Group II introns are mobile genetic elements that have been implicated as agents of genetic diversity, and serve as important model systems for investigating RNA catalysis and pre-mRNA splicing. In the absence of an atomic-resolution structure of the intron, detailed understanding of its catalytic mechanism has remained elusive. Previous identification of a divalent metal ion stabilizing the leaving group in both splicing steps suggested that the group II intron may employ a "two-metal ion" mechanism, a catalytic strategy used by a number of protein phosphoester transfer enzymes. Using metal rescue experiments, we now reveal the presence of a second metal ion required for nucleophile activation in the exon-ligation step of group II intron splicing. Coupled with biochemical and structural evidence of at least two metal ions at the group I intron reaction center, these results suggest a mechanistic paradigm for describing catalysis by large ribozymes.
Bibliographical noteFunding Information:
We thank Cecilia Cortez for oligonucleotide synthesis and members of the Piccirilli lab for helpful discussions, advice, and comments regarding the manuscript. P.M.G. and R.F. acknowledge the Medical Scientist Training Program at the University of Chicago (5T32GM007281-32) for support.