Hammerhead Minizymes with High Cleavage Activity:  A Dimeric Structure as the Active Conformation of Minizymes

A number of minizymes with short oligonucleotide linkers (one to four bases) instead of stem-loop II were constructed. Only two of the minizymes examined had cleavage activity similar to that of the full-sized ribozyme. The minizymes with high activity formed dimeric structures with a common stem II. A significant contribution to stabilization of such a dimeric structure is probably made by nonstandard base pairing and a Mg2+ ion in both catalytic cores of the dimer. Interaction with the substrate also stabilized the dimeric structures. Thus, active minizymes can be designed with a self-complementary linker sequence with G and C on 5‘ and 3‘ sides, respectively, of the linker which induces the intermolecular interactions that lead to formation of active dimers.