Staphylococcus aureus was exposed to polycyclic polyprenylated acylphloroglucinols (PPAP) derivatives at different times of growth. The antimicrobial mechanism of PPAP 23 was investigated by RNA-seq combined with various biochemical approaches. Our results suggest that PPAP 23 interact with the cytoplasmic membrane of Gram-positive bacteria yielding to bacterial cell death.

Recently, a series of endo-type B polycyclic polyprenylated acylphloroglucinols (PPAP) derivatives with high antimicrobial activity were chemically synthesized. One of the derivatives, PPAP 23, which shows high bactericidal activity and moderate toxicity, was chosen for further investigation. PPAP 23 kills MRSA faster than vancomycin and eradicates established biofilms. Moreover, S. aureus does not develop detectable resistance against PPAP 23. The antimicrobial mechanism of PPAP 23 was investigated by RNA-seq combined with various biochemical approaches. RNA-seq suggests that PPAP 23 interferes with iron metabolism. PPAP 23 does not depolarize the membrane, but it affects the membrane integrity. PPAP 23 inhibits bacterial respiration and preferentially Fe-S enzymes. It has a comparable iron chelating activity as doxycycline, and supplementation of exogenous iron diminishes its antimicrobial activities. PPAP 23 induces reactive oxygen species (ROS) and triggers DNA disorganization. Taken together, we speculate that PPAP 23 with its lipophilic side chains interactchains interact with the cytoplasmic membrane of Gram-positive bacteria. In its iron-free form, PPAP 23 may chelate iron from Fe-S enzymes and deactivate the enzymes. In the iron bound form, Fe3+ - PPAP 23 may extract electrons from the menaquinone cycle, thus block the electron flow of the respiratory chain and oxygen consumption. PPAP 23 is not a strong iron chelator, iron may be dissociated from PPAP 23, which may lead to high iron levels in the cell. The elevated iron level signals iron overload and favors iron-based Fenton chemistry to generate toxic level of ROS resulting in cell death.