A circular trans-acting ribozyme designed to adopt the motif of the hepatitis delta virus (HDV) trans-acting ribozyme was produced. The circular form was generated in vitro by splicing a modified group I intron precursor RNA in which the relative order of the 5' and 3' splice sites, flanking the single HDV-like ribozyme sequence-containing exon, is reversed. Trans-cleavage activity of the circular HDV-like ribozyme was comparable to linear permutations of HDV ribozymes containing the same core sequence, and was shown not to be due to linear contaminants in the circular ribozyme preparation. In nuclear and cytoplasmic extracts from HeLa cells, the circular ribozyme had enhanced resistance to nuclease degradation relative to a linear form of the ribozyme, suggesting that circularization may be a viable alternative to chemical modification as a means of stabilizing ribozymes against nuclease degradation.