Inhibitory Effect And Mechanism Of Quercitrin On Staphylococcus Aureus Sortase A

Staphylococcus aureus (S. aureus)，a Gram-positive opportunistic pathogenicbacteria which can cause multiple infectious diseases that threaten human healthseriously, including endocarditis, necrotizing pneumonia and septicemia. Due tothe abuse of antibiotics, the antibiotic-resistant strains were isolated rapidly,especially the methicillin-resistant S. aureus (MRSA) which is reported only twoyears after methicillin was applied for the treatment of penicillin-resistant bacteria.And MRSA spread throughout of world just in10years. The reduced efficacy ofvancomycin and linezolid against MRSA makes research into developing newtreatment options a priority. Therefore, the urgent affair now is to solve the problemof antibiotic-resistant pathogen by developing novel drug targets or antibacterialstrategies.Anti-virulence strategy has been studied broadly. Compared with traditionalstrategies that are aimed at killing bacteria or preventing their growth, anti-virulenceapproach, a way to inhibit the expression and activity of virulence factors, willdiminishes the rate of development of bacterial resistance. Sortase A (SrtA) is acysteine transpeptidase of most Gram-positive bacteria that is responsible for theanchorage of many surface protein virulence factors to the cell wall layer. Thesesurface proteins, displayed by Gram-positive pathogenic bacteria, play a criticalroles in many processes of the establishment of an infection, including adherence,colonization, invasion, signaling and evasion of the host immune system. Therefore,SrtA as a key virulence factor may become an ideal target for the development ofanti-virulence therapeutics for the treatment of the Gram-positive bacterial infections.Actually, the screen of SrtA inhibitors is one of the current research focuses in the world. In this study, S. aureus SrtA was expressed and purified in order to screenSrtA inhibitors by fluorescence resonance energy transfer (FRET). There are4natural compounds had been identified, then quercitrin (QEN), the most effectivecompound, was further chosen for the study of the inhibition mechanism andexperimental therapeutic effects.Quercitrin is a flavonoid glycoside found in Flos Sophorae Immaturus, atraditional Chinese medicine, has been proven to possess a wide array ofpharmacological activities such as anti-inflammatory properties, anti-leishmanialproperties, anti-melanogenic properties, prevention of lipid peroxidation, andprotection against UVB-induced oxidative damage of the skin. In this study, wediscovered that QEN able to effectively inhibit the catalytic activity of S. aureusSrtA. Furthermore, we evaluated this inhibitory effect of QEN against SrtA activitythrough fibrinogen/fibronectin-binding assay,. The bacterial growth curve wasdetermined to assess the antibacterial activity of QEN on S. aureus. Using renalabscess model in mice to study the anti-virulence activity of QEN in vivo. Finally,we discussed the mechanism of QEN against SrtA by MD simulation. The resultsshowed that, QEN can cripple the adhesion of S. aureus SrtA without affect thebacterial growth. QEN dose-dependently inhibited S. aureus binding infibrinogen/fibronectin-binding assay, and the IC50for such inhibition is32.18±5.36μg/ml. Furthermore, QEN significantly alleviated S. aureus-induced renalabscess in mice. At last, MD simulations and mutagenesis assays suggested thatQEN binds to the active region of SrtA (Gly167and Val193). Taken together, thesefindings indicated that QEN is a leading compound for the development ofanti-virulence agents against S. aureus infections.