A deteriorated triple-helical scaffold accelerates formation of the Tetrahymena ribozyme active structure

The Tetrahymena group I ribozyme requires a hierarchical folding process to form its correct three-dimensional structure. Ribozyme activity depends on the catalytic core consisting of two domains, P4-P6 and P3-P7, connected b y a triple-helical scaffold. The folding proceeds in the following order: ( i) fast folding of the P4-P6 domain, (ii) slow folding of the P3-P7 domain, and (iii) structure rearrangement to form the active ribozyme structure. T he third step is believed to directly determine the conformation of the act ive catalytic domain, but as yet the precise mechanisms remain to de elucid ated. To investigate the folding kinetics of this step, we analyzed mutant ribozymes having base substitution(s) in the triple-helical scaffold and fo und that disruption of the scaffold at A105G results in modest slowing of t he P3-P7 folding (1.9-fold) and acceleration of step (iii) by 5.9-fold. The se results suggest that disruption or destabilization of the scaffold is a normal component in the formation process of the active structure of the wi ld type ribozyme. (C) 2001 Published by Elsevier Science B.V. on be half of the Federation of European Biochemical Societies.