Next, we tested the ability of the chemical reaction cycle to induce the folding of the hairpin. We added 20 mM EDC to 0.2 µM H-1 (i.e., a 100.000-fold excess) and monitored the fluorescence intensity of the molecular beacon as a function of time (Fig. ). The experiment was performed at the Tm of H-1 (30 °C), and we can thus assume that roughly 50% of the hairpin was in its folded state before we initiated the experiment. Immediately after the addition of the EDC, the fluorescence intensity dropped drastically, indicating that the population of folded hairpins had increased. Ten minutes after the EDC addition, we added 10 mM N-acetyl aspartate (Ac-D-OH) as a scavenger, i.e., a competing dicarboxylate that can also cyclically react with EDC and rapidly consumes all fuel. Consequently, the hairpin fluorescence returned to a value close to the original level. We attribute the lack of a complete return due to the change in ionic strength of solution caused by the positively charged group present in both EDC and its byproduct of hydration, i.e., the waste EDU (Fig. ).