Thermodynamic contributions of 5’- and 3’-single strand dangling-ends to the stability of short duplex DNAs

Abstract

Differential scanning calorimetry (DSC) melting analysis was performed on 27 short double stranded DNA duplexes containing 15 to 25 base pairs. Experimental duplexes were divided into two categories containing either two 5’ dangling-ends or one 5’ and one 3’ dangling-end. Duplex regions were incrementally reduced from 25 to 15 base pairs with a concurrent increase in length of dangling-ends from 1 to 10 bases. Blunt-ended duplexes from 15 to 25 base pairs served as controls. An additional set of molecules containing 21 base pair duplexes and a single four base dangling-end were also examined. DSC melting curves were measured in varying concentrations of sodium ion (Na + ). From these measurements, thermodynamic parameters for 5’ and 3’ dangling-ends were evaluated as a function of dangling end length. 5’ ends were found to be slightly stabilizing but essentially constant while the 3’ ends were destabilizing with increasing length of the dangling-end. 3’ ends also display a stronger dependence on Na + concentration. In lower Na+ environment, the 3’ ends were more destabilizing than in higher salt environment suggesting a more significant electrostatic component of the destabilizing interactions. Analysis of thermodynamic parameters of dangling ended duplexes as a function of Na + concentration indicated the 3' dangling ends behave differently than 5' dangling ended and blunt-ended duplexes. Molecules with one 5' and one 3' dangling end showed variation in excess specific heat capacity (ΔC p ) when compared to the blunt-ended molecule, while the molecules with two 5’ ends had ΔC p values that were essentially the same as blunt-ended duplexes. These observations suggested differences exist in duplexes with 3’ and 5’ dangling ends, which are interpreted in terms of composite differences in interactions with Na + , solvent, and terminal base pairs of the duplex.