The octahedral rhodium complex, cis-dichloro bis(1,10 phenanthroline)rhodium(III) chloride (BISPHEN), is known to form covalent linkages with DNA involving the attachment of the metal to a base. In order to determine the sequence selectivity of this chemistry, solutions of the complex containing one of the double-stranded DNA plasmids, pBSSK.c-raf (eco) or pBSKS .XE.LTR-F (a construct that contains sequences derived from the long terminal repeat [LTR] region of the human immunodeficiency virus) have been irradiated using UVA light. The DNA samples were denatured after irradiation, a primer was annealed to one of the strands, and a complementary strand was constructed using a polymerase enzyme. Polyacrylamide gel sequencing analysis was used to reveal stops created in the complementary strand caused by the polymerase encountering a metal-bound base. The data indicate that “hot spots” primarily occur at, or adjacent to, guanines (G), with a particularly strong preference for strings of G. In the latter case, the hottest spot is at the 5′G. These results are consistent with our previously postulated mechanism for the covalent binding chemistry which involves photooxidation of deoxyguanosine by the excited state of the metal complex as the primary photochemical step.
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Vol. 72 • No. 6