Expression, Purification, and Characterization of Kynurenine Formamidase from Bacillus cereus.

dc.contributor.advisorBorn, Timothy L.
dc.contributor.authorBougie, James M.
dc.creatorBougie, James M.
dc.date2011-12-08
dc.date.accessioned2012-02-02T18:44:48Z
dc.date.availableNO_RESTRICTION
dc.date.available2012-02-02T18:44:48Z
dc.date.issued2012-02-02
dc.description.abstractThe 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.urihttps://hdl.handle.net/1920/7519
dc.language.isoen_US
dc.subjectKynurenine
dc.subjectTryptophan
dc.subjectFormamidase
dc.subjectQuinolinate
dc.subjectArylformamidase
dc.subjectEnzymology
dc.titleExpression, Purification, and Characterization of Kynurenine Formamidase from Bacillus cereus.
dc.typeThesis
thesis.degree.disciplineChemistry
thesis.degree.grantorGeorge Mason University
thesis.degree.levelMaster's
thesis.degree.nameMasters in Chemistry

Files

Original bundle
Now showing 1 - 1 of 1
Loading...
Thumbnail Image
Name:
Bougie_thesis_2011.pdf
Size:
4.74 MB
Format:
Adobe Portable Document Format
License bundle
Now showing 1 - 1 of 1
No Thumbnail Available
Name:
license.txt
Size:
1.65 KB
Format:
Item-specific license agreed upon to submission
Description: