Two conserved structural components, A-rich bulge and P4 XJ6/7 base-triples, in activating the group I ribozymes

Background: The A-rich bulge of the group I intron ribozyme, a highly conserved structural element in its P5 peripheral region, plays a significant role in activating the ribozyme. The bulge has been known to interact with the P4 stem forming P4 XJ6/7 base-triples in the conserved core. The base-triples by themselves have also been identified as a distinctive element responsible for enhancing the activity of the ribozyme. Results: A weakly active variant of the Tetrahymena ribozyme lacking the P5 extension was dramatically activated by the addition of an A-rich bulge at the peripheral region, or by replacement of the original P4 XJ6/7 base-triples in the core structure with more stabilized isosteric ones. Biochemical analyses showed that the two methods of activation affect the ribozyme differently. Conclusions: The long-range interaction between the A-rich bulge and P4 or additionally stabilized P4 XJ6/7 base-triples can contribute dramatically to activation of the Tetrahymena ribozyme. Both improve the k_cat value, which represents the rate of the limiting step of the ribozyme reaction when its binding site is saturated with GTP. However, the bulge or the modified base-triples gave a moderate reduction or considerable increase, respectively, to the K_m(GTP) value.