Methionine is involved in many cellular processes, several of which produce a feedback inhibitor. 5-methylthioribose (MTR) kinase, one protein involved in the removal of this inhibitor, has a protein kinase fold with conserved kinase motifs and several unique MTR binding motifs. Site-directed mutagenesis and characterization of the Bacillus subtilis enzyme was performed to probe the role of one motif. Active site D233 mutants show an activity profile similar to other protein kinase-like enzymes, suggesting a common mechanism that does not require a catalytic acid. An ordered sequential binding mechanism, with nucleotide binding first, was seen in wild type MTR kinase. Binding studies of the mutant proteins suggest that hydrogen bonding is important for MTR binding. The structures of the mutant proteins also show more differences in MTR binding than nucleotide binding. Overall, D233 is important for increasing the nucleophilicity of MTR, and ensuring its correct position in the active site.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:OTU.1807/32571 |
| Date | 25 July 2012 |
| Creators | Dawson, Karen |
| Contributors | Howell, P. Lynne |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | en_ca |
| Detected Language | English |
| Type | Thesis |
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