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Suppression of DNA-Mediated Charge Transport by BamHI Binding

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Presentation on theme: "Suppression of DNA-Mediated Charge Transport by BamHI Binding"— Presentation transcript:

1 Suppression of DNA-Mediated Charge Transport by BamHI Binding
Kazuhiko Nakatani, Chikara Dohno, Atsushi Ogawa, Isao Saito  Chemistry & Biology  Volume 9, Issue 3, Pages (March 2002) DOI: /S (02)

2 Figure 1 Hydrogen Bonding Contacts between BamHI and G-C Base Pairs in the Recognition Sequence of 5′-GGATCC-3′ and the Effect of Hydrogen Bonding on the Electrostatic Potential Atomic coordinates were taken from the X-ray structure of the BamHI-DNA complex reported by Newman et al. [20]. (A) The protein-DNA interactions involving 5′- and 3′-side G-C base pairs are colored in yellow and blue, respectively. The N7 and O6 atoms of 5′-side G are directly hydrogen bonded to an Arg155 guanidium group that is also bound to Glu161. The O6 of 3′-side G bound to Asn116. For clarity, a hydrogen bonding network involving 5′ side G-C base pair is shown with a dotted line. (B) The electrostatic potential mapped on the surface of a total electron density (0.002 electrons/au3) for free guanine (left) and the guanine-guanidium (Arg155)-carboxylate (Glu161) triad (right). The electron-rich sites are shown in orange, whereas electron-deficient sites are shown in blue. Chemistry & Biology 2002 9, DOI: ( /S (02) )

3 Figure 2 Sequences of Oligomer Duplexes 1/2 and 3/4 Both Containing a BamHI Recognition Sequence, 5′-GGATCC-3′ The modified base (X) denotes p-cyanobenzophenone-substituted 2′-deoxyuridine (CNBPU). G8 and a hole-trapping G triplet (G24G25G26) shown in ODN 3 are located on opposite sides of the BamHI binding site. The BamHI binding site, a modified base, G8, and a G triplet are shown in bold face. Chemistry & Biology 2002 9, DOI: ( /S (02) )

4 Figure 3 Binding of BamHI to ODN 1 and Its Effect on the One-Electron Oxidation of G's (A) An electrophoretic mobility shift assay to detect a BamHI-DNA complex formation [25, 26]. Lane 1, DNA only; lane 2, BamHI, 0.15 U/μl; lane 3, 0.3 U/μl; lane 4, 0.6 U/μl; lane 5, 1.2 U/μl; lane 6, 2.4 U/μl. (B) An autoradiogram of a denaturing polyacrylamide gel for BamHI concentration-dependent photooxidation of duplex 1/2. Lane 1, DNA only; lane 2, 0.15 U/μl; lane 3, 0.3 U/μl, lane 4, 1.2 U/μl; lane 5, Maxam-Gilbert G + A sequencing reactions. The BamHI site and CNBPU (X) are shown in bold face. Chemistry & Biology 2002 9, DOI: ( /S (02) )

5 Figure 4 Autoradiograms of a Denaturing Polyacrylamide Gel for Photooxidation of Duplex 1/2 in the Presence of BamHI ODNs 1 (A) and 2 (B) were separately 5′-32P-end labeled and hybridized to a nonlabeled complementary strand. Lane 1, Maxam-Gilbert G + A sequencing reactions; lane 2, in the absence of BamHI; lanes 3–5, BamHI. ODNs in lanes 2–4 were irradiated at 312 nm. All samples except that in lane 2 were heated with piperidine. The BamHI site and CNBPU (X) are shown in bold face. For clarity, the autoradiogram for ODN 2 was shown upsidedown. Chemistry & Biology 2002 9, DOI: ( /S (02) )

6 Figure 5 An Autoradiogram of a Denaturing Polyacrylamide Gel for Photooxidation of Duplex 3/4 in the Presence of BamHI Lanes 1–3, in the presence of BamHI; lane 4, in the absence of BamHI; lane 5, Maxam-Gilbert G + A sequencing reactions. ODNs in lanes 2–4 were irradiated. All samples except that in lane 2 were heated with piperidine. Chemistry & Biology 2002 9, DOI: ( /S (02) )

7 Figure 6 Illustrations of the Charge Transport in Duplex 3/4 in the Absence and Presence of BamHI (A) In the absense of BamHI. (B) In the presence of BamHI. Horizontal arrows and the numbers shown on the site of guanine oxidation indicate the band intensity relative to that of G24 in the protein bound duplex. Band intensities are determined by densitometry of the lanes 3 (Figure 5A) and 4 (Figure 5B) in Figure 5. Chemistry & Biology 2002 9, DOI: ( /S (02) )


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