RNA folding

RNA plays an indispensable enzymatic and structural role in many critical cellular processes, but understanding of its folding mechanism is relatively incomplete. Here I present studies that combine computer predictions based on a simple model of folding with direct observations of single molecules to bring greater understanding to the dynamics and folding pathways of three different RNAs. Many cellular RNAs require a protein co-factor in order to function, so we first use single molecule experimental techniques to understand RNA folding in the presence of a protein co-factor, in chapter I. In parallel, a Go-based technique was developed for the simulation of RNA folding, allowing some of the first computer simulations of the process. The results of these in silico studies are reported in chapter II, for a very small (and computationally accessible) example RNA, the GCAA tetraloop, demonstrating a "zipper" pathway with a putative intermediate. This methodology is then applied to the hairpin ribozyme in chapter III, to derive a model of the folding transition state for the ribozyme and make predictions about the ribozyme dynamics. In chapter III preliminary observations are also made of the high-speed dynamics of the ribozyme in single molecule experiments using two different types of microscopy.