Natalia Borovok

Tel Aviv University, Tell Afif, Tel Aviv, Israel

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Publications (29)127.4 Total impact

  • Natalia Borovok, Elad Gillon, Alexander Kotlyar
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    ABSTRACT: Here, we present a relatively simple, efficient, and high-yielding polymerase-based method for the synthesis of 15 nm gold nanoparticle conjugates bearing a specific number of 25 base oligonucleotide strands. We have shown that the conjugates bearing one or two oligonucleotide strands per particle, with the conjugates comprising a single complementary strand, self-assemble into nanoparticle dimers and trimers, respectively. Incubation of fully coated AuNPs, containing tens of oligonucleotide strands, with a conjugate bearing a single complementary strand leads to the formation of flower-shaped structures. The assembly of particles into nanoparticle structures shown here is a prerequisite for more complex controlled assembly of particles into three-dimensional macrostructures.
    Bioconjugate Chemistry 04/2012; · 4.58 Impact Factor
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    ABSTRACT: In the present work, we have synthesized conjugates between the 5 nm gold nanoparticles (Au-NP) and 5' thiol-functionalized, 700 bp poly(dG)-poly(dC). We have completely separated and purified to homogeneity conjugates bearing different number of poly(dG)-poly(dC) molecules per Au-NP by electrophoresis and HPLC. The conjugates were directly visualized by atomic force microscopy. We have demonstrated that Au NP-bound poly(dG)-poly(dC) can be considerably extended by Klenow exo(-) polymerase in the presence of dCTP and dGTP.
    Bioconjugate Chemistry 02/2010; · 4.58 Impact Factor
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    ABSTRACT: We describe a method for the preparation of novel long (hundreds of nanometers), uniform, inter-molecular G4-DNA molecules composed of four parallel G-strands. The only long continuous G4-DNA reported so far are intra-molecular structures made of a single G-strand. To enable a tetra-molecular assembly of the G-strands we developed a novel approach based on avidin-biotin biological recognition. The steps of the G4-DNA production include: (i) Enzymatic synthesis of long poly(dG)-poly(dC) molecules with biotinylated poly(dG)-strand; (ii) Formation of a complex between avidin-tetramer and four biotinylated poly(dG)-poly(dC) molecules; (iii) Separation of the poly(dC) strands from the poly(dG)-strands, which are connected to the avidin; (iv) Assembly of the four G-strands attached to the avidin into tetra-molecular G4-DNA. The average contour length of the formed structures, as measured by AFM, is equal to that of the initial poly(dG)-poly(dC) molecules, suggesting a tetra-molecular mechanism of the G-strands assembly. The height of tetra-molecular G4-nanostructures is larger than that of mono-molecular G4-DNA molecules having similar contour length. The CD spectra of the tetra- and mono-molecular G4-DNA are markedly different, suggesting different structural organization of these two types of molecules. The tetra-molecular G4-DNA nanostructures showed clear electrical polarizability. This suggests that they may be useful for molecular electronics.
    Nucleic Acids Research 10/2008; 36(15):5050-60. · 8.28 Impact Factor
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    ABSTRACT: The molecular morphology of long G4-DNA wires made by a novel synthetic method was, for the first time, characterized by high-resolution scanning tunneling microscopy (STM). The STM images reveal a periodic structure seen as repeating "bulbs" along the molecules. These bulbs reflect the helix morphology of the wires. The STM measurements were supported by a statistical morphology analysis of the DNA pitch length and apparent height relative to the surface. In the absence of X-ray and NMR data for these wires, the STM measurements provide a unique alternative to characterize the helix morphology.
    The Journal of Physical Chemistry B 08/2008; 112(31):9267-9. · 3.61 Impact Factor
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    ABSTRACT: Here we describe a novel and efficient procedure for preparation of long uniform G4-DNA wires. The procedure includes (i) enzymatic synthesis of double-stranded DNA molecules consisting of long (up to 10,000 bases), continuous G strands and chains of complementary (dC)20-oligonucleotides, poly(dG)-n(dC)20; (ii) size exclusion HPLC separation of the G strands from the (dC)20 oligonucleotides in 0.1M NaOH; and (iii) folding of the purified G strands into G4-DNA structures by lowering the pH to 7.0. We show by atomic force microscopy (AFM) that the preparation procedure yielded G4-DNA wires with a uniform morphology and a narrow length distribution. The correlation between the total amount of nucleotides in the G strands and the contour length of the G4-DNA molecules estimated by AFM suggests monomolecular folding of the G strands into quadruplex structures. The folding takes place either in the presence or in the absence of stabilizing ions (K+ or Na+). The addition of these cations leads to a dramatic change in the circular dichroism spectrum of the G4-DNA.
    Analytical Biochemistry 04/2008; 374(1):71-8. · 2.58 Impact Factor
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    ABSTRACT: Abstract— Intense illumination (60-120 MW/cm2) of an oxygen-free aqueous solution of pyranine (8-hydroxypyrene-l,3,6-tri-sulfonate) by the third harmonic frequency of an Nd-Yag laser (355 nm) drives a two successive-photon oxidative process of the dye. The first photon excites the dye to its first electronic singlet state. The second photon interacts with the excited molecule, ejects an electron to the solution and deactivates the molecule to a ground state of the oxidized dye (φ+). The oxidized product, φ+, is an intensely colored compound (Λmax= 445 nm, ε= 43 000 ± 1000 M−1 cm−1) that reacts with a variety of electron donors like quinols, ascorbate and ferrous compounds. In the absence of added reductant, φ+ is stable, having a lifetime of -10 min. In acidic solutions the solvated electrons generated by the photochemical reaction react preferentially with H+. In alkaline solution the favored electron acceptor is the ground-state pyranine anion and a radical, φ, of the reduced dye is formed. The reduced product is well distinguished from the oxidized one, having its maximal absorption at 510 nm with e = 25 000 ± 2000 M-l cm−1. The oxidized radical can be reduced either by φ- or by other electron donors. The apparent second-order rate constants of these reactions, which vary from 106 up to 109M−1 s−1, are slower than the rates of diffusion-controlled reactions. Thus the redox reactions are limited by an energy barrier for electron transfer within the encounter complex between the reactants.
    Photochemistry and Photobiology 01/2008; 63(4):448 - 454. · 2.29 Impact Factor
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    ABSTRACT: Three types of DNA: approximately 2700 bp polydeoxyguanylic olydeoxycytidylic acid [poly(dG)-poly(dC)], approximately 2700 bp polydeoxyadenylic polydeoxythymidylic acid [poly(dA)-poly(dT)] and 2686 bp linear plasmid pUC19 were deposited on a mica surface and imaged by atomic force microscopy. Contour length measurements show that the average length of poly(dG)-poly(dC) is approximately 30% shorter than that of poly(dA)-poly(dT) and the plasmid. This led us to suggest that individual poly(dG)-poly(dC) molecules are immobilized on mica under ambient conditions in a form which is likely related to the A-form of DNA in contrast to poly(dA)-poly(dT) and random sequence DNA which are immobilized in a form that is related to the DNA B-form.
    FEBS Letters 01/2008; 581(30):5843-6. · 3.58 Impact Factor
  • Irit Lubitz, Natalia Borovok, Alexander Kotlyar
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    ABSTRACT: Interaction of meso-tetrakis(4-N-methylpyridyl)porphyrin (TMPyP) with G4-wires composed of approximately 1000 stacked tetrads (Kotlyar, A. B., Borovok, N., Molotsky, T., Cohen, H., Shapir, E., and Porath, D. (2005) Long monomolecular G4-DNA nanowires, Adv. Mater. 17, 1901-1905) was studied. These wires exist in either K (Na)-free or K forms in contrast to short telomeric G-quadruplexes, which are stable only in the presence of monovalent cations. We showed that a stable complex between K-free G4-wires and the porphyrin is formed at a TMPyP to tetrad molar ratio of 0.5. A 19 nm shift and a hypochromicity of 58% in the absorption spectrum, the induced CD of the porphyrin, and efficient energy transfer between TMPyP and K-free G4-wires suggest an intercalative mechanism of TMPyP binding. The K form interacts with TMPyP much weaker than the K-free form of the wires. Binding of TMPyP to the K form is characterized by a small (3 nm) shift of the Soret band, a weak positive induced CD in the Soret region, and the absence of energy transfer between the G-bases and the porphyrin. These parameters reflect a nonintercalative binding of TMPyP to the K form of the wires. We suggest that K ions positioned in the center space between the adjacent tetrads limit the access of TMPyP and other organic molecules to this region, thus enabling only nonintercalative modes of ligand binding to G-quadruplex DNAs.
    Biochemistry 12/2007; 46(45):12925-9. · 3.38 Impact Factor
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    ABSTRACT: Efficient attachment of DNA to metal surfaces or electrodes is essential for charge-transport measurements, scanning tunneling microscopy, and for devices and sensors. To optimize DNA deposition on Au-based surfaces and electrodes, we synthesized DNA with phosphorothioate (PT) groups attached to the G strand of poly(deoxyguanine)-poly(deoxycytosine) [poly(dG)-poly(dC)]. This procedure strongly improves the DNA anchoring to Au-based surfaces by sulfur-gold interaction. Much higher molecular surface density on Au substrates was observed for PT poly(dG)-poly(dC) compared to “bare” molecules. Deposition of PT poly(dG)-poly(dC) on Au-based electrodes, followed by thorough washing, showed that they specifically attach to the electrodes and are not spread on the surrounding SiO2 surface.
    Applied Physics Letters 10/2007; 91(17):173101-173101-3. · 3.79 Impact Factor
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    ABSTRACT: Double-stranded poly(dG)–poly(dC) and triple-stranded poly(dG)–poly(dG)–poly(dC) DNA were deposited on the modified surface of highly oriented pyrolitic graphite (HOPG) and visualized using atomic force microscopy with high-resolution (radius of ~1 nm) tips. The high resolution attained by this technique enabled us to detect single-stranded regions in double-stranded poly(dG)–poly(dC) and double-stranded and single-stranded regions in poly(dG)–poly(dG)–poly(dC) triplexes, as well as to resolve the helical pitch of the triplex molecules. We could also follow the reaction of G-strand extension in poly(dG)–poly(dC) by the Klenow exo− fragment of DNA polymerase I. This approach to molecular visualization could serve as a useful tool for the investigation of irregular structures in canonical DNA and other biopolymers, as well as studies of the molecular mechanisms of DNA replication and transcription.
    Nanotechnology 05/2007; 18(22):225102. · 3.84 Impact Factor
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    ABSTRACT: G4-DNA, a quadruple helical motif of stacked guanine tetrads, is stiffer and more resistant to surface forces than double-stranded DNA (dsDNA), yet it enables self-assembly. Therefore, it is more likely to enable charge transport upon deposition on hard supports. We report clear evidence of polarizability of long G4-DNA molecules measured by electrostatic force microscopy, while coadsorbed dsDNA molecules on mica are electrically silent. This is another sign that G4-DNA is potentially better than dsDNA as a conducting molecular wire.
    Nano Letters 05/2007; 7(4):981-6. · 13.03 Impact Factor
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    ABSTRACT: High-resolution scanning tunneling microscopy (STM) imaging of single double-stranded poly(G)-poly(C) DNA molecules, made by a novel synthesis method, shows the molecules morphology. The STM images reveal a periodic structure of approximately 4 nm, seen as repeating "bulbs" along the molecules. These "bulbs" are associated with the molecule helix (the major grooves). "Nicks", two per 100 nm on the average, are observed along the DNA as well. The STM measurements were supported by a morphological statistics of the DNA molecule groove length and apparent height relative to the surface.
    The Journal of Physical Chemistry B 04/2006; 110(9):4430-3. · 3.61 Impact Factor
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    ABSTRACT: Various DNA-based structures (single-, double-, triple-stranded and quadruplex-DNA) were characterized using non-contact atomic-force microscopy on two substrates: modified highly-oriented pyrolitic graphite (HOPG) and mica. Deposition on mica, a conventional substrate used in studies of bio-molecules, results in strong deformation of all above types of molecules while deposition on modified HOPG affects the morphology of DNA much less compared to mica. This is demonstrated by a larger measured height of the DNA molecules deposited on HOPG, as compared to mica, and an increased flexibility of the molecules, evidenced by a shorter molecular end-to-end distance on HOPG. The estimated heights of the triplex and the quadruplex DNA measured on HOPG are similar to the diameter of these molecules in liquid. We thus conclude that modified HOPG is a substrate more suitable than mica for AFM characterization of DNA morphology.
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    ABSTRACT: The photoinduced covalent redox label 8-thiouredopyrene-1,3,6-trisulfonate (TUPS) has been attached to two lysine residues (K8 and K39) at opposite sides of horse heart cytochrome c, as well as to cysteines, at the same positions, introduced by site-directed mutagenesis. Electron transfer between TUPS and the heme of cytochrome c deviates from the expected monoexponential kinetic behavior. Neither the overall rate nor the individual exponential components of electron transfer, as followed by kinetic absorption spectroscopy, correlate with the length of the covalent link connecting the dye with the protein. Molecular dynamics calculations show that TUPS can approach the protein surface and occupy several such positions. This heterogeneity may explain the multiexponential electron-transfer kinetics. The calculated optimal electron-transfer pathways do not follow the covalent link but involve through space jumps from the dye to the protein moiety, effectively decoupling the length of the covalent link and the electron-transfer rates.
    Journal of Chemical Information and Modeling 10/2005; 45(6):1520-6. · 4.30 Impact Factor
  • Advanced Materials 06/2005; 17(15):1901 - 1905. · 14.83 Impact Factor
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    ABSTRACT: In this paper, we describe a production procedure of the one-to-one double helical complex of poly(dG)-poly(dC), characterized by a well-defined length (up to 10 kb) and narrow size distribution of molecules. Direct evidence of strands slippage during poly(dG)-poly(dC) synthesis by Klenow exo(-) fragment of polymerase I is obtained by fluorescence resonance energy transfer (FRET). We show that the polymer extension results in an increase in the separation distance between fluorescent dyes attached to 5' ends of the strands in time and, as a result, losing communication between the dyes via FRET. Analysis of the products of the early steps of the synthesis by high-performance liquid chromatography and mass spectroscopy suggest that only one nucleotide is added to each of the strand composing poly(dG)-poly(dC) in the elementary step of the polymer extension. We show that proper pairing of a base at the 3' end of the primer strand with a base in sequence of the template strand is required for initiation of the synthesis. If the 3' end nucleotide in either poly(dG) or poly(dC) strand is substituted for A, the polymer does not grow. Introduction of the T-nucleotide into the complementary strand to permit pairing with A-nucleotide results in the restoration of the synthesis. The data reported here correspond with a slippage model of replication, which includes the formation of loops on the 3' ends of both strands composing poly(dG)-poly(dC) and their migration over long-molecular distances (microm) to 5' ends of the strands.
    Nucleic Acids Research 02/2005; 33(2):525-35. · 8.28 Impact Factor
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    ABSTRACT: The extension of the G-strand of long (700 bp) poly(dG)-poly(dC) by the Klenow exo(-) fragment of DNA polymerase I yields a complete triplex structure of the H-DNA type. High-performance liquid chromatography analysis demonstrates that the length of the G-strand is doubled during the polymerase synthesis. Fluorescence resonance energy transfer analysis shows that the 5' ends of the G- and the C-strands, labeled with fluorescein and TAMRA, respectively, are positioned close to each other in the product of the synthesis. Atomic force microscopy morphology imaging shows that the synthesized structures lack single-stranded fragments and have approximately the same length as the parent 700 bp poly(dG)-poly(dC). CD spectrum of the polymer has a large negative peak at 278 nm, which is characteristic of the poly(dG)-poly(dG)-poly(dC) triplex. The polymer is resistant to DNase and interacts much more weakly with ethidium bromide as compared with the double-stranded DNA.
    Nucleic Acids Research 02/2005; 33(20):6515-21. · 8.28 Impact Factor
  • Advanced Materials - ADVAN MATER. 01/2005; 17(15).
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    ABSTRACT: 1-Thiouredopyrene-3,6,8-trisulfonate (TUPS) has recently been used as a photoinduced covalent redox label capable of reducing various cofactors of proteins. A new reaction of this dye, whereby its excited triplet state oxidizes suitable electron donors, is now reported. The characteristic difference spectrum of the reduced radical of TUPS is determined. We also observe the self-exchange electron transfer between two TUPS molecules in their triplet excited states and determine the reaction scheme and the rate constants of the various pathways in the process of triplet depletion. The ability of photoexcited TUPS to withdraw an electron from reduced cytochrome-c is also observed. It is thus demonstrated that TUPS is an appropriate photoinduced covalent redox label for initiating both the oxidative and reductive phases of electron transfer processes in biological macromolecules.
    Photochemistry and Photobiology 07/2004; 79(6):489-93. · 2.29 Impact Factor
  • Alexander B Kotlyar, Natalia Borovok
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    ABSTRACT: Tightly coupled inside-out vesicles were prepared from Paracoccus denitrificans cells (SPP, sub-Paracoccus particles) and characterized kinetically. The rate of NADH oxidation, catalysed by SPP, increases 6-8 times on addition of gramicidin. The vesicles are capable of catalysing Delta micro H+-dependent reverse electron transfer from quinol to NAD+. The kinetic parameters of the NADH-oxidase and the reverse electron transfer carried out by membrane-bound P. denitrificans complex I were estimated and compared with those of the mitochondrial enzyme. The data demonstrate that catalytic properties of the dinucleotide-binding site of the bacterial and mitochondrial complex I are almost identical, pointing out similar organization of the site in mammals and P. denitrificans. Inhibition of the bacterial complex I by a specific inhibitor of Q reduction, rotenone, is very different from that of the mitochondrial enzyme. The inhibitor is capable of suppressing the NADH oxidation reaction only at micromolar concentrations, while the activity of mitochondrial enzyme is suppressed by nanomolar concentrations of rotenone. In contrast to the mitochondrial enzyme, rotenone, even at concentrations as high as 10 micro m, does not inhibit the reverse, Delta micro H+-dependent NAD+-reductase reaction on SPP.
    European Journal of Biochemistry 09/2002; 269(16):4020-4. · 3.58 Impact Factor

Publication Stats

184 Citations
127.40 Total Impact Points


  • 1995–2012
    • Tel Aviv University
      • • Department of Chemistry
      • • Center for Nanoscience and Nanotechnology
      Tell Afif, Tel Aviv, Israel
  • 2006–2007
    • Hebrew University of Jerusalem
      • Department of Physical Chemistry
      Jerusalem, Jerusalem District, Israel