BENDING OF DNA BY THE MITOMYCIN C-INDUCED, GPG INTRASTRAND CROSS-LINK

Mitomycin C (MC) forms interstrand and intrastrand cross-link adducts and monoalkylation products (monoadducts) with DNA. Each of the three types of adducts was incorporated site-specifically into both a 15-mer and a 21-mer oligodeoxyribonucleotide duplex. The adduct-containing d uplexes were P-32-phosphorylated and ligated to form multimers, which were then analyzed for anomalous electrophoretic mobility by nondenatu ring polyacrylamide gel electrophoresis, using the method of Koo and C rothers [(1988) Proc. Natl. Acad. Sci. U.S.A. 85, 1763-1767] in order to detect DNA curvature caused by the adducts. The intrastrand crossli nk adduct was found to induce a 14.6 +/- 2.0 degrees DNA bend per lesi on (minimum value) while no DNA bending was detected for either the in terstrand cross-link or the monoadduct. Molecular mechanics modeling i ndicated that the possible origin of the bend lies in a considerable d eviation from parallel of the normals to the best planes of the intras trand cross-linked guanines, due to a shorter than normal distance bet ween their N-2 atoms forced upon them by the cross-link. The observed bending by the MC intrastrand lesion may be the cause of the increased flexibility of MC-modified DNA, localized to distinct regions, as obs erved in earlier work by hydrodynamic methods and electron microscopy. The MC adduct-caused DNA bend may serve as a recognition site for cer tain DNA-binding proteins.