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Title: Structure and Mechanism of Isopropylmalate Dehydrogenase from Arabidopsis thaliana: Insights on Leucine and Aliphatic Glucosinolate Biosynthesis

Abstract

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 Mg2+ 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. Furthermore, 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.

Authors:
 [1];  [1];  [1]
  1. Washington Univ., St. Louis, MO (United States)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
National Science Foundation (NSF); USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
OSTI Identifier:
1267478
Grant/Contract Number:  
MCB-0904215; AC02–06CH11357
Resource Type:
Accepted Manuscript
Journal Name:
Journal of Biological Chemistry
Additional Journal Information:
Journal Volume: 291; Journal Issue: 26; Journal ID: ISSN 0021-9258
Publisher:
American Society for Biochemistry and Molecular Biology
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; 37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; enzyme; plant biochemistry; protein evolution; protein structure; x-ray crystallography; reaction mechanism

Citation Formats

Lee, Soon Goo, Nwumeh, Ronald, and Jez, Joseph M. Structure and Mechanism of Isopropylmalate Dehydrogenase from Arabidopsis thaliana: Insights on Leucine and Aliphatic Glucosinolate Biosynthesis. United States: N. p., 2016. Web. doi:10.1074/jbc.M116.730358.
Lee, Soon Goo, Nwumeh, Ronald, & Jez, Joseph M. Structure and Mechanism of Isopropylmalate Dehydrogenase from Arabidopsis thaliana: Insights on Leucine and Aliphatic Glucosinolate Biosynthesis. United States. doi:10.1074/jbc.M116.730358.
Lee, Soon Goo, Nwumeh, Ronald, and Jez, Joseph M. Mon . "Structure and Mechanism of Isopropylmalate Dehydrogenase from Arabidopsis thaliana: Insights on Leucine and Aliphatic Glucosinolate Biosynthesis". United States. doi:10.1074/jbc.M116.730358. https://www.osti.gov/servlets/purl/1267478.
@article{osti_1267478,
title = {Structure and Mechanism of Isopropylmalate Dehydrogenase from Arabidopsis thaliana: Insights on Leucine and Aliphatic Glucosinolate Biosynthesis},
author = {Lee, Soon Goo and Nwumeh, Ronald and Jez, Joseph M.},
abstractNote = {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 Mg2+ 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. Furthermore, 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.},
doi = {10.1074/jbc.M116.730358},
journal = {Journal of Biological Chemistry},
number = 26,
volume = 291,
place = {United States},
year = {2016},
month = {5}
}

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