Date of Award
1-2016
Degree Type
Dissertation
Degree Name
Doctor of Philosophy (PhD)
Department
Biochemistry
First Advisor
Barbara L. Golden
Committee Chair
Barbara L. Golden
Committee Member 1
Frederick S. Gimble
Committee Member 2
Christine A. Hrycyna
Committee Member 3
Elizabeth J. Tran
Abstract
Ribozymes, just like protein enzymes, catalyze diverse chemical reactions. The first goal of this dissertation is to understand the mechanism of ribozyme-mediated phosphodiester cleavage reaction. Biochemical assays and X-ray crystallography were used for probing the active site of two small self-cleaving ribozymes, the hepatitis delta virus (HDV) ribozyme and the hammerhead ribozyme. Results presented here suggest that divalent metal ions play critical roles in the catalytic mechanisms of both the HDV and the hammerhead ribozymes. In the HDV ribozyme, the result is consistent with an active site Mg2+ being directly involved in catalysis. In the hammerhead ribozyme, however, Mg2+ ions likely indirectly influence catalysis through binding to the active site nucleotides. The second goal of this dissertation is to engineer an artificial ribozyme that mimics the function of the natural aminoacyl-tRNA synthetase. The design was based on the sequence of a naturally occurring riboswitch and an in vitro selected ribozyme that could charge tRNA with chemically synthesized unnatural amino acid substrates. The engineered ribozyme reported here serves as a prototype that could ultimately be used for site-specific incorporation of unnatural amino acids into proteins in cells.
Recommended Citation
Chen, Ji, "Investigating and expanding the functionality of RNA catalysts: Studies of the hepatitis delta virus, the hammerhead, and the aminoacyl-tRNA synthetase-like ribozymes" (2016). Open Access Dissertations. 632.
https://docs.lib.purdue.edu/open_access_dissertations/632