title: Gapped DNA is anisotropically bent.
abstract: Ionizing radiation damages DNA in several ways, including through formation of a single-nucleoside gap in one DNA strand. We have developed a two-dimensional gel electrophoresis method to investigate the effect of a strand gap on DNA structure. We generate a library of gapped DNA molecules by treating a DNA restriction fragment with the hydroxyl radical, generated by the reaction of Fe(II) EDTA with hydrogen peroxide. The DNA molecule studied contains a fixed bend produced by a set of phased adenine tracts. The A-tract bend serves as a reference bend for investigating the conformational nature of a strand gap. In the first electrophoretic dimension, a bent DNA molecule that has been treated with the hydroxyl radical is electrophoresed on a native gel. Smearing of the band on the native gel indicates that the library of gapped DNA molecules contains a variety of DNA conformations. In the second electrophoretic dimension, gapped DNA molecules having different native gel mobilities are electrophoresed on separate lanes of a denaturing gel to reveal how each strand gap affects the native gel mobility (and thus shape) of the DNA. Our results demonstrate that a single-nucleoside gap in a DNA duplex leads to an anisotropic, directional bend in the DNA helix axis. The implications of our findings for recognition of this lesion by DNA repair proteins are discussed.
authors.affiliation: Protometrix, Inc., 66 High Street, Guilford, CT 06437, USA.
authors.forename: Hong
authors.initials: H
authors.lastname: Guo
authors.forename: Thomas D
authors.initials: TD
authors.lastname: Tullius
mesh_terms: D016880:Anisotropy; D001483:Base Sequence; D004247:DNA / Q000737:chemistry*; D015180:Electrophoresis, Gel, Two-Dimensional; D006878:Hydroxides; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D010957:Plasmids / Q000737:chemistry*; D011089:Polydeoxyribonucleotides / Q000737:chemistry*
chemical_list: D006878:Hydroxides; D011089:Polydeoxyribonucleotides; D004247:DNA; C031356:hydroxide ion