Date of Award
Doctor of Philosophy (PhD)
Biotechnology Science and Engineering
Luis R. Cruz-Vera
Ribosomes., Allosteric proteins., Proteins--Synthesis., Gene expression., Amino acids., Tryptophan.
Ribosomes often times work in unison with both nascent peptides and accessory molecules to regulate expression of genes by controlling protein synthesis. This allows for fine regulation of gene activity based on current cellular physiology. In prokaryotes, there are numerous differences among regulatory nascent peptides that induce ribosome arrest, thus allowing for specificity in regulation. In the tna operon the amino acid L-tryptophan, L-Trp, can selectively interact with the ribosome translating tnaC mRNA, and thereby control the expression of the enzyme tryptophanase and an L-Trp transporter. It has been suggested that the combined action of L-Trp and the regulatory TnaC nascent peptide affects the catalytic site of the ribosome known as peptide transfer center (PTC). Since the nature of substrates entering the PTC and their molecular mechanism of reaction are greatly variable, it is possible that the action of L-Trp and TnaC would also be variable and perhaps selective to the substrates at the PTC. This study focused on determining the nature of the inhibition of the ribosome function produced by L-Trp and TnaC as explored using biochemical analyses to better understand regulatory nascent peptides. The results showed that L-Trp performs as an allosteric inhibitor, requiring the binding of RF2, puromycin, or aminoacyl-tRNAs to the ribosome concurrently with L Trp. Hydrolysis reactions induced by RF2 during translation termination were more efficiently inhibited than was the peptidyl-transferase reaction induced by aminoacyl-tRNAs during translation elongation. L-Trp operates allosterically to inhibit accommodation of selective nucleophilic molecules in the PTC and thus hydrolysis of the peptide or full peptidyl transfer. In conclusion, L-Trp and TnaC are specific regulators of the translation termination process.
Gordon, Emily R., "Allosteric regulation of the ribosome by L-tryptophan" (2015). Dissertations. 65.