The long-term goal of the proposed research is to elucidate fundamental molecular principles that govern catalysis in enzymes, in the context of metal-radical pair generation and coupled radical-mediated reactions. This goal will be enacted through a comprehensive program of biochemical and physical studies of the coenzyme B12 (adenosylcobalamin) - dependent enzyme, ethanolamine ammonia-lyase, from Salmonella typhimurium. We have developed new methods, software and hardware for pulsed-electron paramagnetic resonance (EPR) spectroscopy. In conjunction with biochemical techniques, we have developed low-temperature systems and time- resolved EPR approaches that enable pioneering experimental efforts that address, directly, covalent bond- breaking/bond-making reactions in ethanolamine ammonia-lyase at single-step resolution. The general aims are to define the free energy landscape that guides the reactions, identify the origins and characterize the roles of the protein and coupled solvent fluctuations that propel the system over this landscape, and resolve the Angstrom-scale molecular structures that frame the free energy landscape. The projects develop a proposed "configurational catalysis" model for enzyme catalysis, and seek to establish the radical rearrangement as a paradigm for defining landscape and protein-solvent motional contributions to chemical steps in enzyme reactions. Contributions of solvent structure and dynamics to the enzyme reactions are also addressed by a suite of EPR spectroscopic and relaxation techniques. The outcomes of the research will impact molecular approaches in biomedicine and human health, including anti- microbial therapies, rational design of catalysts, inhibitors, and bio-nanocompartments, and control of free radical reactions.

Public Health Relevance

The long-term goal of the proposed research is to elucidate fundamental principles that govern catalysis in enzymes. Pioneering studies of contributions of molecular structure and dynamics to radical reactivity in B12 enzymes is enabled by development of new spectroscopic methods and unique low temperature biophysical approaches. Outcomes impact molecular approaches in biomedicine and human health, including anti-microbial therapies, rational design of catalysts, inhibitors, and bionanocompartments, and control of free radical reactions.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
High Priority, Short Term Project Award (R56)
Project #
2R56DK054514-15
Application #
8918207
Study Section
Macromolecular Structure and Function A Study Section (MSFA)
Program Officer
Sechi, Salvatore
Project Start
1998-03-15
Project End
2015-08-31
Budget Start
2014-09-18
Budget End
2015-08-31
Support Year
15
Fiscal Year
2014
Total Cost
$91,444
Indirect Cost
$28,634
Name
Emory University
Department
Physics
Type
DUNS #
066469933
City
Atlanta
State
GA
Country
United States
Zip Code
30322