FORMATION OF A CATALYTICALLY ACTIVE DIMER BY TRNA(VAL)-DRIVEN SHORT RIBOZYMES

A minizyme is a hammerhead ribozyme with a short oligonucleotide linke r instead of stem/loop II. Minizymes with low activity as monomers for m active dimeric structures with a common stem. We explored the use of dimeric minizymes as gene-inactivating agents by placing minizymes un der the control of a tRNA(Val) promoter. The tRNA(Val) portion of the transcript did not hinder dimerization as the tRNA-embedded minizyme f ormed an active dimeric structure. The cleavage activity of this miniz yme that had been expressed either in vitro or in HeLa cells was almos t one order of magnitude higher than that of the tRNA(Val)-embedded co nventional hammerhead ribozyme. The tRNA(Val)-driven minizyme inhibite d reporter gene activity (95%) whereas the tRNA(Val)-driven hammerhead ribozyme resulted in approximately 55% inhibition.