The Effect of Site-Specific Modifications of DNA on Thermodynamic Stability, Ion Binding and Hydration

Abstract

Cations, which associate with DNA in both the major and minor grooves, play a significant role in determining DNA conformation. In the major groove, cations are associated with the N7/O6 edge of guanines, while in the minor groove they are found at A-T pairs. Both G-C and A-T have potential cation binding sites that when modified should result in the reorganization of salts and water, which in turn would affect local conformation and stability. We report herein the biophysical characterization of DNA duplexes in which we altered the N-7 position in the major groove of purines (7-deaza-guanine, 7-aminomethyl-7-deazaguanine, 7-hydroxymethyl-7-deazaguanine and 7-deaza-adenine) and at N-3 position of adenine in the minor groove (3-deazaadenine and 3-methyl-3-deazaadenine). These modifications alter the electronic properties of the heterocyclic bases and specifically eliminate DNA cation binding sites in the different grooves, or in the case of 7-aminomethyl-7-deazaguanine places a cationic group in the major groove at the edge of a G-C pair. The low temperature NMR and x-ray crystal structures of some of these DNA appear identical to unmodified DNA; however, the thermodynamic analyses show that these modified bases have a significant impact on the dynamic structure of DNA. In most cases, a reduction in thermodynamic stability driven by enthalpy changes was observed. The only modification that is thermodynamically as, or more, stable than the corresponding unmodified DNA is the 7-aminomethyl-7-deaza-guanine. The thermodynamic effects of the different substitutions are associated with the folding enthalpies and hydration. Interpretation of how these base modifications affect DNA structure and stability will be discussed.Supported by RO1CA29088 from NIH and MCB-0315746 from NSF.