaction with the observed rate constants, the electron transfer is found to be mechanistically coupled with the proton transfer.     

Solid-phase synthesis of phenolic steroids: from optimization studies to a convenient procedure for combinatorial synthesis of biologically relevant estradiol derivatives

During the course of our studies on therapeutic agents for the treatment of breast cancer, we became interested in the solid-phase combinatorial synthesis of estradiol derivatives that contain a functionalized side chain at either position 16 beta or 7 alpha. Both types of compounds have already demonstrated inhibitory activity toward both biosynthesis and action of estradiol. As a first step, two versatile precursors bearing an azidoalkyl side chain at either position 16 beta or 7 alpha of estradiol were synthesized using standard solution-phase methods. Afterward, the effectiveness of five linkers to attach the phenolic function of these estradiol derivatives to a polystyrene resin was investigated; they were benzylic ether (Merrifield), 4-alkoxy-benzylic ethers (Wang, Sheppard), tetrahydropyranyl ether (Ellman), benzoic ester, and o-nitrobenzyl ether. To test the linker in a synthetic context, a short sequence of reactions, including reduction of the azide and acylation of the corresponding amine, was performed on the polymer-bound estradiol derivative. While all of the tested linkers proved effective in attaching the phenol functionality of the precursor, only the o-nitrobenzyl ether photolabile linker enabled the release of the final products in acceptable purities. Consequently, this linker was used to perform successfully the solid-phase synthesis of four different classes of estradiol derivatives in acceptable yields and excellent purities. This study was preliminary to the combinatorial synthesis of larger libraries of biologically relevant estradiol derivatives.     

DNA cleavage induced by thermal electron transfer from a dimeric NADH analogue to acridinium ions in the presence of oxygen

Acridinium ions, which can intercalate to DNA, act as thermal DNA cleavers in the presence of a dimeric NADH analogue and O2 in an aqueous solution via thermal electron transfer from a dimeric NADH analogue to acridinium ions, followed by the electron-transfer reduction of O2 to O2*- by the resulting NAD radicals.     

Excess electron transfer from an internally conjugated aromatic amine to 5-bromo-2'-deoxyuridine in DNA

DNA duplexes containing an N,N,N',N'-tetramethyl-1,5-diaminonaphthalene analogue and 5-bromo-2'-deoxyuridine (BrdU) provide a readily accessible system for investigating excess electron transfer in DNA. Photoexcitation of the aromatic amine (lambda > 335 nm) induces reductive electron transfer as observed by strand cleavage adjacent to the BrdU residue. The weak exponential distance dependence (0.3 A-1) of electron transfer determined for this system of mixed dA-T and dG-dC base pairs suggests that thermally activated electron hopping is competitive with proton transfer within the dG.dC radical anion. The UV-dependent transfer of excess electrons and subsequent strand cleavage proceeds equivalently under anaerobic and aerobic conditions and is not sensitive to e-(aq) or hydroxyl radical trapping agents.     

Autoxidation of adamantylideneadamantane in a protic solvent. Facile proton-induced electron transfer from the olefin to oxygen

Addition of trifluoroacetic acid to adamantylideneadamantane in dichloromethane solution under oxygen in the dark results in a rapid oxygenation of the olefin. The reaction is proposed to proceed through a mechanism involving proton-induced electron transfer from the olefin to oxygen giving its radical cation and a hydroperoxy radical followed by their subsequent reactions.     

Selective DNA detection at Zeptomole level based on coulometric measurement of gold nanoparticle-mediated electron transfer across a self-assembled monolayer

A selective DNA sensing with zeptomole detection level is developed based on coulometric measurement of gold nanoparticle (AuNPs)-mediated electron transfer (ET) across a self-assembled monolayer on the gold electrode. After immobilization of a thiolated hairpin-structured DNA probe, an alkanethiol monolayer was self-assembled on the resultant electrode to block [Fe(CN)6 ]-3-/4in a solution from accessing the electrode. In the presence of DNA target, hybridization between the DNA probe and the DNA target breaks the stem duplex of DNA probe. Consequently, stem moiety at the 3′-end of the DNA probes was removed from the electrode surface and made available for hybridization with the reporter DNA-AuNPs conjugates (reporter DNA-AuNPs). The thiolated reporter DNA matches the stem moiety at the 3′-end of the DNA probe. AuNPs were then enlarged by immersing the electrode in a growth solution containing HAuCl 4 and H2O2 after the reporter DNA-AuNPs bound onto the electrode surface. The enlarged AuNPs on the electrode restored the ET between the electrode and the [Fe(CN)6]3 -/4- , as a result, amplified signals were achieved for DNA target detection using the coulometric measurement of Fe(CN)6 3- electro-reduction by prolonging the electrolysis time. The quantities of ET on the DNA sensor increased with the increase in DNA target concentration through a linear range of 3.0 fM to 1.0 pM when electrolysis time was set to 300 s, and the detection limit was 1.0 fM. Correspondingly, thousands of DNA (zeptomole) copies were detected in 10L samples. Furthermore, the DNA sensor showed excellent differentiation ability for single-base mismatch.     

Photoinduced electron transfer from N,N-dimethylaniline to 7-amino coumarins in protein-surfactant complex: slowing down of electron transfer dynamics compared to micelles

Photoinduced electron transfer from N,N-dimethylaniline to different Coumarin dyes has been investigated in dodecyl trimethyl ammonium bromide (DTAB) micelles and in Bovine serum albumin (BSA)-DTAB protein-surfactant complex using steady-state and picosecond time-resolved fluorescence spectroscopy. We observed a slower fluorescence quenching rate in the DTAB micelles and in the protein-surfactant complex as compared to that in pure acetonitrile solution. Moreover, the observed fluorescence quenching in BSA-DTAB complex was found to be slower than that in DTAB micelles. In the correlation of free-energy change with the fluorescence quenching constant we observed a deviation in the fluorescence quenching electron transfer rate for Coumarin 151 (C-151) from the normal Marcus curve. This observation is ascribed to the stronger interaction of C-151 with the surfactant molecules present in the micelles. This is evident from the slower translation diffusion (D(L)) of Coumarin 151 compared to other probe molecules.