Limits...
Zn2+ selectively stabilizes FdU-substituted DNA through a unique major groove binding motif.

Ghosh S, Salsbury FR, Horita DA, Gmeiner WH - Nucleic Acids Res. (2011)

Bottom Line: Mg(2+) neither inhibited EtBr complexation nor had as strong of a stabilizing effect.DNA sequences that did not contain consecutive FdU were not stabilized by Zn(2+).A lipofectamine preparation of the Zn(2+)-DNA complex displayed enhanced cytotoxicity toward prostate cancer cells relative to the individual components prepared as lipofectamine complexes indicating the potential utility of Zn(2+)-DNA complexes for cancer treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Biology, Program in Molecular Genetics, Wake Forest University School of Medicine, Winston-Salem, NC 27157, USA.

ABSTRACT
We report, based on semi-empirical calculations, that Zn(2+) binds duplex DNA containing consecutive FdU-dA base pairs in the major groove with distorted trigonal bipyramidal geometry. In this previously uncharacterized binding motif, O4 and F5 on consecutive FdU are axial ligands while three water molecules complete the coordination sphere. NMR spectroscopy confirmed Zn(2+) complexation occurred with maintenance of base pairing while a slight hypsochromic shift in circular dichroism (CD) spectra indicated moderate structural distortion relative to B-form DNA. Zn(2+) complexation inhibited ethidium bromide (EtBr) intercalation and stabilized FdU-substituted duplex DNA (ΔT(m) > 15 °C). Mg(2+) neither inhibited EtBr complexation nor had as strong of a stabilizing effect. DNA sequences that did not contain consecutive FdU were not stabilized by Zn(2+). A lipofectamine preparation of the Zn(2+)-DNA complex displayed enhanced cytotoxicity toward prostate cancer cells relative to the individual components prepared as lipofectamine complexes indicating the potential utility of Zn(2+)-DNA complexes for cancer treatment.

Show MeSH

Related in: MedlinePlus

MTS assay results demonstrating the cytotoxicity of Zn2+, the 3′-FdU DNA hairpin and the Zn2+ complex of the 3′-FdU hairpin, towards PC3 prostate cancer cells. The Zn2+–DNA complex (mDNA) displays the greatest cytotoxicity.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3105383&req=5

Figure 6: MTS assay results demonstrating the cytotoxicity of Zn2+, the 3′-FdU DNA hairpin and the Zn2+ complex of the 3′-FdU hairpin, towards PC3 prostate cancer cells. The Zn2+–DNA complex (mDNA) displays the greatest cytotoxicity.

Mentions: Although normal prostate tissue has the highest Zn2+ concentration in the human body, prostate cancer (PCa) cells have extremely low intracellular Zn2+ and Zn2+ is highly cytotoxic to PCa cells (15). Thus, Zn2+ complexes of FdU-containing DNA sequences may be cytotoxic to PCa cells both from the intracellular release of Zn2+ and from the release of the cytotoxic nucleotide analog FdUMP (6,7). The 3′-FdU DNA hairpin was prepared with and without Zn2+ using lipofectamine to promote cellular internalization. Cytotoxicity was evaluated using a modified clonogenic assay with an MTS readout. The results are shown in Figure 6. As is evident from the data, the 3′-FdU hairpin sequence is cytotoxic toward PCa cells. The lipofectamine complex containing Zn2+, but did not include the FdU-substituted DNA, was also cytotoxic toward PCa cells. The cytotoxicity of the Zn2+–DNA complex prepared with lipofectamine displayed the greatest cytotoxicity toward PC3 cells (Figure 6). All samples were prepared under identical conditions. We conclude that inclusion of Zn2+ in lipofectamine complexes of FdU-containing DNA sequences enhances the cytotoxicity of FdU-substituted DNA toward prostate cancer cells.Figure 6.


Zn2+ selectively stabilizes FdU-substituted DNA through a unique major groove binding motif.

Ghosh S, Salsbury FR, Horita DA, Gmeiner WH - Nucleic Acids Res. (2011)

MTS assay results demonstrating the cytotoxicity of Zn2+, the 3′-FdU DNA hairpin and the Zn2+ complex of the 3′-FdU hairpin, towards PC3 prostate cancer cells. The Zn2+–DNA complex (mDNA) displays the greatest cytotoxicity.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3105383&req=5

Figure 6: MTS assay results demonstrating the cytotoxicity of Zn2+, the 3′-FdU DNA hairpin and the Zn2+ complex of the 3′-FdU hairpin, towards PC3 prostate cancer cells. The Zn2+–DNA complex (mDNA) displays the greatest cytotoxicity.
Mentions: Although normal prostate tissue has the highest Zn2+ concentration in the human body, prostate cancer (PCa) cells have extremely low intracellular Zn2+ and Zn2+ is highly cytotoxic to PCa cells (15). Thus, Zn2+ complexes of FdU-containing DNA sequences may be cytotoxic to PCa cells both from the intracellular release of Zn2+ and from the release of the cytotoxic nucleotide analog FdUMP (6,7). The 3′-FdU DNA hairpin was prepared with and without Zn2+ using lipofectamine to promote cellular internalization. Cytotoxicity was evaluated using a modified clonogenic assay with an MTS readout. The results are shown in Figure 6. As is evident from the data, the 3′-FdU hairpin sequence is cytotoxic toward PCa cells. The lipofectamine complex containing Zn2+, but did not include the FdU-substituted DNA, was also cytotoxic toward PCa cells. The cytotoxicity of the Zn2+–DNA complex prepared with lipofectamine displayed the greatest cytotoxicity toward PC3 cells (Figure 6). All samples were prepared under identical conditions. We conclude that inclusion of Zn2+ in lipofectamine complexes of FdU-containing DNA sequences enhances the cytotoxicity of FdU-substituted DNA toward prostate cancer cells.Figure 6.

Bottom Line: Mg(2+) neither inhibited EtBr complexation nor had as strong of a stabilizing effect.DNA sequences that did not contain consecutive FdU were not stabilized by Zn(2+).A lipofectamine preparation of the Zn(2+)-DNA complex displayed enhanced cytotoxicity toward prostate cancer cells relative to the individual components prepared as lipofectamine complexes indicating the potential utility of Zn(2+)-DNA complexes for cancer treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Biology, Program in Molecular Genetics, Wake Forest University School of Medicine, Winston-Salem, NC 27157, USA.

ABSTRACT
We report, based on semi-empirical calculations, that Zn(2+) binds duplex DNA containing consecutive FdU-dA base pairs in the major groove with distorted trigonal bipyramidal geometry. In this previously uncharacterized binding motif, O4 and F5 on consecutive FdU are axial ligands while three water molecules complete the coordination sphere. NMR spectroscopy confirmed Zn(2+) complexation occurred with maintenance of base pairing while a slight hypsochromic shift in circular dichroism (CD) spectra indicated moderate structural distortion relative to B-form DNA. Zn(2+) complexation inhibited ethidium bromide (EtBr) intercalation and stabilized FdU-substituted duplex DNA (ΔT(m) > 15 °C). Mg(2+) neither inhibited EtBr complexation nor had as strong of a stabilizing effect. DNA sequences that did not contain consecutive FdU were not stabilized by Zn(2+). A lipofectamine preparation of the Zn(2+)-DNA complex displayed enhanced cytotoxicity toward prostate cancer cells relative to the individual components prepared as lipofectamine complexes indicating the potential utility of Zn(2+)-DNA complexes for cancer treatment.

Show MeSH
Related in: MedlinePlus