Phosphorothioates in pre-tRNAs can change the specificities of RNAses P or reduce the cleavage efficiencies.

Several phosphorothioate-modified E coli and yeast pre-tRNAs were synthesized. If this modification included the phosphodiester at the RNase P cleavage site, two different effects were observed. With some pre-tRNAs the RNase P cleavage efficiency was severely reduced, whereas with other pre-tRNAs a new reaction type for RNase P was observed. Unlike the previously studied base or ribose modifications, phosphorothioates resulted in aberrant cleavages at unmodified phosphodiesters. These new sites could be located in the 5'-flank or in the acceptor stem of the tRNA domain. Modified mutants of E coli pre-tRNA(Tyr) with different base pairs at the RNase P cleavage site were cleaved with reduced efficiencies, but no aberrant products were observed.