Antimicrobial resistance (AMR) is a global health threat. The dramatic increase of Methicillin-resistant Staphylococcus aureus (MRSA) infections emphasizes the need to find new anti-infective agents with a novel mode of action. The Caseinolytic protease (ClpP) is a central virulence factor in stress survival, virulence, and antibiotic resistance of MRSA. Here, we found ayanin, a flavonoid isolated from Callicarpa nudiflora, was an inhibitor of MRSA ClpP with an IC₅₀ of 19.63 μM. Using quantitative real-time PCR, ayanin reduced the virulence of Staphylococcus aureus (S. aureus) by down-regulating the level of some important virulence factors, including agrA, RNAⅢ, hla, pvl, psmα and spa. The results of cellular thermal shift assay and thermal shift assay revealed a binding between ayanin and ClpP. Molecular docking showed that ASP-168, ASN-173 and ARG-171 were the potential binding sites for ClpP binding to ayanin. ClpP mutagenesis study further indicated that ARG-171 and ASN-173 were the main active sites of ClpP. The affinity constant (K_D) value of ayanin with ClpP was 3.15 × 10⁻⁵ M measured by surface plasmon resonance. In addition, ayanin exhibited a significant therapeutic effect on pneumonia infection induced by S. aureus in mice in vivo, especially in combination with vancomycin. This is the first report of ayanin with in vivo and in vitro efficacy against S. aureus infection. In conclusion, ayanin is a promising therapeutic agent to combat MRSA infections by targeting ClpP.

抗菌素耐药性（AMR）是一个全球性的健康威胁。耐甲氧西林金黄色葡萄球菌(MRSA) 感染的急剧增加强调需要寻找具有新颖作用方式的新型抗感染药物。酪蛋白分解酶 (ClpP) 是 MRSA 应激存活、毒力和抗生素耐药性的核心毒力因子。在这里，我们发现 ayanin（一种从裸花紫珠中分离出来的黄酮类化合物）是 MRSA ClpP 的抑制剂，IC ₅₀为 19.63 μM。利用实时定量PCR，绫青素通过下调一些重要毒力因子（包括agrA、RNAⅢ、hla、pvl、psmα和spa ）的水平来降低金黄色葡萄球菌（S. aureus）的毒力。细胞热位移测定和热位移测定的结果揭示了 ayanin 和 ClpP 之间的结合。分子对接表明ASP-168、ASN-173和ARG-171是ClpP与 ayanin 结合的潜在结合位点。ClpP诱变研究进一步表明ARG-171和ASN-173是ClpP的主要活性位点。通过表面等离振子共振测定，花青素与ClpP的亲和常数( K _D )值为3.15×10 ^{-5 M。}此外，花青素在体内对小鼠金黄色葡萄球菌引起的肺炎感染表现出显着的治疗效果，特别是与万古霉素联合使用。这是首次报道绫氨酸对金黄色葡萄球菌感染具有体内和体外功效。总之， ayanin 是一种有前途的治疗剂，可以通过靶向 ClpP 来对抗 MRSA 感染。