Cellular Small Molecules Contribute to Twister Ribozyme Catalysis

The number of self-cleaving small ribozymes has increased sharply in recent years. Advances have been made in describing these ribozymes in terms of four catalytic strategies: α describes in-line attack, β describes neutralization of the nonbridging oxygens, γ describes activation of the nucleophile, and δ describes stabilization of the leaving group. Current literature presents the rapid self-cleavage of the twister ribozyme in terms of all four of these classic catalytic strategies. Herein, we describe the nonspecific contribution of small molecules to ribozyme catalysis. At biological pH, the rate of the wild-type twister ribozyme is enhanced up to 5-fold in the presence of moderate buffer concentrations, similar to the 3-5-fold effects reported previously for buffer catalysis for protein enzymes. We observe this catalytic enhancement not only with standard laboratory buffers, but also with diverse biological small molecules, including imidazole, amino acids, and amino sugars. Brønsted plots suggest that small molecules assist in proton transfer, most likely with δ catalysis. Cellular small molecules provide a simple way to overcome the limited functional diversity of RNA and have the potential to participate in the catalytic mechanisms of many ribozymes in vivo.