Information on EC 4.2.1.33 - 3-isopropylmalate dehydratase

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea

EC NUMBER
COMMENTARY
4.2.1.33
-
RECOMMENDED NAME
GeneOntology No.
3-isopropylmalate dehydratase
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
(2R,3S)-3-isopropylmalate = (2S)-2-isopropylmalate
show the reaction diagram
overall reaction
-
-
-
(2R,3S)-3-isopropylmalate = 2-isopropylmaleate + H2O
show the reaction diagram
stoichiometry of reaction, enzyme also hydrates 2-isopropylmaleate to 3-hydroxy-4-methyl-3-carboxypentanoate, interconversion between the 2 isomers
Neurospora sp.
-
(2R,3S)-3-isopropylmalate = 2-isopropylmaleate + H2O
show the reaction diagram
equilibrium concentration ratios
-
(2R,3S)-3-isopropylmalate = 2-isopropylmaleate + H2O
show the reaction diagram
feedback regulation by leucine
-
(2R,3S)-3-isopropylmalate = 2-isopropylmaleate + H2O
show the reaction diagram
mechanism, equilibrium in vitro favors reaction opposite the direction of leucine biosynthesis
-
(2R,3S)-3-isopropylmalate = 2-isopropylmaleate + H2O
show the reaction diagram
transcriptional feedback regulation by leucine
-
(2R,3S)-3-isopropylmalate = 2-isopropylmaleate + H2O
show the reaction diagram
(1a)
-
-
-
2-isopropylmaleate + H2O = (2S)-2-isopropylmalate
show the reaction diagram
(1b)
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
condensation
Neurospora sp., Salmonella sp.
-
-
elimination
-
of H2O, C-O bond cleavage, , trans elimination
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Biosynthesis of secondary metabolites
-
-
leucine metabolism
-
-
Metabolic pathways
-
-
Valine, leucine and isoleucine biosynthesis
-
-
SYSTEMATIC NAME
IUBMB Comments
(2R,3S)-3-isopropylmalate hydro-lyase (2-isopropylmaleate-forming)
Forms part of the leucine biosynthesis pathway. The enzyme brings about the interconversion of the two isomers of isopropylmalate. It contains an iron-sulfur cluster.
SYNONYMS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
3-isopropylmalate dehydratase
-
-
3-isopropylmalate isomerase
Q58673
-
Alpha-IPM isomerase
-
-
-
-
alpha-isopropylmalate isomerase
-
-
-
-
beta-Isopropylmalate dehydratase
-
-
-
-
dehydratase,beta-isopropylmalate
-
-
-
-
IPM dehydratase
-
-
-
-
IPM isomerase
Q58673
-
IPMI
-
-
-
-
IPMI
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
-
IPMI
P9WK95, P9WQF5
-
IPMI
P9WK95, P9WQF5
-
-
IPMI SSU1
-
isopropylmalate isomerase small subunit1
isopropylmalate isomerase
-
-
-
-
isopropylmalate isomerase
-
-
isopropylmalate isomerase
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
-
isopropylmalate isomerase
P81291
-
isopropylmalate isomerase
Q58673
-
isopropylmalate isomerase
-
-
isopropylmalate isomerase
P9WK95, P9WQF5
-
isopropylmalate isomerase
P9WK95, P9WQF5
-
-
leuC
P81291
gene name
leuC
Q58673
large subunit
LeuCD
P9WK95, P9WQF5
-
LeuCD
P9WK95, P9WQF5
-
-
LeuD
Q58673
small subunit
SOI10
-
-
-
-
Superoxide-inducible protein 10
-
-
-
-
MJ0499
P81291
gene name, large subunit of 3-isopropylmalate dehydratase
additional information
P9WK95, P9WQF5
the enzyme belongs to the aconitase superfamily
additional information
P9WK95, P9WQF5
the enzyme belongs to the aconitase superfamily
-
CAS REGISTRY NUMBER
COMMENTARY
37290-72-5
-
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
large subunit
UniProt
Manually annotated by BRENDA team
the Arabidopsis genome encodes one potential large subunit and three potential small subunits, encoded by AtleuC and AtleuD genes
-
-
Manually annotated by BRENDA team
Corynebacterium glutamicum B-6
strain B-6
-
-
Manually annotated by BRENDA team
large subunit of 3-isopropylmalate dehydratase
SwissProt
Manually annotated by BRENDA team
large subunit LeuC
UniProt
Manually annotated by BRENDA team
small subunit LeuD
UniProt
Manually annotated by BRENDA team
large subunit LeuC
UniProt
Manually annotated by BRENDA team
small subunit LeuD
UniProt
Manually annotated by BRENDA team
wild type and complementing Leu-2 mutants
-
-
Manually annotated by BRENDA team
Neurospora sp.
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
malfunction
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
for IPMI small subunit 1, no plants homozygous for the T-DNA allele can be obtained in either analyzed line (ipmi ssu1-1 or ipmi ssu1-2)
malfunction
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
homozygous ipmi 3-1mutants can be established. In leaves and seeds of the ipmi ssu3-1 mutant few significant changes in amino acid content. Met levels are unchanged in ipmi ssu3-1 and S-methylmethionine can not be detected
malfunction
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
homozygous ipmi ssu2-1 mutants can be established. Free amino acid content of the leaves and seeds of ipmi ssu2-1 plants is not substantially different from that of the wild-type. S-methylmethionine is not detectable in the ipmi ssu2-1 mutant. Profile of the methionine-derived glucosinolates in leaves of ipmi ssu2-1 does not show substantial variations from that of wild-type plants
malfunction
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
in leaves, ipmi lsu1-2 plants exhibit the weakest chemical phenotype in comparison to wild-type: transport or storage form of Met is barely detectable. Many more changes between the ipmi lsu1-1 and 1-3 plants and wild-type: accumulation of S-methylmethionine in rosette leaves of both mutants which is accompanied by 2fold increased levels of Met. Average 2-isopropylmalate content of 0.42 and 0.02 mg/g dry weight in leaves of ipmi lsu1-3 and ipmi lsu1-1 mutants, respectively, while this metabolite is undetectable in the ipmi lsu1-2 plants. Relative levels of 2-(3'-methylsulfinyl)propylmalate are 10/1/14 in ipmi lsu1-1, 1-2 and 1-3 plants
malfunction
-
lethal phenotype of the atleud3 mutant with a defect in female gametophyte development
malfunction
-
knockdown plants contain chloroplasts with abnormal morphology and show a severe macroscopic phenotype with narrow, often undulated leaves with yellow pale green leaf blades except at the vascular bundles, which show normal green color
metabolism
-
isopropylmalate isomerase large subunit and small subunits form heterodimers to catalyze the isomerization of 2-isopropylmalate to 3-isopropylmalate in leucine biosynthesis in bacteria and archaea. Reverse genetics and metabolite profiling show that AtLeuD1 and AtLeuD2 function redundantly in aliphatic glucosinolate biosynthesis, but AtLeuD3 is not likely to be involved in this pathway
physiological function
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
an intact IPMI small subunit 1 gene seems to be crucial for the development of mature viable seeds
physiological function
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
function of IPMI large subunit 1 in both the biosynthesis of Leu and in the Met chain elongation pathway of glucosinolate formation
physiological function
-
isopropylmalate isomerase large subunit and small subunits form heterodimers to catalyze the isomerization of 2-isopropylmalate to 3-isopropylmalate in leucine biosynthesis in bacteria and archaea. AtLeuD3 plays an essential role in leucine biosynthesis and female gametophyte development
physiological function
-
the small subunit 1 of the Arabidopsis isopropylmalate isomerase is required for normal growth and development and the early stages of glucosinolate formation
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
(2R,3S)-3-isopropylmalate
(2S)-2-isopropylmalate
show the reaction diagram
O59393
the isopropylmalate isomerase small subunit of the hyperthermophilic archaea Pyrococcus horikoshii (PhIPMI-s) functions as isopropylmalate isomerase in the leucine biosynthesis pathway, and as homoaconitase (HACN) in the lysine biosynthesis pathway via alpha-aminoadipic acid
-
-
?
(2R,3S)-3-isopropylmalate
2-isopropylmaleate + H2O
show the reaction diagram
-
-
-
-
?
(2R,3S)-3-isopropylmalate
2-isopropylmaleate + H2O
show the reaction diagram
Q58673
-
-
-
?
(2S)-2-isopropylmalate + H2O
(2R,3S)-3-isopropylmalate
show the reaction diagram
P9WK95, P9WQF5
-
-
-
?
(2S)-2-isopropylmalate + H2O
(2R,3S)-3-isopropylmalate
show the reaction diagram
P9WK95, P9WQF5
-
-
-
?
2-isopropylmaleate + H2O
3-isopropylmalate
show the reaction diagram
-
-
-
-
2-isopropylmaleate + H2O
3-isopropylmalate
show the reaction diagram
-
-
-
-
2-isopropylmaleate + H2O
3-isopropylmalate
show the reaction diagram
-
-
-
-
2-isopropylmaleate + H2O
3-isopropylmalate
show the reaction diagram
Neurospora sp., Salmonella sp.
-
-
-
-
2-isopropylmaleate + H2O
?
show the reaction diagram
Neurospora sp.
-
high activities in leu-1 mutants which have accumulated great amounts of inducer
-
-
-
2-isopropylmaleate + H2O
?
show the reaction diagram
-
second of three reactions of leucine biosynthesis
-
-
-
2-isopropylmaleate + H2O
?
show the reaction diagram
-
second of three reactions of leucine biosynthesis
-
-
-
2-isopropylmaleate + H2O
?
show the reaction diagram
Neurospora sp., Salmonella sp.
-
second of three reactions of leucine biosynthesis
-
-
-
3-isopropylmalate
2-isopropylmaleate + H2O
show the reaction diagram
-
-
-
-
3-isopropylmalate
2-isopropylmaleate + H2O
show the reaction diagram
-
-
-
-
3-isopropylmalate
2-isopropylmaleate + H2O
show the reaction diagram
-
-
-
-
3-isopropylmalate
2-isopropylmaleate + H2O
show the reaction diagram
Neurospora sp., Salmonella sp.
-
-
-
-
additional information
?
-
Neurospora sp., Salmonella sp.
-
citrate, aconitate, isocitrate are not substrates
-
-
-
additional information
?
-
P9WK95, P9WQF5
the functional LeuCD complex catalyzes the stereospecific conversion reaction of alpha-isopropylmalate to beta-isopropylmalate, the active site is only completely formed after LeuC and LeuD have assembled
-
-
-
additional information
?
-
P9WK95, P9WQF5
the functional LeuCD complex catalyzes the stereospecific conversion reaction of alpha-isopropylmaleate to beta-isopropylmalate, the active site is only completely formed after LeuC and LeuD have assembled. The LeuD residues 30-37 form the substrate discriminating loop, and LeuD residues 70-74 the substrate binding loop
-
-
-
additional information
?
-
P9WK95, P9WQF5
the functional LeuCD complex catalyzes the stereospecific conversion reaction of alpha-isopropylmaleate to beta-isopropylmalate, the active site is only completely formed after LeuC and LeuD have assembled. The LeuD residues 30-37 form the substrate discriminating loop, and LeuD residues 70-74 the substrate binding loop
-
-
-
additional information
?
-
P9WK95, P9WQF5
the functional LeuCD complex catalyzes the stereospecific conversion reaction of alpha-isopropylmalate to beta-isopropylmalate, the active site is only completely formed after LeuC and LeuD have assembled
-
-
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
(2R,3S)-3-isopropylmalate
(2S)-2-isopropylmalate
show the reaction diagram
O59393
the isopropylmalate isomerase small subunit of the hyperthermophilic archaea Pyrococcus horikoshii (PhIPMI-s) functions as isopropylmalate isomerase in the leucine biosynthesis pathway, and as homoaconitase (HACN) in the lysine biosynthesis pathway via alpha-aminoadipic acid
-
-
?
(2R,3S)-3-isopropylmalate
2-isopropylmaleate + H2O
show the reaction diagram
-
-
-
-
?
(2R,3S)-3-isopropylmalate
2-isopropylmaleate + H2O
show the reaction diagram
Q58673
-
-
-
?
(2S)-2-isopropylmalate + H2O
(2R,3S)-3-isopropylmalate
show the reaction diagram
P9WK95, P9WQF5
-
-
-
?
(2S)-2-isopropylmalate + H2O
(2R,3S)-3-isopropylmalate
show the reaction diagram
P9WK95, P9WQF5
-
-
-
?
2-isopropylmaleate + H2O
?
show the reaction diagram
Neurospora sp.
-
high activities in leu-1 mutants which have accumulated great amounts of inducer
-
-
-
2-isopropylmaleate + H2O
?
show the reaction diagram
-
second of three reactions of leucine biosynthesis
-
-
-
2-isopropylmaleate + H2O
?
show the reaction diagram
-
second of three reactions of leucine biosynthesis
-
-
-
2-isopropylmaleate + H2O
?
show the reaction diagram
Neurospora sp., Salmonella sp.
-
second of three reactions of leucine biosynthesis
-
-
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Iron-sulfur cluster
P81291
the structure of the protein has unbound Fe-S clusters and contains a fourth cysteine in the active site
additional information
Neurospora sp., Salmonella sp.
-
no requirement
additional information
-
no requirement
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
(NH4)2SO4
-
recovery after restoration of lower ionic strength shows hysteresis effect
1,10-phenanthroline
-
16% inhibition at 0.1 mM
8-hydroxyquinolinesulfonate
-
16% inhibition at 0.1 mM
beta-carboxy-beta-hydroxyisocaproate
-
synthetic, racemic mixture
Cd2+
-
complete inhibition at 1 mM
Co2+
-
3% inhibition at 1 mM
Cu2+
Neurospora sp.
-
complete inhibition at 10 mM
Cu2+
-
complete inhibition at 1 mM
cysteine
Neurospora sp.
-
complete inhibition at 0.004-0.02 mM
cysteine
-
55% inhibition at 20 mM, protected by substrate
dimethylmesaconate
Neurospora sp.
-
-
Fe2+
-
13% inhibition at 1 mM
Hg2+
Neurospora sp.
-
complete inhibition at 10 mM
Hg2+
-
complete inhibition at 1 mM
isopropylmalate
Neurospora sp., Salmonella sp.
-
-
KCN
-
inactivation prevented by ammonium sulfate
KCN
-
complete inactivation at 10 mM; inactivation prevented by ammonium sulfate
KF
-
most potent inhibitor
Mg2+
-
26% inhibition at 1 mM
N-ethylmaleimide
Neurospora sp.
-
65% inhibition after preincubation in 1 mM
N-ethylmaleimide
-
-
sulfhydryl compounds
-
protection by substrate
Zn2+
-
complete inhibition at 1 mM
Mn2+
-
45% inhibition at 1 mM
additional information
-
inhibition at pH 4.5-5.0, reactivation at neutral pH
-
additional information
-
additional compounds tested, without effect
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
Neurospora sp., Salmonella sp.
-
no requirement
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.078
alpha-hydroxy-beta-carboxyisocaproate
-
-
4.57
alpha-hydroxy-beta-carboxyisocaproate
-
-
0.75
beta-carboxy-beta-hydroxyisocaproate
-
-
0.95
beta-carboxy-beta-hydroxyisocaproate
-
-
0.083
citraconate
-
crude enzyme preparation
1.79
citraconate
-
crude enzyme preparation
0.0216
dimethylcitraconate
-
-
0.048
dimethylcitraconate
-
crude enzyme preparation
0.18
dimethylcitraconate
-
-
additional information
additional information
Neurospora sp.
-
Ks for standard substrates
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
12.3
Neurospora sp.
-
-
additional information
-
-
additional information
-
enzyme activity with citraconate 4.8-fold and 7.4-fold higher than with standard substrates
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6
-
enzyme assay at
7 - 8
Neurospora sp.
-
-
7.5
P9WK95, P9WQF5
assay at
8.5
-
enzyme assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
4 - 9.5
P9WK95, P9WQF5
-
6 - 8
Neurospora sp., Salmonella sp.
-
at pH 6.0, 20% of optimum activity, activity varies only about 20% between pH 7.0 and 8.0
6 - 8
-
at pH 6.0, 20% of optimum activity, activity varies only about 20% between pH 7.0 and 8.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
30
-
enzyme assay at
34
-
enzyme assay at
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
15 - 95
P9WK95, P9WQF5
-
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
-
expression in all floral tissues including sepal, stamen, pistil as well as pollen grains, but it is only present at both ends of developing siliques
Manually annotated by BRENDA team
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
-
Manually annotated by BRENDA team
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
-
Manually annotated by BRENDA team
additional information
-
semi-quantitative RT-PCR analysis of temporal and spatial expression of AtLeuC and AtLeuD genes, overview. The tissue-specifis expression analysis reveals that the patterns of small subunits AtLeuD1 and AtLeuD2 expression are similar, but distinct from that of AtLeuD3
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
-
presence of different heterodimeric IPMIs in chloroplasts with distinct substrate specificities for Leu or glucosinolate metabolism
Manually annotated by BRENDA team
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
presence of different heterodimeric IPMIs in chloroplasts with distinct substrate specificities for Leu or glucosinolate metabolism. Import of the IPMI small subunit 2:GFP fusion protein into chloroplasts
Manually annotated by BRENDA team
additional information
-
analysis of subcellular localization of AtLeuC and AtLeuDs within Arabidopsis chloroplast compartments, overview
-
Manually annotated by BRENDA team
PDB
SCOP
CATH
ORGANISM
Campylobacter jejuni subsp. jejuni (strain IA3902)
Methanocaldococcus jannaschii (strain ATCC 43067 / DSM 2661 / JAL-1 / JCM 10045 / NBRC 100440)
Methanocaldococcus jannaschii (strain ATCC 43067 / DSM 2661 / JAL-1 / JCM 10045 / NBRC 100440)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain CDC 1551 / Oshkosh)
Mycobacterium tuberculosis (strain CDC 1551 / Oshkosh)
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
Streptococcus mutans serotype c (strain ATCC 700610 / UA159)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
70000 - 90000
-
sucrose density gradient centrifugation
5661
88500
-
sucrose density gradient centrifugation, gel filtration
5660
90000
Neurospora sp., Salmonella sp.
-
sucrose density gradient centrifugation, gel filtration
5658
90000
-
sucrose density gradient centrifugation, gel filtration
5659, 5663
additional information
-
complementation pair enzymes have higher molecular weights of 169000-185000
5660
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
?
Q58673
x * 46000 + x * 18000, SDS-PAGE
heterodimer
P9WK95, P9WQF5
IPMI exists as a functional LeuCD complex of two subunits: the large LeuC and the small LeuD subunit, structure determination and modeling, overview
heterodimer
P9WK95, P9WQF5
IPMI exists as a functional LeuCD complex of two subunits: the large LeuC and the small LeuD subunit, structure determination and modeling, overview. Presence of two LeuD subfamilies, structure modeling, overview
heterodimer
-
IPMI exists as a functional LeuCD complex of two subunits: the large LeuC and the small LeuD subunit, structure determination and modeling, overview. Presence of two LeuD subfamilies, structure modeling, overview, IPMI exists as a functional LeuCD complex of two subunits: the large LeuC and the small LeuD subunit, structure determination and modeling, overview
-
monomer
-
1*90000, SDS-PAGE
monomer
-
variants LeuD-1-156 and LeuD-1-168, gel-filtration
multimer
-
active as monomer, can aggregate to dimers or multimers in cell
tetramer
Neurospora sp.
-
trimer or tetramer of 2 different polypeptide subunits
trimer
Neurospora sp.
-
trimer or tetramer of 2 different polypeptide subunits
monomer
-
variants LeuD-1-156 and LeuD-1-168, gel-filtration
-
additional information
-
amino acid analysis, protein contains hinge regions that are very sensitive to proteolytic attack
additional information
P9WK95, P9WQF5
structure and oligomeric state of Mtb-LeuD in the crystal structure, overview
additional information
-
structure and oligomeric state of Mtb-LeuD in the crystal structure, overview
-
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
sitting drop vapor diffusion method, using 25% (w/v) polyethylene glycol monomethyl ether 2000 and 0.1 MTris-HCl (pH 7.9)
Q58673
sitting-drop vapour-diffusion methodand hanging-drop vapour diffusion at 20C, structures of oxidized and reduced forms of the large subunit of isopropylmalate isomerase (ox-MJ0499 and red-MJ0499, respectively) are reported at 1.8 and 2.7 A resolution, respectively. Significant large conformational changes are observed in the active site of red-MJ0499 when compared with ox-MJ0499
P81291
small subunit LeuD variants, X-ray diffraction structure determination and analysis at resolutions of 2.0 A for LeuD_1-156, 1.2 A for LeuD_1-168, and 2.5 A for LeuD_1-186, respectively
P9WK95, P9WQF5
variants LeuD-1-156 and LeuD-1-168, by sitting-drop vapour-diffusion method, crystals of LeuD-1-156 belong to the hexagonal system (space group P6122 or P6522) with up to four subunits in the asymmetric unit, whereas the crystals of LeuD-1-168 belong to the monoclinic system (space group P21) with two subunits in the asymmetric unit. Both crystals diffract to beyond 2.0 A resolution
-
crystal structure isopropylmalate isomerase small subunit. Four molecules create an interlocked assembly with intermolecular disulfide linkages having a skewed 222 point-group symmetry. The structure reveals the formation of intermolecular disulfide linkages, and it provides insight into the dual substrate specificity of the enzyme
O59393
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6 - 6.5
Neurospora sp.
-
unstable at pH 7.0
5658
6
-
unstable at pH 7.0 or above
5659
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
45
Neurospora sp., Salmonella sp.
-
95% loss of activity after 10 min
5658
45
-
50% loss of activity after 2 min
5661
55
-
half-life: 4.5 h, in stabilizing ammonium sulfate buffer
5659
62
-
half-life: 1 min, in stabilizing ammonium sulfate buffer
5659
additional information
-
very heat sensitive, high ionic strenght stabilizes enzyme against heat inactivation
5659
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
10% inactivation at 0C in phosphate buffer at pH 7 or above
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loss of activity after repeated freezing and thawing
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loss of activity after repeated freezing and thawing
Neurospora sp.
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substrate stabilizes
Neurospora sp.
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unstable in phosphate buffer at pH 7.0, 10% loss of activity per h at 0C
Neurospora sp.
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half-life 2-3 h in potassium phosphate buffer, ph 6.8., 0-5C, beta-isopropylmalate at 1 mM increases half-life to 15 h
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stabilized by high concentrations of glycerol and ammonium sulfate which also inhibit activity, half-life with 50% glycerol, 2.07 M ammonium sulfate, and 30% glycerol plus 830 mM ammonium sulfate: 50 h, 57 h, and 90 h, respectively, instead of 2-3 h without, stabilization probably due to changes in intramolecular interactions
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loss of activity after repeated freezing and thawing
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substrate stabilizes
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unstable in phosphate buffer at pH 7.0, 10% loss of activity per h at 0C
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, sodium phosphate buffer, pH 7.2, 6 M urea, several days
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
nickel-affinity column chromatography and Superdex 200 gel filtration
Q58673
variants LeuD-1-156 and LeuD-1-168, by centrifugation, on Ni-NTA column and by gel filtration, more than 95% pure
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Neurospora sp.
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression of FLAG-tagged LeuC and of the three LeuD isoforms in transgenic Arabidosis thaliana plants, interaction analysis, overview
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full length reading frame of IPMI small subunit 2 fused in frame to the 5' end of the gene encoding the green fluorescent protein in the vector psmGFP4. Construct transiently expressed in tobacco protoplasts and stably transformed into Arabidopsis plants
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
expressed in Escherichia coli BL21(DE3) cells
Q58673
expression in Escherichia coli BL21(DE3) Star cells
P81291
expression of His6-tagged LeuD in Escherichia coli strain BL21(DE3)
P9WK95, P9WQF5
variants LeuD-1-156 and LeuD-1-168, ligated into the pETM-11 expression vector, expressed in Escherichia coli BL21(DE3) RP
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
in ipmi ssu2-1 plants, absence of mature mRNAs in homozygous plants and the knockout of these genes by T-DNAs
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
in ipmi ssu3-1 plants, absence of mature mRNAs in homozygous plants and the knockout of these genes by T-DNAs
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
in the ipmilsu1-2mutant mRNA levels are reduced to 65 and 50% of the wild-type standard in seedlings and rosette leaves, respectively, while IPMI large subunit 1 transcript levels are reduced to 40 and 27% of wild-type in these tissues of ipmi lsu1-1 plants. Strongest reduction in ipmi lsu1-3 seedlings (21% of wild-type), while the IPMI large subunit 1 mRNA level in ipmi lsu1-3 reaches about 29% of the wild-type level in leaves
Q94AR8, Q9LYT7, Q9ZW84, Q9ZW85
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
G456D
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mutation in the 3-isopropylmalate dehydratase large subunit gene (leuC). Introduction of the leuC mutation into a defined L-lysine producer, AHD-2 (hom59 and lysC311), by allelic replacement leads to the phenotype of a partial requirement for L-leucine and approximately 14% increased L-lysine production
additional information
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construction of AtLeuD1 T-DNA knockout mutant and AtLeuD -RNAi plants, i.e. specific AtLeuD2 -RNAi, AtLeuD -RNAi, atleud1 /AtLeuD2 -RNAi and atleud1 /AtLeuD -RNAi. Lethal phenotype of the atleud3 mutant
G456D
Corynebacterium glutamicum B-6
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mutation in the 3-isopropylmalate dehydratase large subunit gene (leuC). Introduction of the leuC mutation into a defined L-lysine producer, AHD-2 (hom59 and lysC311), by allelic replacement leads to the phenotype of a partial requirement for L-leucine and approximately 14% increased L-lysine production
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additional information
P9WK95, P9WQF5
construction of three C-terminally truncated variants of small subunit LeuD
additional information
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construction of three C-terminally truncated variants of small subunit LeuD
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