Mechanism of substrate recognition and PLP-induced conformational changes in LL-diaminopimelate aminotransferase from Arabidopsis thaliana

J Mol Biol. 2008 Dec 31;384(5):1314-29. doi: 10.1016/j.jmb.2008.10.022. Epub 2008 Oct 15.


LL-Diaminopimelate aminotransferase (LL-DAP-AT), a pyridoxal phosphate (PLP)-dependent enzyme in the lysine biosynthetic pathways of plants and Chlamydia, is a potential target for the development of herbicides or antibiotics. This homodimeric enzyme converts L-tetrahydrodipicolinic acid (THDP) directly to LL-DAP using L-glutamate as the source of the amino group. Earlier, we described the 3D structures of native and malate-bound LL-DAP-AT from Arabidopsis thaliana (AtDAP-AT). Seven additional crystal structures of AtDAP-AT and its variants are reported here as part of an investigation into the mechanism of substrate recognition and catalysis. Two structures are of AtDAP-AT with reduced external aldimine analogues: N-(5'-phosphopyridoxyl)-L-glutamate (PLP-Glu) and N-(5'-phosphopyridoxyl)- LL-Diaminopimelate (PLP-DAP) bound in the active site. Surprisingly, they reveal that both L-glutamate and LL-DAP are recognized in a very similar fashion by the same sets of amino acid residues; both molecules adopt twisted V-shaped conformations. With both substrates, the alpha-carboxylates are bound in a salt bridge with Arg404, whereas the distal carboxylates are recognized via hydrogen bonds to the well-conserved side chains of Tyr37, Tyr125 and Lys129. The distal C(epsilon) amino group of LL-DAP is specifically recognized by several non-covalent interactions with residues from the other subunit (Asn309*, Tyr94*, Gly95*, and Glu97* (Amino acid designators followed by an asterisk (*) indicate that the residues originate in the other subunit of the dimer)) and by three bound water molecules. Two catalytically inactive variants of AtDAP-AT were created via site-directed mutagenesis of the active site lysine (K270N and K270Q). The structures of these variants permitted the observation of the unreduced external aldimines of PLP with L-glutamate and with LL-DAP in the active site, and revealed differences in the torsion angle about the PLP-substrate bond. Lastly, an apo-AtDAP-AT structure missing PLP revealed details of conformational changes induced by PLP binding and substrate entry into the active site.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoenzymes / chemistry
  • Apoenzymes / metabolism
  • Arabidopsis / enzymology*
  • Arabidopsis Proteins / chemistry*
  • Arabidopsis Proteins / metabolism*
  • Catalysis
  • Catalytic Domain
  • Crystallography, X-Ray
  • Diaminopimelic Acid / chemistry
  • Diaminopimelic Acid / metabolism
  • Glutamic Acid / chemistry
  • Glutamic Acid / metabolism
  • Lysine / biosynthesis
  • Models, Molecular
  • Mutant Proteins / chemistry
  • Mutant Proteins / metabolism
  • Oxidation-Reduction
  • Protein Structure, Secondary
  • Pyridoxal Phosphate / chemistry
  • Pyridoxal Phosphate / metabolism*
  • Static Electricity
  • Substrate Specificity


  • Apoenzymes
  • Arabidopsis Proteins
  • Mutant Proteins
  • Glutamic Acid
  • Diaminopimelic Acid
  • Pyridoxal Phosphate
  • Lysine

Associated data

  • PDB/3EI5
  • PDB/3EI6
  • PDB/3EI7
  • PDB/3EI8
  • PDB/3EI9
  • PDB/3EIA
  • PDB/3EIB