Coordinated Mechanistic Approaches to Desulfonation in Two-component FMN Monooxygenases

East Carolina University, Greenville NC

Investigators

Abstract

With support from the Chemistry of Life Processes Program in the Division of Chemistry, Professor Holly Ellis and her research team at East Carolina University will study the interesting chemistry carried out by some of the enzymes involved in the acquisition of the element sulfur that bacteria need for survival. A fascinating feature of these enzymes is their ability to change their structures to facilitate the reactions they catalyze. Many of these crucial structural changes and accompanying catalytic events are poorly understood. The two-component enzyme systems selected for this study utilize various structural changes to promote sulfur acquisition. Therefore, these enzyme systems will be ideal to evaluate a multitude of different structural strategies that would be essential for catalysis. The project will also be integrated with the Brody RISE Pre-College Program to train high school students from underrepresented groups in the sciences in different biochemical techniques used to understand enzyme function. An undergraduate student fellowship will be provided in the summer to mentor and encourage participation of underrepresented students in scientific research. Two-component flavin mononucleotide(FMN)-dependent monooxygenases are essential for sulfur acquisition in bacteria when preferred sulfur sources are limiting. These enzymes utilize an FMN reductase and monooxygenase to carry out their critical functional roles; roles that require an elegant coordinated mechanistic strategy for desulfonation. The described objectives will evaluate if the two-component flavin-dependent systems involved in sulfur acquisition have evolved common structural features and mechanistic steps to carry out their complex functional role. Objective 1 is to identify the mechanistic strategies of two-component FMN-dependent monooxygenases that promote desulfonation. Objective 2 is to determine how the structural properties of two-component NAD(P)H:FMN reductases dictate function. Both objectives utilize complementary spectroscopic, kinetic, and biophysical approaches to provide evidence for the desulfonation mechanism and identify oligomeric changes that contribute to flavin transfer in the NAD(P)H:FMN reductases. 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.