FOLDING INTERMEDIATES OF A SELF-SPLICING RNA - MISPAIRING OF THE CATALYTIC CORE

The Tetrahymena thermophila self-splicing RNA is trapped in an inactiv e conformation during folding reactions at physiological temperatures. The structure of this metastable intermediate was probed by chemical modification interference and site-directed mutagenesis. Ln the inacti ve structure, an incorrect base-pairing, which we call Alt P3, displac es the P3 helix in the catalytic core of the intron. Mutations that st abilize Alt P3 increase the fraction of pre-rRNA that becomes trapped in the inactive structure, whereas mutations that destabilize Alt P3 r educe accumulation of this conformer. At high concentrations of Mg2+, the yield of correctly folded mutant pre-rRNAs is similar to wild-type RNA. Under these conditions, the rate of folding for mutant RNAs is s lower than for the wild-type, but is increased by addition of urea. Th e results show that slow folding of the Tetrahymena pre-rRNA is a cons equence of non-native secondary structure in the catalytic core of the intron, which is linked to an alternative hairpin in the 5' exon. Thi s illustrates how kinetically stable, long-range interactions shape RN A folding pathways. (C) 1998 Academic Press.