Investigation on the Antibacterial Activity and Molecular Mechanism of Rifamycin Derivatives Against Methicillin-resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA) poses a severe clinical drug resistance challenge. Point mutations in RpoB, the core target of rifamycins, have caused widespread of MRSA, highlighting the urgent need for developing novel rifamycin derivatives against resistant bacteria. In this study, 34a -rifamycin W-M1-hemiacetal ( 3c ), an anti-MRSA active compound, was screened from rifamycin biosynthetic intermediates and derivatives. 3c possesses a C5/C11 seco-rifamycin W skeleton and a hemiacetal six-membered ring in the polyketide side chain. Its mechanism of anti-MRSA biological activity was investigated by Scanning Electron Microscopy, Transmission Electron Microscopy, Confocal Laser Scanning Microscopy 3D and RNA Sequencing. Morphological observations revealed that rifampicin induced metabolic damage in MRSA via transcription inhibition, while 3c could significantly disrupt the integrity of MRSA cell membranes, causing damages such as membrane shrinkage and blebbing. RNA-seq further confirmed the distinct mechanisms: rifampicin affected core metabolic pathways (e.g., glycolysis, ribosome), whereas 3c regulated membrane synthesis and quorum sensing pathways, achieving dual bactericidal and anti-biofilm effects. Collectively, this provides new insights and experimental basis for addressing MRSA drug resistance, and offers theoretical support for the functional expansion of the rifamycin biosynthetic pathway.