RUI: Biosynthetic Diversification of Diketopiperazine Natural Products

With support from the Chemistry of Life Processes Program in the Division of Chemistry, Amy Lane from University of North Florida (UNF) aims to decipher Nature’s methods for creating a group of molecules known as 2,5-diketopiperazines (DKPs). These naturally occurring compounds offer a variety of applications in agriculture and other commercial enterprises. Microorganisms assemble DKPs through enzyme-catalyzed reactions, in which ubiquitous chemical building blocks are transformed into unique DKPs. Dr. Lane and her team seek to develop a molecular-level understanding of these enzymes to unleash the potential for biocatalytic synthesis of the next generation of DKPs. This project will be integrated into undergraduate education, with the project acting as a vehicle for providing hands-on research training to undergraduates. Impact on the future STEM (Science, Technology, Engineering and Mathematics) workforce will be magnified beyond direct research participants via the Creating Leaders in Multidisciplinary Basic Science (CLIMBS) program. The CLIMBS program implements professional development workshops as “ladders” to prepare UNF’s diverse student population for the pursuit of careers in science and to facilitate their entrance into graduate programs as a first step on that journey.Cyclodipeptide synthases (CDPSs) catalyze the formation of DKP scaffolds from two aminoacyl-tRNA substrates. These DKP scaffolds are structurally diversified and equipped for biological function via tailoring reactions. Nearly all of the >800 experimentally-characterized or bioinformatics-predicted CDPSs originate from bacterial gene clusters that also encode predicted DKP tailoring enzymes with diverse and unknown functions. These cryptic biosynthetic pathways are predicted to yield a plethora of uniquely functionalized DKPs, but remain poorly explored relative to pathways yielding other common natural product classes. The goals of this project are to establish molecular-level understanding of selected biosynthetic pathways that contain CDPSs and to diversify DKP natural products obtained from these pathways. To accomplish these goals, the Lane research group will apply an in silico-in vitro approach to characterize molecular details of DKP isomerization, develop an in vivo biosynthetic platform for the assembly of DKP analogs from unnatural amino acids, and establish isotope-labeled metabolomics approaches to identify new DKPs. This work is expected both advance fundamental understanding of diketopiperazine biosynthesis and yield strategies to more fully unleash the power of mechanistically-guided synthetic biology in natural product chemistry.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.