Lysine 2,3-aminomutase catalyzes the interconversion of L-lysine and L- beta-lysine. This reaction is the first committed step in the pathway that allows Clostridia to use Lysine as a source of carbon, nitrogen, and energy. Clostridia also use beta-L-lysine for the biosynthesis of a number of antibiotics. Numerous studies indicate that several intermediates on the reaction pathway contain carbon-centered radicals. The purpose of this study is to determine how the enzyme in combination with its cofactors (pyridoxal phosphate, an iron-sulfur cluster [Fe-S], and S- adenosylmethionine (SAM)) generates these radicals. The working hypothesis is that upon electron transfer to SAM from [Fe-S], SAM homolyzes to generate methionine and a 5'-deoxyadenosyl radical (5'-dA.) which is responsible for initiating catalysis. In order to test this model, electron paramagnetic resonance and Mossbauer spectroscopies will be used to assess a change in the oxidation state of the [Fe-S] during catalysis. In addition, the kinetic competence of this change will be verified using rapid kinetics methods. Lastly, analogs of SAM will be synthesized and used as probes for communication between the [Fe-S] and SAM during the cleavage event, as well as traps to assess the production of 5'-dA.. This proposed radical generating system is also present in the ribonucleotide reductase from anaerobically growing E. coli, as well as pyruvate formate lyase, and research in this area may offer clues for new methods of inhibiting anaerobic bacteria.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32GM018282-03
Application #
2770823
Study Section
Physical Biochemistry Study Section (PB)
Project Start
1998-09-01
Project End
Budget Start
1998-09-01
Budget End
1999-02-28
Support Year
3
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Wisconsin Madison
Department
Biochemistry
Type
Other Domestic Higher Education
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715