The dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase from Haemophilus influenzae contains two active-site histidine residues

  • Danuta M. Gillner
  • David L. Bienvenue
  • Boguslaw P. Nocek
  • Andrzej Joachimiak
  • Vincentos Zachary
  • Brian Bennett
  • Richard C. Holz
Original Paper


The catalytic and structural properties of the H67A and H349A dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) from Haemophilus influenzae were investigated. On the basis of sequence alignment with the carboxypeptidase from Pseudomonas sp. strain RS-16, both H67 and H349 were predicted to be Zn(II) ligands. The H67A DapE enzyme exhibited a decreased catalytic efficiency (180-fold) compared with wild-type (WT) DapE towards N-succinyldiaminopimelic acid. No catalytic activity was observed for H349A under the experimental conditions used. The electronic paramagnetic resonance (EPR) and electronic absorption data indicate that the Co(II) ion bound to H349A-DapE is analogous to that of WT DapE after the addition of a single Co(II) ion. The addition of 1 equiv of Co(II) to H67A DapE provides spectra that are very different from those of the first Co(II) binding site of the WT enzyme, but that are similar to those of the second binding site. The EPR and electronic absorption data, in conjunction with the kinetic data, are consistent with the assignment of H67 and H349 as active-site metal ligands for the DapE from H. influenzae. Furthermore, the data suggest that H67 is a ligand in the first metal binding site, while H349 resides in the second metal binding site. A three-dimensional homology structure of the DapE from H. influenzae was generated using the X-ray crystal structure of the DapE from Neisseria meningitidis as a template and superimposed on the structure of the aminopeptidase from Aeromonas proteolytica (AAP). This homology structure confirms the assignment of H67 and H349 as active-site ligands. The superimposition of the homology model of DapE with the dizinc(II) structure of AAP indicates that within 4.0 Å of the Zn(II) binding sites of AAP all of the amino acid residues of DapE are nearly identical.


Biomedicine Biosynthesis Electron paramagnetic resonance Enzyme kinetics Homology model 



Aminopeptidase from Aeromonas proteolytica


Carboxypeptidase from Pseudomonas sp. strain RS-16


dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase


Electron paramagnetic resonance


4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid


Inductively coupled plasma atomic emission spectrometry




N-Succinyl-l,l-diaminopimelic acid


Wild type



This work was supported by the National Science Foundation (CHE- 0652981, RCH) and the National Institutes of Health (AI056321, RR001980 BB). The Bruker ESP-300E EPR spectrometer was purchased with funds provided by the National Science Foundation (BIR-9413530).


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Copyright information

© SBIC 2008

Authors and Affiliations

  • Danuta M. Gillner
    • 1
    • 2
  • David L. Bienvenue
    • 1
  • Boguslaw P. Nocek
    • 3
  • Andrzej Joachimiak
    • 3
  • Vincentos Zachary
    • 1
  • Brian Bennett
    • 4
  • Richard C. Holz
    • 1
  1. 1.Department of ChemistryLoyola University ChicagoChicagoUSA
  2. 2.Department of ChemistrySilesian University of TechnologyGliwicePoland
  3. 3.The Midwest Center for Structural GenomicsArgonne National LaboratoryArgonneUSA
  4. 4.The National Biomedical EPR CenterBiophysics Research Institute, Medical College of WisconsinMilwaukeeUSA

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