DapE can function as an aspartyl peptidase in the presence of Mn2+

J Bacteriol. 2003 Aug;185(16):4748-54. doi: 10.1128/JB.185.16.4748-4754.2003.


Extracts of a multiply peptidase-deficient (pepNABDPQTE iadA iaaA) Salmonella enterica serovar Typhimurium strain contain an aspartyl dipeptidase activity that is dependent on Mn(2+). Purification of this activity followed by N-terminal sequencing of the protein suggested that the Mn(2+)-dependent peptidase is DapE (N-succinyl-L,L-diaminopimelate desuccinylase). A dapE chromosomal disruption was constructed and transduced into a multiply peptidase-deficient (MPD) strain. Crude extracts of this strain showed no aspartyl peptidase activity, and the strain failed to utilize Asp-Leu as a leucine source. The dapE gene was cloned into expression vectors in order to overproduce either the native protein (DapE) or a hexahistidine fusion protein (DapE-His(6)). Extracts of a strain carrying the plasmid overexpresssing native DapE in the MPD dapE background showed a 3,200-fold elevation of Mn(2+)-dependent aspartyl peptidase activity relative to the MPD dapE(+) strain. In addition, purified DapE-His(6) exhibited Mn(2+)-dependent peptidase activity toward aspartyl dipeptides. Growth of the MPD strain carrying a single genomic copy of dapE on Asp-Leu as a Leu source was slow but detectable. Overproduction of DapE in the MPD dapE strain allowed growth on Asp-Leu at a much faster rate. DapE was found to be specific for N-terminal aspartyl dipeptides: no N-terminal Glu, Met, or Leu peptides were hydrolyzed, nor were any peptides containing more than two amino acids. DapE is known to bind two divalent cations: one with high affinity and the other with lower affinity. Our data indicate that the form of DapE active as a peptidase contains Zn(2+) in the high-affinity site and Mn(2+) in the low-affinity site.

MeSH terms

  • Amidohydrolases / antagonists & inhibitors
  • Amidohydrolases / chemistry
  • Amidohydrolases / genetics*
  • Amidohydrolases / metabolism*
  • Aspartic Acid Endopeptidases / antagonists & inhibitors
  • Aspartic Acid Endopeptidases / chemistry
  • Aspartic Acid Endopeptidases / genetics
  • Aspartic Acid Endopeptidases / metabolism*
  • Binding Sites
  • Kinetics
  • Manganese / metabolism
  • Mutation
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Salmonella typhimurium / enzymology*
  • Salmonella typhimurium / genetics
  • Salmonella typhimurium / growth & development
  • Substrate Specificity
  • Zinc / metabolism


  • Recombinant Proteins
  • Manganese
  • Aspartic Acid Endopeptidases
  • Amidohydrolases
  • succinyldiaminopimelate desuccinylase
  • Zinc