A Thermodynamic Approach to Control Gene Expression: Targeting of
DNA Intramolecular Structures with Complementary Strands

Dr. Luis A. Marky, PhD
Pharmaceutical Sciences, Biochemistry & Molecular Biology and
Eppley Institute for Cancer Research
University of Nebraska Medical Center

Antisense, antigene and siRNA strategies are currently used to control the expression of genes. To this end, our laboratory is currently mimicking the targeting of mRNA by targeting DNA hairpin loops with their partially complementary strands. We use a combination of differential scanning calorimetry (DSC) and UV melting techniques to investigate the reaction of a variety of DNA hairpin loops (single end loops, dumbbell, threeway junction with two loops and a hairpin with a long bulge) with single strands that are complementary to the bases in the loop and to one strand of their stem, forming duplex products with dangling ends, nicks or with a displaced strand. We determine standard thermodynamic profiles for the unfolding of the hairpin loop and duplex product of each reaction.

The DSC and UV melts show monophasic transitions for all DNA single hairpin loops (reactants) and biphasic transitions for the double hairpin loops (reactants) and duplex products. The resulting DSC data is then used to create thermodynamic (Hess) cycles that correspond to each targeting reaction.

All targeting reactions yielded favorable ΔG contributions that were enthalpy driven. These favorable heat contributions result from the formation of basepair stacks involving the unpaired bases of the loops.