RNA structural studies : implications in HIV-1 replication and the origins of life
Abstract
Structure and function relationships are important to understand many biological processes. Biological components are not one dimensional and their structures may drive key aspects of their function. This dissertation focuses on understanding key structural elements involved in HIV-1 replication and elucidating the structure of RNA aptamers that could enhance our understanding of early life. In this dissertation we examined a key interaction in HIV-1 reverse transcription initiation, the interaction between the A-rich loop of the HIV-1 5' untranslated region (5'UTR) and the anticodon of tRNALys3. We showed that this interaction is important for viral infectivity. Using NMR structural techniques, we provided indirect evidence of this interaction and showed that it important for HIV-1 gRNA integrity. Using a combination of in vitro and cell-based assays we showed that this interaction serves to protect the HIV-1 gRNA from degradation by HIV-1 RT before reverse transcription can occur. We also examined another important host factor involved in HIV-1 replication, RNA Helicase A (RHA). RHA has been shown to be important for HIV-1 infectivity and acts as a processivity enhancement factor during reverse transcription. We showed that RHA is recruited to the virion through interactions with the primer binding site (PBS) segment of the HIV-1 5'UTR. We showed that this highly conserved region adopts a three-way junction structure that is recognized and is necessary for its recruitment. We also examined novel RNA aptamers that shift the reduction potential of bound flavin. This is a common strategy for highly reactive flavoenzymes and understanding the molecular mechanisms of flavin recognition may provide insights into early life. We were able to solve a preliminary solution structure of one of these aptamers and showed that it recognizes flavin using a scaffold of base triples. In summary, this work shows how structural features can influence biological function.
Degree
Ph. D.