Discovery of coniferaldehyde as an inhibitor of caseinolytic protease to combat Staphylococcus aureus infections

The rising incidence of methicillin-resistant Staphylococcus aureus (MRSA) poses a significant threat to global public health, highlighting the urgent need for novel therapies and treatments in clinical settings. Caseinolytic protease P (ClpP) serves as a key component of bacterial degradation systems, playing a crucial role in maintaining cellular homeostasis and contributing to pathogenicity. Targeting ClpP function inhibition has demonstrated potential in combating antibiotic resistance and offers a promising therapeutic strategy for treating S. aureus infections. In this study, coniferaldehyde (CA) was identified as a ClpP inhibitor through ClpP peptidase inhibition assay. CA reduced the hemolysis activity, protease hydrolysis and bacterial invasion ability via regulating the transcription of main virulence factors. Furthermore, CA treatment led to a decreased resistance of S. aureus to adverse stimuli, including heat, acidic pH, high osmotic environment, hydrogen peroxide and NaClO stress assays. Notably, CA enhanced the efficacy of the bactericidal antibiotic tigecycline against growing S. aureus in time-killing assays. Molecular simulations and mutagenesis analyses revealed that the amino acids M31 and G33 were critical for the interaction between CA and ClpP. Importantly, CA exhibited excellent protective efficacy against S. aureus pneumonia in murine infection models. Our findings confirm that CA is an effective ClpP inhibitor with potential as a therapeutic agent for S. aureus infections.