Fast folding of a ribozyme by stabilizing core interactions: Evidence for multiple folding pathways in RNA

Folding of the Tetrahymena ribozyme under physiological conditions in vitro is Limited by slow conversion of long-lived intermediates to the active st ructure. These intermediates arise because the most stable domain of the ri bozyme folds 10-50 times more rapidly than the core region containing helix P3. Native gel electrophoresis and time-resolved X-ray-dependent hydroxyl radical cleavage revealed that mutations that weaken peripheral interaction s between domains accelerated folding fivefold, while a point mutation that stabilizes P3 enabled 80 % of the mutant RNA to reach the native conformat ion within 30 seconds at 22 degrees C. The P3 mutation increased the foldin g rate of the catalytic core as much as 50-fold, so that both domains of th e ribozyme were formed at approximately the same rate. The results show tha t the ribozyme folds rapidly without significantly populating metastable in termediates when native interactions in the ribozyme core are stabilized re lative to peripheral structural elements. (C) 2000 Academic Press.