| Staphylococcus aureus(S.aureus),a common gram-positive micrococci,is one of the most famous zoonotic pathogen and a critical cause of various diseases,including skin infections,soft tissue infections,endocarditis,sepsis and necrotizing pneumonia.The drug resistance of S.aureus has always been the barrier to the treatment of the infectious diseases,especially the spread of MRSA strains,which has brought great difficulties to clinical treatment.Previous studies have reported that nearly 90% of MRSA strains have multidrug resistance mechanisms,which can resistant to variety of common antibiotics including ?-lactam,aminoglycoside tetracycline and fluoroquinolones.Vancomycin is regarded as the last option for the treatment of MRSA infections.However,in recent years,the markedly decreasing efficiency of vancomycin on MRSA suggests that MRSA infections may be incurable in the future.?-lactamase is one of the main reasons for the drug resistance of S.aureus.It can be combined with the carbonyl group of ?-lactam ring to catalyze the hydrolysis,and inactivate the antibacterial ability.The encoding genes of ?-lactamase can be carried by plasmids and spread to many kinds of pathogenic bacteria.Previous studies have shown that nearly 95% of the S.aureus clinical isolates have the ability secreting ?-lactamase,and it has seriously affected the use of ?-lactam antibiotics.However,?-lactam antibiotic is always one of the most widely used antibacterial in the world,and accounts for 60% of total antibiotic usage each year.For now,only few kinds of ?-lactamase inhibitors have been put into use in clinical practice,and all of them can only be used for class A,C,D ?-lactamase.For now,the spread of class B ?-lactamase has become a huge challenge for the treatment,but there is still no class B ?-lactamase inhibitors used in clinical practice.Accordingly,it is an urgent need to develop class B ?-lactamase inhibitors.In this study,we found that Theaflavin-3,3?-gallate(TFDG)can effectively inhibit the hydrolysis of class B ?-lactamase and increase the antibacterial activity of ?-lactam antibiotics against MRSA strains.MRSA BAA1717 was used as main experimental strain in this study,and its typical class B ?-lactamase,?-lactamase N1,was used as the main target protein.Further,we used theoretical chemistry methods including homology modeling,molecular docking and molecular simulation assays to calculate the sites of?-lactamase N1 binding with TFDG,and verified the amino acid sites by site-directed mutagenesis assay.The mouse model of S.aureus infection was established to revealed that TFDG can significantly increase the protective effect of ?-lactam antibiotics on mice,including reducing the mortality rate of mice,decreasing the colonization of bacteria in lungs and alleviating the tissue lesions of lungs.China is the hometown of tea,and tea culture is also an important embodiment of Chinese spiritual culture.In recent years,the research of active constituents in tea has attracted much attention.Furthermore,the theaflavin which has many kinds of biological function has become a hot area of reseach.In conclusion,our study provides an important experimental basis for the mechanism of TFDG inhibiting the activity of MBLs,and also provides experimental evidence and innovative compounds for the research of MBLs inhibitors. |
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