The Sizeof the Internal Loop in DNA Hairpins InfluencesTheir Targeting with Partially Complementary Strands

摘要

Targeting of noncanonical DNA structures, such as hairpin loops, may have significant diagnostic and therapeutic potential. Oligonucleotides can be used for binding to mRNA, forming a DNA/RNA hybrid duplex that inhibits translation. This kind of modulation of gene expression is called the antisense approach. In order to determine the best strategy to target a common structural motif in mRNA, we have designed a set of stem-loop DNA molecules with sequence: d(GCGC>T>nGTAAT5GTTAC>T>nGCGC), where n = 1, 3, or 5, “T5” is an end loop of five thymines. We used a combination of calorimetric and spectroscopy techniques to determine the thermodynamics for the reaction of a set of hairpins containing internal loops with their respective partially complementary strands. Our aim was to determine if internal- and end-loops are promising regions for targeting with their corresponding complementary strands. Indeed, all targeting reactions were accompanied by negative changes in free energy, indicating that reactions proceed spontaneously. Further investigation showed that these negative free energy terms result from a net balance of unfavorableentropy and favorable enthalpy contributions. In particular, unfoldingof hairpins and duplexes is accompanied by positive changes in heatcapacity, which may be a result of exposure of hydrophobic groupsto the solvent. This study provides a new method for the targetingof mRNA in order to control gene expression.