title: Cloning, expression and purification of extracellular serine protease Esp, a biofilm-degrading enzyme, from Staphylococcus epidermidis.
abstract: OBJECTIVE
  Staphylococcus epidermidis Esp, an extracellular serine protease, inhibits Staphylococcus aureus biofilm formation and nasal colonization. To further expand the biotechnological applications of Esp, we developed a highly efficient and economic method for the purification of recombinant Esp based on a Brevibacillus choshinensis expression-secretion system.
  
  
  RESULTS
  The esp gene was fused with the N-terminal Sec-dependent signal sequence of the B. choshinensis cell wall protein and a C-terminal hexa-histidine-tag gene. The recombinant Esp was expressed and secreted into the optimized medium as an immature form and subsequently activated by thermolysin. The mature Esp was easily purified by a single purification step using nickel affinity chromatography and showed proteolytic activity as well as Staph. aureus biofilm destruction activity.
  
  
  CONCLUSIONS
  The purification yield of the developed extracellular production system was 5 mg recombinant mature Esp per 20-ml culture, which was much higher than that of an intracellular production system in Escherichia coli (3 mg recombinant Esp per 1-l culture).
  
  
  CONCLUSIONS
  Our findings will be a powerful tool for the production and purification of recombinant Esp and also applicable to a large variety of recombinant proteins used for basic researches and biotechnological applications.
authors.affiliation: Department of Bacteriology, The Jikei University School of Medicine, Minato-Ku, Tokyo, Japan. ssugimoto@jikei.ac.jp
authors.forename: S
authors.initials: S
authors.lastname: Sugimoto
authors.forename: T
authors.initials: T
authors.lastname: Iwase
authors.forename: F
authors.initials: F
authors.lastname: Sato
authors.forename: A
authors.initials: A
authors.lastname: Tajima
authors.forename: H
authors.initials: H
authors.lastname: Shinji
authors.forename: Y
authors.initials: Y
authors.lastname: Mizunoe
mesh_terms: D018441:Biofilms; D058675:Brevibacillus / Q000235:genetics / Q000378:metabolism*; D002364:Caseins / Q000378:metabolism; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006639:Histidine / Q000737:chemistry; D010957:Plasmids; D021382:Protein Sorting Signals; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D057057:Serine Proteases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D013211:Staphylococcus aureus / Q000502:physiology; D013212:Staphylococcus epidermidis / Q000201:enzymology* / Q000235:genetics; D013820:Thermolysin / Q000378:metabolism
chemical_list: D002364:Caseins; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; C026018:azocasein; D006639:Histidine; D057057:Serine Proteases; D013820:Thermolysin