Structural And Function Of Bacterial Multidrug-resistance Pump NorA And NorB

The antibiotic has saved countless lives since its discovery However,the drug resistance has emerged due to the wide use of antibiotics.Quinolone,a chemical synthesized antibiotic family,is remarkably effective in treatment of infection.Quinolone resistance has been found in Staphylococcus aureus since its widespread applications in the early 1980s.NorA and NorB are believed to play critical roles in resistance of fluoroquinolone in Staphylococcus aureus.Both NorA and NorB are anitpoters.They pump out fluoroquinolones diffused into cell via an electrochemical proton gradient mechanism resulting in reducing the intracellular antibiotic concentration.NorA is a 12 transmembrane protein composed of 388 amino acids,while NorB possess 14 transmembrane helices.Due to their complexity of substrate selection and the difficulty of the expression,purification and crystallization of membrane protein,their structures and physiological functions have not been fully understood.The crystal structure of NorA and NorB can help us to understand the mechanism of substrate specificity and the transport mechanism.We screened the expression of the homologous of NorA and NorB in the thermophilic and halophilic bacteria,respectively.We obtained several high expression,high stability and high purity membrane proteins.Moreover,by screening different salt concentrations,pH values and detergents,we optimized the purification conditions of protein.Finally,the diffractive crystals of membrane protein have been obtained,but the diffraction resolution need further to improve.At the same time,using a variety of biochemical experiments,we investigated the mechanism of transportation,these results will provide the structure and function study of these transporters.