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Binding of the competitive inhibitor L-captopril to the dapE-encoded Nsuccinyl- L,L-diaminopimelic acid desuccinylase from Neisseria meningitidis (NmDapE) was examined by kinetic, spectroscopic, and crystallographic methods. L-Captopril, an angiotensinconverting enzyme (ACE) inhibitor, was previously shown to be a potent inhibitor of the DapE from Haemophilus influenzae (HiDapE) with an IC50 of 3.3 μM and a measured fQ of 1.8 //M and displayed a dose-responsive antibiotic activity toward Escherichia coli. L-Captopril is also a competitive inhibitor of NmDapE with Ki of 2.8 μM. To examine the nature of the interaction of L-captopril with the dinuclear active site of DapE, we have obtained electron paramagnetic resonance (EPR) and magnetic circular dichroism (MCD) data for the enzymatically hyperactive Co(ll)-substituted forms of both HiDapE and NmDapE. EPR and MCD data indicate that the two Co(Il) ions in DapE are antiferromagnetically coupled, yielding an S' = 0 ground state, and suggest a thiolate bridge between the two metal ions. Verification of a thiolate-bridged dinuclear complex was obtained by determining the three-dimensional X-ray crystal structure of NmDapE in complex with l - captopril at 1.8 Å resolution. Combination of these data provides new insights into binding of L-captopril to the active site of DapE enzymes as well as important inhibitor--active site residue interaction's. Such information is critical for the design of new, potent inhibitors of DapE enzymes. [ABSTRACT FROM AUTHOR]