Extraction, Purification, and Characterization of 2'hydroxybipenyl-2-Sulfanate (HBPS) Desulfinase (DszB) from Rhodococcus Erythropolis

Shannon G. Woolridge, Chemical Engineering, University of Kentucky College of Engineering Paducah Campus, Paducah, KY and Derek L. Englert, Chemical and Materials Engineering, University of Kentucky, Paducah, KY and Christina M. Payne, Chemical and Materials Engineering, University of Kentucky, Lexington, KY

The transportation industry consumes 94 percent of the global liquid petroleum products for fuel usage.  These refined fuel products are derived from crude oil which contains various concentrations of sulfur, depending on its origin.  Residual sulfur in finished fuels convert to sulfur oxides (SO_x) upon combustion which are emitted as a toxic pollutant to the environment causing acid deposition (acid rain), smog, and respiratory irritation in humans.  The low-sulfur crude oil supply is dwindling since demand for liquid fuels is ever-increasing, consequently to meet this critical demand, impurity-rich reserves, such as heavy North American oil, are used.  Though these non-traditional sources help meet the demand, the impurity-rich raw materials have an abundant amount of thiophenic sulfur compounds.  The most widely adopted technology for removing sulfur compounds or desulfurization, is known as hydrodesulfurization, which while effective in sulfur removal in light refinery fractions; it leaves a substantial amount of the thiophenic sulfur within the middle distillate and heavy oil fractions.  Also, hydrodesulfurization breaks carbon-carbon bonds, reducing the amount of energy produced per gallon of fuel.  Alternatively, biodesulfurization offers a complimentary approach to conventional desulfurization methods, producing high-energy and low-sulfur levels while the carbon-carbon bonds remain intact.  The research presented in this poster examines modifications made to the desulfinase (DszB) enzyme.  The best species for this particular type of mechanism is the Rhodococcus species; they are not restricted by mass transport rate limitations seen from other bacteria.  A particular strain of soil bacteria, Rhodococcus erythropolis, was found to produce a family of enzymes that cleave the sulfur-carbon molecule without altering the carbon-carbon bonds, proving ideal for this process.   DNA extracted from R. erythropolis, using polymerase chain reaction (PCR) techniques, set the foundation for the DszB enzyme implantation at specific locations that would affect its activity.  Results from this research presents that the DszB enzyme DNA was successfully placed into Escherichia coli (E. coli) which will act as the host strain for further analysis.  By identifying distinctive behaviors within the active site interactions compared to those of bound molecules, chemical and dynamic relationships can be recognized. Future design plans are to reduce competitive binding at certain sites and improve performance of the DszB enzyme for applications in biosulfurization.