title: Efficient expression of sortase A fromStaphylococcus aureusinEscherichia coliand its enzymatic characterizations.
abstract: BACKGROUND
  Sortase A (SrtA) is a transpeptidase found inStaphylococcus aureus, which is widely used in site-specific protein modification. However, SrtA was expressed inEscherichia coli(E. coli) in rather low level (ranging from several milligrams to 76.9 mg/L at most). The present study aims to optimize fermentation conditions for improving SrtA expression inE. coli.
  
  
  RESULTS
  Under the optimized media (0.48 g/L glycerol, 1.37 g/L tryptone, 0.51 g/L yeast extract, MOPS 0.5 g/L, PBS buffer 180 mL/L) and condition (30 °C for 8 h) in a 7-L fermentor, the enzyme activity and the yield of SrtA reached 2458.4 ± 115.9 U/mg DCW and 232.4 ± 21.1 mg/L, respectively, which were higher by 5.8- and 4.5-folds compared with initial conditions, respectively. The yield of SrtA also represented threefold increase than the previously reported maximal level. In addition, the enzymatic characterizations of SrtA (optimal temperature, optimal pH, the influence of metal irons, and tolerance to water-soluble organic solvents) were determined.
  
  
  CONCLUSIONS
  Enhanced expression of SrtA was achieved by optimization of medium and condition. This result will have potential application for production levels of SrtA on an industry scale. Moreover, the detailed enzymatic characterizations of SrtA were examined, which will provide a useful guide for its future application.
authors.affiliation: The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, China.|State Key Laboratory of Natural and Biomimetic Drugs, Peking University, Beijing, 100191 China.
authors.forename: Zhimeng
authors.initials: Z
authors.lastname: Wu
authors.affiliation: The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, China.
authors.forename: Haofei
authors.initials: H
authors.lastname: Hong
authors.affiliation: The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, China.
authors.forename: Xinrui
authors.initials: X
authors.lastname: Zhao
authors.affiliation: The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, China.
authors.forename: Xun
authors.initials: X
authors.lastname: Wang
keywords: 7-L fermentor; Enzymatic characterizations; Escherichia coli; Response surface methodology; Sortase A