During protein synthesis, elongation factor Tu (EF-Tu) delivers aminoacyl-tRNA (aa-tRNA) to the A-site of mRNA-programmed ribosomes in a GTP-dependent manner. To enable future studies on the functional and structural requirement of EF-Tu’s function, a Cysteine-free variant of EF-Tu was constructed suitable for subsequent labelling of the protein and use in kinetic studies. Here, the kinetic properties of three Cysteine-less EF-Tu variants are reported, demonstrating that only the variant with the Alanine substitution in position 81 retains wild-type activity with respect to the interaction with guanine nucleotides, aa-tRNA and the ribosome. To explore a possible tRNA independent pathway for the GTPase activation signal, three residues in domain II of EF-Tu (Arginine 279, Glycine 280, Arginine 283) were mutated; the activity of EF-Tu variants were analyzed. Results suggest that these residues are indeed required for efficient ribosome-dependent stimulation of the GTPase activity of EF-Tu. / x, 85 leaves : ill. (some col.) ; 29 cm
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:ALU.w.uleth.ca/dspace#10133/3107 |
| Date | January 2011 |
| Creators | Mo, Fan |
| Contributors | Wieden, Hans-Joachim |
| Publisher | Lethbridge, Alta. : University of Lethbridge, Dept. of Chemistry and Biochemistry, c2011, Arts and Science, Department of Chemistry and Biochemistry |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | en_US |
| Detected Language | English |
| Type | Thesis |
| Relation | Thesis (University of Lethbridge. Faculty of Arts and Science) |
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