Coenzyme B12-Dependent Enzyme Catalysis
The long-term goal of our research on coenzyme B12 (adenosylcobalamin)-dependent enzyme catalysis is to reveal fundamental principles that govern radical-mediated catalyses in metalloenzymes. Specifically, we investigate the molecular mechanism of catalysis in the B12-dependent ethanolamine ammonia-lyase enzyme from Salmonella typhimurium. High-resolution (sub-Angstrom) molecular structure determinations are performed by using low temperature pulsed-EPR spectroscopy of frozen samples. Reaction kinetics and dynamics are probed on time scales from 10-13 to >104 s, in solid and fluid phases, and from <190 K to ambient temperatures, by using a combination of EPR and optical absorption techniques. We use isotope substitution, different substrate molecules, and site directed mutations to rationally modify the native system.