BASE-CATALYSIS OF IMINO PROTON-EXCHANGE IN DNA - EFFECTS OF CATALYST UPON DNA-STRUCTURE AND DYNAMICS

Characterization of the kinetics and energetics of base-pair opening i n nucleic acids relies upon measurements of the rates of exchange of i mino protons with water protons at high concentrations of the exchange catalyst. Under these conditions, the exchange catalyst may affect st ructural or dynamic properties of the nucleic acid molecule and thus, limit the significance of the exchange data. To address this problem, we have used NMR spectroscopy to characterize the effects of a catalys t of imino proton exchange, namely, ammonia upon the structure and dyn amics of the self-complementary DNA dodecamer [d(CGCAGATCTGCG)](2). Th e changes in structure were monitored in proton NOESY and DQF-COSY exp eriments and in phosphorus spectra at 15 degrees C and at ammonia conc entrations ranging from 0.002 to 0.5 M. The results indicate that ammo nia induces subtle chang es in the solution conformation of the dodeca mer, but the overall structure is maintained close to the B-type DNA s tructure. However, the relaxation rates (i.e., transverse, longitudina l, and cross relaxation rates) of several non-exchangeable protons wer e found to increase by similar to 50% upon changing ammonia concentrat ion from 0.002 to 0.5 M. The increases were comparable for all protons investigated suggesting that they originate from an ammonia-induced i ncrease in the overall correlation time of the DNA dodecamer. Numerica l analysis revealed that the catalyst-induced enhancements in proton r elaxation can alter significantly the calculated values of the exchang e rates of imino protons, especially those obtained from measurements of the line widths of these proton resonances.