Computer-Aided Calculation of the Local Folding Potential of Target RNA and Its Use for Ribozyme Design

Abstract

Ribozymes first bind their target via complementary sequences like a conventional antisense strand and, subsequently, catalyze the hydrolyses of the cleavable motif. In the case of hammerhead ribozymes, this is a specific base triplet. Even though it has been described that there are 12 conceivable motifs recognized by hammerhead ribozymes that are cleaved with different efficiencies (1,2), a typical long-chain target RNA provides a large number of potential cleavage sites. Given this flexibility, one important step in ribozyme design is the selection of an appropriate cleavable motif within the target. For ribozyme-mediated inhibition in living cells, the contribution of antisense effects and catalytic effects to the overall extent of inhibition remains unresolved, and may well be dependent on the specific constructs and biological systems. However, it is reasonable to assume that the first step, the binding of the ribozyme with its target, is crucial. Therefore, the local target sequence against which a ribozyme is directed does not only have to contain a cleavable triplet, but should also be accessible for the ribozyme.