title: The leucine aminopeptidase of Staphylococcus aureus is secreted and contributes to biofilm formation.
abstract: BACKGROUND
  Staphylococcus aureus has emerged as a major drug-resistant pathogen in hospital- and community-acquired infections. Leucine aminopeptidase (LAP) is known to be essential for survival of the bacteria; however the LAP of S. aureus has not been extensively characterized. In this study, we report a detailed characterization of the S. aureus LAP.
  
  
  METHODS
  LAP from S. aureus was cloned, purified, and further biochemically characterized. The expression of LAP was analyzed by Western blotting. Growth and biofilm formation were analyzed spectrophotometrically.
  
  
  RESULTS
  LAP was cloned from S. aureus and expressed as a 55 kDa protein, whereas the molecular weight of the native protein is approximately 600 kDa. LAP showed amidolytic activity against l-leucine p-nitroanilide. Optimal activity was observed at pH 8.5 and 37°C with a V(max) of 2500μmol/min/mg protein. LAP enzymatic activity was inhibited by ion chelators and enhanced by divalent metal ions, specifically Ni. LAP is secreted by laboratory as well as clinical strains. Bestatin, an inhibitor of LAP, inhibits S. aureus growth and biofilm formation.
  
  
  CONCLUSIONS
  To our knowledge, this is the first detailed characterization of LAP from S. aureus and suggests its importance in survival and pathogenesis.
authors.affiliation: Department of Cell Biology, School of Biological Sciences and Biotechnology, Indian Institute of Advanced Research, Koba Institutional Area, Gandhinagar 382007, Gujarat, India. rsingh@iiar.res.in
authors.forename: Arun Kumar
authors.initials: AK
authors.lastname: Singh
authors.forename: Rochika
authors.initials: R
authors.lastname: Singh
authors.forename: Dhanendra
authors.initials: D
authors.lastname: Tomar
authors.forename: Chirayu D
authors.initials: CD
authors.lastname: Pandya
authors.forename: Rajesh
authors.initials: R
authors.lastname: Singh
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018441:Biofilms / Q000187:drug effects / Q000254:growth & development*; D020128:Inhibitory Concentration 50; D007700:Kinetics; D007930:Leucine / Q000031:analogs & derivatives / Q000494:pharmacology; D007931:Leucyl Aminopeptidase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid; D013211:Staphylococcus aureus / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D007931:Leucyl Aminopeptidase; D007930:Leucine; C012211:ubenimex