UC San Diego

Group I introns are a class of catalytic RNAs that can perform a self-splicing reaction. This thesis will examine the large-scale conformational changes of three Tetrahymena group I intron (TET) states. A combination of mutational analysis and cryogenic electron microscopy (cryo-EM) was used to determine the dynamics associated with the L9/P5 mutation (∆L9) in which a long-range tertiary contact that affects the structure-function of TET is knocked out. This mutation allowed for the visualization of dynamics that were not seen in the previously published models of the wild-type (wt) construct. There are three different states of TET modeled: an intermediate form in which the intron is connected to the 5` exon, but the 3` exon is spliced; a second step (S2) state where the intron is base pairing to the ligated exons, and finally, a post-second step (post-S2) state in which the intron is fully spliced. Understanding the structural changes during the splicing of ∆L9 TET provides specific insights into the structure-function relationships of group I introns and general mechanisms of RNA catalysis and dynamics. The structural biology methods developed to facilitate these experiments have also expanded the use of cryo-EM as a general tool for studying many different RNAs. Furthermore, by increasing our knowledge of RNA catalysis, we gain a more comprehensive grasp of these essential biological macromolecules and their role in the origins of life on Earth.