The hammerhead ribozyme reaction is more complex than might have been expected, perhaps because of the flexibility of RNA, which would have enhanced the potential of RNA during evolution of and in the RNA world. Divalent Mg2+ ions can increase the rate of the ribozyme-catalyzed reaction by approximately 10(9)-fold as compared to the background rate under standard conditions. However, the role of Mg2+ ions is controversial since the reaction can proceed in the presence of high concentrations of monovalent ions, such as Li+, Na+, and NH4+ ions, in the absence of divalent ions. We thus carried out ribozyme reactions under various conditions, and we obtained parameters that explain the experimental data. On the basis of the analysis, we propose a new pathway in the hammerhead ribozyme reaction in which divalent metal ions and monovalent ions act cooperatively.