3EI9

Crystal structure of K270N variant of LL-diaminopimelate aminotransferase from Arabidopsis thaliana complexed with L-Glu: External aldimine form

Summary for 3EI9

• Entry DOI: 10.2210/pdb3ei9/pdb
• Related: 3EI5 3EI6 3EI7 3EI8 3EIA 3EIB
• Descriptor: LL-diaminopimelate aminotransferase, (E)-N-({3-hydroxy-2-methyl-5-[(phosphonooxy)methyl]pyridin-4-yl}methylidene)-L-glutamic acid, GLYCEROL, ... (5 entities in total)
• Functional Keywords: aminotransferase, lysine biosynthesis, pyridoxal 5' phosphate, external aldimine, ll-diaminopimelate, chloroplast, pyridoxal phosphate, transferase, transit peptide
• Biological source: Arabidopsis thaliana (mouse-ear cress,thale-cress)
• Cellular location: Plastid, chloroplast: Q93ZN9
• Total number of polymer chains: 2
• Total formula weight: 96598.40

Authors

Watanabe, N.,Clay, M.D.,van Belkum, M.J.,Cherney, M.M.,Vederas, J.C.,James, M.N.G. (deposition date: 2008-09-15, release date: 2008-10-14, Last modification date: 2023-08-30)

Primary citation

Watanabe, N.,Clay, M.D.,van Belkum, M.J.,Cherney, M.M.,Vederas, J.C.,James, M.N.
Mechanism of substrate recognition and PLP-induced conformational changes in LL-diaminopimelate aminotransferase from Arabidopsis thaliana.
J.Mol.Biol., 384:1314-1329, 2008

PubMed Abstract: 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.

PubMed: 18952095
DOI: 10.1016/j.jmb.2008.10.022

Experimental method

X-RAY DIFFRACTION (1.55 Å)