Variations in Stability of Polynucleotide Hairpins Due to Stem-Loop Conformation

Ultraviolet (UV) spectroscopy is used to examine the transition from hairpin conformation to random coil of self-complementary single stranded DNA (ssDNA) and RNA (ssRNA). The transition is driven by heating. The Melting Temperature (Tm) of a polynucleotide hairpin is the temperature at which half of the molecules are in the hairpin conformation and half in a random coil conformation. The hairpin is a secondary structure formed by a self-complementary single stranded nucleotide sequence and is composed of two parts, a stem and a loop. A longer stem contains more paired nucleotides, which stabilize the secondary structure, while a longer loop lowers the stress on the stem. Our results indicate that for a DNA sequence with constant stem nucleotide number and varying loop length from zero to three nucleotides, Tm increases significantly with the size of the loop. These results indicate that a nucleotide sequence in the hairpin conformation experiences a mechanical stress produced by the loop on the stem structure. This stress on the stem decreases the melting temperature of the structure. The goal of these experiments is to quantify this mechanical stress, in order to probe the mechanical response of DNA and RNA to sharp bends. Studies are now underway to examine the effect of varying stem lengths on the stability of DNA and RNA hairpins.