Antimicrobial resistance (AMR) poses a significant global public health threat, necessitating the discovery of new antimicrobials. Natural products (NPs) are important reservoirs for such molecules. Among them, 2,5-diketopiperazines (DKPs) stand out due to their remarkable biological activities. DKP biosynthesis typically involves a core enzyme known as cyclodipeptide synthase (CDPS), which forms a cyclodipeptide scaffold, followed by one or more tailoring enzymes that introduce chemical modifications, leading to more complex DKPs. While the diversity of DKPs obtained is substantial, it remains limited since the initial cyclodipeptide scaffolds are predominantly composed of aromatic and hydrophobic amino acids.
Recently, novel core enzymes termed RCDPSs have been identified, showing no sequence homology to CDPSs. Notably, these RCDPSs utilize aminoacyl-tRNAs as substrates to synthesize cyclodipeptide scaffolds containing arginine.
This project proposes to investigate these RCDPSs, aiming to enable the biosynthesis of diverse DKPs containing arginine and other charged amino acids. The objectives are to establish the natural repertoire of cyclodipeptide scaffolds produced by these enzymes, understand the molecular basis of their substrate specificity, and ultimately perform enzymatic and metabolic engineering to generate a broader diversity of non-natural DKPs with charged amino acids. The project will be carried out using a range of biological (molecular biology, biochemistry, biophysics) and analytical chemistry (LC-MS) methods, with collaborations involving experts in structural biology and synthetic chemistry. If the project's progress allows, a collaboration will be established with an already identified platform to test the biological activity of the generated compounds.