With this award, the Chemistry of Life Processes program is funding Dr. Denyce K. Wicht at Suffolk University and Dr. Daniel P. Dowling at UMass-Boston to research proteins involved in extracting sulfur from naturally occurring organic molecules. The cycling of sulfur-containing substances is important to understanding how living organisms (including humans) play a role in moving this specific element between earth's water, atmosphere, and land components. The interconversions of compounds containing carbon-sulfur bonds and sulfur-oxygen bonds are observed across a range of microorganisms, but the order in which the chemistry proceeds only recently was discovered. Therefore, these chemical transformations remain largely unexplored. This research entails a collaborative effort by Dr. Wicht and Dr. Dowling to perform fundamental biochemical analyses and obtain structural information for how these events take place. Research assistants at the undergraduate and graduate level gain experience with enzymology, biochemistry, and protein crystallography. Additionally, participants engage in outreach activities with middle school students and receive specific feedback on lifelong learning skills important to the production of a modern STEM workforce. Strong scientific mentorship provided by PI Wicht and co-PI Dowling contributes to the development of a diverse, globally competitive STEM workforce.
The recently reported bacterial sulfur assimilation pathway from naturally occurring dimethylsulfone to inorganic sulfite is the central focus of the work. There are three monooxygenases involved in the biochemical reactions: SfnG (for sulfone utilization), MsuC (for methanesulfinate utilization), and MsuD (for methanesulfonate ultilization). All three enzymes are reduced flavin dependent monooxygenases that require three substrates: an organosulfur molecule, O2, and FMNH2. The kinetic landscape of the novel biochemical pathway from dimethylsulfone to sulfite is unknown, therefore the immediate goal of the proposed research activities is the determination of the apparent steady-state kinetic parameters for each protein in the pathway using visible spectroscopy. The kinetic analyses of purified recombinant enzymes are coupled with substrate specificity data determined by 1H and 13C NMR spectroscopy. For each monooxygenase, a maximum rate of enzyme turnover and the specificity constant for the preferred organosulfur substrate is defined. High-resolution X-ray crystallography is used to determine the molecular details of enzyme active site(s) and reaction chemistry. Together, the experimental results are used to formulate a proposed overall enzyme mechanism consistent with the cleavage of two C-S bonds and the formation of one S-O bond. Mapping the active site environment of these enzymes guides future mechanistic studies by identifying important amino acid residues involved in bond-making and bond-breaking to sulfur.
This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.
