Isonitriles are organic molecules that contain a carbon-nitrogen triple bond. These molecules, which are involved in radical chemistry and are important reagents for multi-component chemical reactions, are both difficult to synthesize chemically and possess an extremely foul odor. These properties have likely limited the study and use of these interesting compounds. Surprisingly, there are many examples of biological isonitrile compounds, found in both terrestrial and marine environments. These compounds are commonly produced by bacterial pathogens, including species that infect fish, plants, insects, and mammals. The mechanistic basis for biosynthesis of isonitrile natural products has not been studied. This project will investigate the enzymology of PvcA, the isonitrile-synthesizing enzyme from a Pseudomonas aeruginosa pathway that is responsible for the synthesis of a novel natural product. This enzyme converts the amino group of tyrosine to an isonitrile, using the C2 carbon of ribulose-5-phosphate as the source of the carbon atom in the isonitrile. This research project will examine the structural and functional mechanisms of the PvcA enzymes from three homologous bacterial pathways. A catalytic mechanism for the PvcA reaction will be tested through the collaborative enzymatic and structural approaches using the crystal structure of the PvcA enzyme from Pseudomonas aeruginosa as an experimental framework. These isonitrile clusters additionally contain PvcB, a non-heme oxygenase that catalyzes pathway-specific two- or four-electron oxidations. The structural and chemical basis for differences in reactions catalyzed by PvcB will also be examined. These experiments will provide detailed mechanistic insights into the structure and function of these novel enzymes, expanding our knowledge of the synthetic capabilities of diverse microorganisms.
Broader Impact This research project will explore the enzymatic synthesis of isonitriles, an area of research that is virtually unexplored and will therefore provide an exciting research opportunity to graduate and undergraduate trainees. Specifically, this project will support the training of two graduate students. It will additionally support multiple undergraduate students through established, highly successful summer and academic year research programs. Undergraduate students will fully participate in lab activities, will present their work and write lab reports, and will be integrated into the lab so that they understand the contribution made to the larger research study. Students will be trained to identify and address specific hypothesis-driven questions and will receive broad interdisciplinary training in the important areas of chemical, biochemical, and structural biology.
This project is jointly supported by the Biomolecular Dynamics, Structure and Function Cluster in the Division of Molecular and Cellular Biosciences and the Chemistry of Life Processes program in the Chemistry Division.
