Diaminopimelate dehydrogenase (DAP-DH) catalyzes the single step conversion of L-2-amino-6-ketopimeIate to meso-DAP, the direct precursor of L-lysine in the bacterial lysine synthetic pathway. The enzyme catalyzes the oxidative deamination of an amino acid of D configuration, and must additionally distinguish between two chiral centers on the same symmetric substrate, we have therefore initiated crystallographic studies to probe the structural basis for this stereospecific discrimination. DAP-DH has been cloned, expressed in E.coli and purified to homogeneity using standard biochemical procedures The crystalline structure of the complex DAPDH-NADP(+) has been solved using multiple isomorphous replacement procedures and non-crystallographic symmetry averaging. The resulting model has been refined against 2.4 A diffraction data to a crystallographic R-factor of 15.1%. DAP-DH is a homodimer of structurally not-identical subunits, Each subunit is composed of two domains, with a deep cleft in between. The N-terminal domain contains a modified dinucleotide binding domain or Rossman fold (6 central β-strands in a 213456 topology, surrounded by 4 α-helices). The C-terminal domain is composed of 6 β-strands and 5 α-helices. The dinier interface involves mainly residue from the C-terminal portion of the protein. The relative positions of the two domains in the two monomers are different, possibly due to different lattice contacts. The nucleotide is bound in an extended conformation across the C-terminal portion of the β-sheet of the Rossman fold, with its C4 facing the cleft between the two domains. The structure of DAP-DH closely resembles the structure of dihydrodipicolinate reductase, the enzyme immediately preceding DAP-DH in the DAP/lysine biosynthetic pathway.
|Original language||English (US)|
|State||Published - Dec 1 1996|
ASJC Scopus subject areas
- Molecular Biology