A wide range of experimental methods have been used to explore the structure and function of the hairpin ribozyme. In vitro selection methods has provided comprehensive information on RNA secondary structure and the identity of essential bases within the two internal loops of the ribozyme-substrate complex. Crosslinking analysis has identified a tertiary structure motif within ribozyme loop B that is also found in rRNA. The ribozyme contains two domains that fold independently. A sharp bend at the junction between the two domains is necessary for the two domains to interact in the active complex. Biphasic kinetic behavior in pre-steady state cleavage kinetics results from partitioning of the ribozyme between the active, bent conformer, and an inactive conformer in which the two domains are coaxially stacked, thus preventing their interaction. Novel strategies have been used to reconstruct the ribozyme in such a way that folding into the active conformer is strongly favored.

Peer reviewed