Expression, Purification, and Characterization of Kynurenine Formamidase from Bacillus cereus.
| dc.contributor.advisor | Born, Timothy L. | |
| dc.contributor.author | Bougie, James M. | |
| dc.creator | Bougie, James M. | |
| dc.date | 2011-12-08 | |
| dc.date.accessioned | 2012-02-02T18:44:48Z | |
| dc.date.available | NO_RESTRICTION | |
| dc.date.available | 2012-02-02T18:44:48Z | |
| dc.date.issued | 2012-02-02 | |
| dc.description.abstract | The kynurenine pathway of tryptophan catabolism in eukaryotes has long been known to be a source of the NAD precursor molecule quinolinic acid as well as immunomodulatory and neuroactive molecules which have been implicated in a wide variety of metabolic disorders and pathologies. Until fairly recently, the kynurenine pathway as it exists in its quinolinate producing role in eukaryotes was thought to be absent in prokaryotes. With the discovery of bacterial genes associated with some of the major enzymes in the pathway in several bacterial species, a “new” source of potential therapeutic targets was revealed. The complete function of the kynurenine pathway in bacteria is poorly understood due to its fairly recent discovery and resultant lack of mechanistic and structural data regarding its constituent enzymes. In this work, the second enzyme of the kynurenine pathway, kynurenine formamidase (EC 3.5.1.9), was cloned from the genome of Bacillus cereus into the pET100/D-TOPO expression vector, expressed in Escherichia coli, purified via metal affinity chromatography, and assayed in order to determine basic kinetic and mechanistic information. While determining the kinetic parameters for the enzyme and comparing them to those found in the literature, the behavior of the enzyme was found to deviate in some cases from the published values. These discrepancies have been noted and will serve as the basis for further study of the enzyme’s structure and mechanism. | |
| dc.identifier.uri | https://hdl.handle.net/1920/7519 | |
| dc.language.iso | en_US | |
| dc.subject | Kynurenine | |
| dc.subject | Tryptophan | |
| dc.subject | Formamidase | |
| dc.subject | Quinolinate | |
| dc.subject | Arylformamidase | |
| dc.subject | Enzymology | |
| dc.title | Expression, Purification, and Characterization of Kynurenine Formamidase from Bacillus cereus. | |
| dc.type | Thesis | |
| thesis.degree.discipline | Chemistry | |
| thesis.degree.grantor | George Mason University | |
| thesis.degree.level | Master's | |
| thesis.degree.name | Masters in Chemistry |