Optimal Distance between the Termini of Stem P3 and P4 in Kin.46 Kinase Ribozyme

摘要

The Kin.46 kinase ribozyme catalyzes the transfer of the thiophosphate from ATP-y-S to its own 5' hydroxyl end in the presence of oligonucleotide effector which is complementary to its 3' primer binding sequence (PBS) used in the amplification steps during the original selection for activity. Omitting the oligonucleotides reduces the observed catalytic rate constant (k_(obs)) by 10 to 10 -fold and addition of the deoxyoligonucleotide effector is necessary for its activity. The activator helix formed by the PBS and the oligo effector is connected by a 5nt "linker" region, J1/4, to the substrate-binding internal guide sequence, stem P1 and stabilizes the active conformation of the ribozyme by stabilizing the interaction between the linker, J1/4 and complementary nucleotides, J3/2, within the active site (Fig. 1).