PBP2/MreC: an ideal target to identify new antibacterial agents – BacWallScreen

The peptidoglycan, a heteropolymer and major component of the bacterial cell wall, is essential for cell integrity. It surrounds completely the cell and its synthesis is coordinated with cell growth and division. Its assembly is accomplished by periplasmic protein complexes involving synthetases, scaffolding proteins and hydrolases readily accessible from the environment. We propose to develop original antibacterials that block these protein complexes, in particular, the PBP2 and MreC proteins which complex formation is essential for the synthesis of the peptidoglycan. This novel strategy aimed at finding inhibitors of the PBP2:MreC complex will be based on two complementary assays: 1) a high-throughput HTRF (Homogeneous Time Resolved Fluorescence) screen of chemical libraries enriched for inhibitors of protein:protein interactions; 2) an in silico screen of the MolPort catalog against our PBP2:MreC 3D structure of Helicobacter pylori. The best candidates are then tested in the HTRF assay. Molecules identified through these two complementary approaches will then be 1) tested in a BRET (Bioluminescence Resonance Energy Transfer) assay that allows assessing their in cellulo efficacy followed by 2) a medicinal chemistry approach based on structure-activity relationship to improve their inhibitory potency. In parallel, we will model, synthesize and evaluate by HTRF followed by BRET small synthetic peptides mimicking the PBP2:MreC interface. Finally, the best molecules and peptidomimetics will be tested against H. pylori alone or in combination. Additionally, we will associate them to antimicrobial peptides and cell wall active antibiotics. Inhibition assays will be extended to Gram-negative pathogens of the ESKAPE group responsible for most multidrug resistant infections.