Studies of nucleic acid structure, stability, and recognition using transition metal oxidants

Redox reactions of metal complexes Ru(tpy)(bpy)O{dollar}sp{lcub}2+{rcub}{dollar} (bpy = 2,2{dollar}spprime{dollar}-bipyridine; tpy = 2,2{dollar}spprime{dollar},2{dollar}sp{lcub}primeprime{rcub}{dollar}-terpyridine) and Pt{dollar}sb2{dollar}(pop){dollar}sb4sp{lcub}4-{rcub}{dollar} (pop = P{dollar}sb2{dollar}O{dollar}sb5{dollar}H{dollar}sb2sp{lcub}2-{rcub}{dollar}) with nucleic acids were used to obtain information on oligonucleotide structures, thermal stabilities, protein binding sites, ion binding affinities, and structure-selective reaction rates.; Pt{dollar}sb2{dollar}(pop){dollar}sb4sp{lcub}4-{rcub}{dollar} oxidizes the ribose moiety of oligodeoxynucleotides in single-stranded regions more efficiently than in double- or triple-stranded regions. Thermal transitions for duplex DNA and triplex DNA were observed in Chapter 1 by plotting the cleavage intensity for the region of interest as a function of reaction temperature. This approach resolved a transition that could not be resolved with absorbance. In addition, Pt{dollar}sb2{dollar}(pop){dollar}sb4sp{lcub}4-{rcub}{dollar} was used to photofootrprint the basic fibroblast growth factor protein on its DNA aptamer and confirms the aptamer secondary structure in Chapter 2.; The extent of guanine oxidation by Ru(tpy)(bpy)O{dollar}sp{lcub}2+{rcub}{dollar} was determined in Chapter 3 for four derivatives of the iron responsive element (IRE). Oxidation patterns obtained for the loop region of the RNA derivatives were consistent with their known secondary structures and were dissimilar to two DNA analogs. Band-shift studies using rabbit reticulocyte lysates showed that the RNA oligomer but not the DNA analog bound the IRE binding protein.; Divalent metal ion interactions with DNA were explored using both Pt{dollar}sb2{dollar}(pop){dollar}sb4sp{lcub}4-{rcub}{dollar} and Ru(tpy)(bpy)O{dollar}sp{lcub}2+{rcub}.{dollar} Inhibition of DNA cleavage by Ru(tpy)(bpy)O{dollar}sp{lcub}2+{rcub}{dollar} was observed for Mn{dollar}sp{lcub}2+{rcub}{dollar}, Mg{dollar}sp{lcub}2+{rcub}{dollar}, Ni{dollar}sp{lcub}2+{rcub}{dollar}, and Co{dollar}sp{lcub}2+{rcub}{dollar} in Chapter 4. The reaction of Mn{dollar}sp{lcub}2+{rcub}{dollar} with Ru(tpy)(bpy)O{dollar}sp{lcub}2+{rcub}{dollar} produced a transient intermediate at 531 nm and a second-order rate constant of 1300 {dollar}pm{dollar} 200 M{dollar}sp{lcub}-1{rcub}{dollar}s{dollar}sp{lcub}-1{rcub}{dollar}, which is fast enough to inhibit guanine oxidation. Mg{dollar}sp{lcub}2+{rcub}{dollar}, Ni{dollar}sp{lcub}2+{rcub}{dollar}, and Co{dollar}sp{lcub}2+{rcub}{dollar}, which do not react with Ru(tpy)(bpy)O{dollar}sp{lcub}2+{rcub}{dollar}, inhibit oxidation only in the double-stranded region, while Mn{dollar}sp{lcub}2+{rcub}{dollar} was a better inhibitor in the single-stranded regions. Phosphorescence emission quenching by Pt{dollar}sb2{dollar}(pop){dollar}sb4sp{lcub}4-{rcub}{dollar} was used in Chapter 5 to measure the binding affinity of Mn{dollar}sp{lcub}2+{rcub}{dollar} to calf thymus DNA. Application of polyelectrolyte theory resulted in an effective charge for Mn{dollar}sp{lcub}2+{rcub}{dollar} of 1.23 {dollar}pm{dollar} 0.12 and {dollar}Delta{dollar}G{dollar}spcircsb{lcub}rm t{rcub}{dollar} = 3.6 kcal/mol.