Structure and Mechanism of Isopropylmalate Dehydrogenase from Arabidopsis thaliana: INSIGHTS ON LEUCINE AND ALIPHATIC GLUCOSINOLATE BIOSYNTHESIS

J Biol Chem. 2016 Jun 24;291(26):13421-30. doi: 10.1074/jbc.M116.730358. Epub 2016 May 2.


Isopropylmalate dehydrogenase (IPMDH) and 3-(2'-methylthio)ethylmalate dehydrogenase catalyze the oxidative decarboxylation of different β-hydroxyacids in the leucine- and methionine-derived glucosinolate biosynthesis pathways, respectively, in plants. Evolution of the glucosinolate biosynthetic enzyme from IPMDH results from a single amino acid substitution that alters substrate specificity. Here, we present the x-ray crystal structures of Arabidopsis thaliana IPMDH2 (AtIPMDH2) in complex with either isopropylmalate and Mg(2+) or NAD(+) These structures reveal conformational changes that occur upon ligand binding and provide insight on the active site of the enzyme. The x-ray structures and kinetic analysis of site-directed mutants are consistent with a chemical mechanism in which Lys-232 activates a water molecule for catalysis. Structural analysis of the AtIPMDH2 K232M mutant and isothermal titration calorimetry supports a key role of Lys-232 in the reaction mechanism. This study suggests that IPMDH-like enzymes in both leucine and glucosinolate biosynthesis pathways use a common mechanism and that members of the β-hydroxyacid reductive decarboxylase family employ different active site features for similar reactions.

Keywords: enzyme; plant biochemistry; protein evolution; protein structure; reaction mechanism; x-ray crystallography.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3-Isopropylmalate Dehydrogenase / chemistry*
  • 3-Isopropylmalate Dehydrogenase / genetics
  • 3-Isopropylmalate Dehydrogenase / metabolism
  • Arabidopsis / enzymology*
  • Arabidopsis / genetics
  • Arabidopsis Proteins / chemistry*
  • Arabidopsis Proteins / genetics
  • Arabidopsis Proteins / metabolism
  • Catalytic Domain
  • Crystallography, X-Ray
  • Glucosinolates / biosynthesis*
  • Glucosinolates / chemistry
  • Glucosinolates / genetics
  • Leucine / biosynthesis*
  • Leucine / chemistry
  • Leucine / genetics
  • Structure-Activity Relationship


  • Arabidopsis Proteins
  • Glucosinolates
  • 3-Isopropylmalate Dehydrogenase
  • Leucine

Associated data

  • PDB/3R8W
  • PDB/5J32
  • PDB/5J33
  • PDB/5J34