Purification and characterisation of a lactococcal aminoacylase

Arch Microbiol. 2003 Jun;179(6):402-8. doi: 10.1007/s00203-003-0544-5. Epub 2003 Apr 8.


The amd1-encoded aminoacylase from Lactococcus lactis MG1363 was cloned and overexpressed in Escherichia coli and purified. The assumed dimeric enzyme has a subunit molecular mass of about 42 kDa and contains 2.0+/-0.1 g-atoms of zinc and cobalt, in equimolar amounts, per subunit of Amd1. The enzyme was characterised with respect to substrate specificity, pH, temperature and metal dependence. Amd1 exhibited a broad activity range towards N-acetylated- l-amino acids with a strong preference towards those containing neutral aliphatic and aromatic side chains. It hydrolysed N-acetyl- l-alanine most efficiently, and exhibited temperature and pH optima of 30 degrees C and 7.0, respectively. The activity of Amd1 towards N-acetyl- l-alanine was enhanced by the divalent cation Co(2+), while Cd(2+ )inhibited activity. Interestingly, Amd1 was shown to catalyse the hydrolysis of several dipeptides at pH 7.0, although with reduced V(max) values as compared to hydrolysis of N-acetylated- l-amino acids. This characteristic has also biological significance since Amd1 was able to complement a growth deficiency in a L. lactis triple peptidase mutant.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amidohydrolases / genetics*
  • Amidohydrolases / isolation & purification
  • Amidohydrolases / metabolism*
  • Cloning, Molecular
  • DNA, Bacterial / analysis
  • Electrophoresis, Polyacrylamide Gel
  • Hydrogen-Ion Concentration
  • Lactococcus lactis / enzymology*
  • Lactococcus lactis / genetics
  • Lactococcus lactis / growth & development
  • Metals / metabolism
  • Mutation
  • Peptide Hydrolases / metabolism
  • Plasmids / genetics
  • Polymerase Chain Reaction
  • Substrate Specificity
  • Temperature


  • DNA, Bacterial
  • Metals
  • Peptide Hydrolases
  • Amidohydrolases
  • aminoacylase I