Information on EC 4.2.1.42 - galactarate dehydratase

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The expected taxonomic range for this enzyme is: Bacteria

EC NUMBER
COMMENTARY
4.2.1.42
-
RECOMMENDED NAME
GeneOntology No.
galactarate dehydratase
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
galactarate = (2R,3S)-2,3-dihydroxy-5-oxohexanedioate + H2O
show the reaction diagram
-
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
elimination
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
D-galactarate degradation I
-
-
D-galactarate degradation II
-
-
degradation of sugar acids
-
-
Ascorbate and aldarate metabolism
-
-
SYSTEMATIC NAME
IUBMB Comments
galactarate hydro-lyase (5-dehydro-4-deoxy-D-glucarate-forming)
-
SYNONYMS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
A9CG74
Agrobacterium tumefaciens C58
-
-
-
Dehydratase, galactarate
-
-
-
-
Galactarate dehydrase
-
-
-
-
galactarate dehydratase
-
galactarate dehydratase
Oceanobacillus iheyensis DSM 14371
-
-
galactarate dehydratase III
-
-
galactarate dehydratase III
Agrobacterium tumefaciens C58
-
-
-
GalcD
-
-
-
-
GalrD-II
Oceanobacillus iheyensis DSM 14371
-
-
GalrD-III
Agrobacterium tumefaciens C58
-
-
-
m-galactarate dehydratase
-
-
m-galactarate dehydratase
Agrobacterium tumefaciens C58
-
-
-
CAS REGISTRY NUMBER
COMMENTARY
37290-78-1
-
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
Agrobacterium tumefaciens C58
-
-
-
Manually annotated by BRENDA team
strain CR63MA
-
-
Manually annotated by BRENDA team
Escherichia coli CR63MA
strain CR63MA
-
-
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
3-galactarate
2-dehydro-3-deoxy-D-glucarate + H2O
show the reaction diagram
Agrobacterium tumefaciens, Agrobacterium tumefaciens C58
-
-
-
?
D-Galactarate
5-Dehydro-4-deoxy-D-glucarate + H2O
show the reaction diagram
-
-
-
-
D-Galactarate
5-Dehydro-4-deoxy-D-glucarate + H2O
show the reaction diagram
-
-
-
-
D-Galactarate
5-Dehydro-4-deoxy-D-glucarate + H2O
show the reaction diagram
Escherichia coli CR63MA
-
-
-
-
D-Galactarate
5-Dehydro-4-deoxy-D-glucarate + H2O
show the reaction diagram
Escherichia coli CR63MA
-
-
-
-
D-Galactarate
?
show the reaction diagram
Escherichia coli, Escherichia coli CR63MA
-
enzyme of D-glucarate catabolism
-
-
-
D-galacturonate
3-deoxy-D-xylo-hexarate + 3-deoxy-D-lyxo-hexarate + H2O
show the reaction diagram
Agrobacterium tumefaciens, Agrobacterium tumefaciens C58
-
low level activity
-
?
galactarate
5-dehydro-4-deoxy D-glucarate + H2O
show the reaction diagram
-
-
?
galactarate
5-dehydro-4-deoxy D-glucarate + H2O
show the reaction diagram
Oceanobacillus iheyensis DSM 14371
-
-
-
?
galactarate
5-dehydro-4-deoxy-D-glucarate + H2O
show the reaction diagram
galactarate i.e. (2R,3S,4R,5S)-2,3,4,5-tetrahydroxyhexanedioate. The enzyme also catalyzes dehydration of L-talarate. The dehydration of L-talarate is accompanied by competing epimerization to galactarate. Little epimerization to L-talarate is observed in the dehydration of galactarate. On the basis of (1) structures of the wild type enzyme complexed with L-lyxarohydroxamate, an analogue of the enolate intermediate, and of the K197A mutant complexed with L-glucarate, a substrate for exchange of the R-proton, and (2) incorporation of solvent deuterium into galactarate in competition with dehydration, it is concluded that Lys197 functions as the galactarate-specific base and His328 functions as the L-talarate-specific base. The epimerization of L-talarate to galactarate that competes with dehydration can be rationalized by partitioning of the enolate intermediate between dehydration (departure of the 3-OH group catalyzed by the conjugate acid of His328) and epimerization (protonation on C2 by the conjugate acid of Lys197). Only galactarate and L-talarate are completely dehydrated by the enzyme. No other acid sugar results in detectable turnover
-
?
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
3-galactarate
2-dehydro-3-deoxy-D-glucarate + H2O
show the reaction diagram
Agrobacterium tumefaciens, Agrobacterium tumefaciens C58
-
-
-
?
D-Galactarate
?
show the reaction diagram
Escherichia coli, Escherichia coli CR63MA
-
enzyme of D-glucarate catabolism
-
-
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Mg2+
structure contains two Mg2+ ions located 10.4 A from one another, with one located in the canonical position in the (beta/alpha)7beta-barrel domain, the second is located in a site within the capping domain
additional information
-
no metal requirement
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
cacodylate
-
-
citrate
-
-
diphosphate
-
-
phosphate
-
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.08
3-galactarate
-
at pH 7.9 and 22C
-
0.4
galactarate
-
-
0.62
galactarate
wild-type, in 50 mM Hepes, pH 8.0, 5 mM MgCl2
0.8
galactarate
-
-
2
galactarate
mutant Y90F, in 50 mM Hepes, pH 8.0, 5 mM MgCl2
4.4
galactarate
mutant H45Q, in 50 mM Hepes, pH 8.0, 5 mM MgCl2
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.12
3-galactarate
Agrobacterium tumefaciens
-
at pH 7.9 and 22C
-
0.0013
galactarate
Oceanobacillus iheyensis
Q8EMJ9
mutant H45Q, in 50 mM Hepes, pH 8.0, 5 mM MgCl2
0.0061
galactarate
Oceanobacillus iheyensis
Q8EMJ9
mutant Y90F, in 50 mM Hepes, pH 8.0, 5 mM MgCl2
3.6
galactarate
Salmonella enterica subsp. enterica serovar Typhimurium str. LT2
Q8ZL58
pH 8.0, 25C
6.8
galactarate
Oceanobacillus iheyensis
Q8EMJ9
wild-type, in 50 mM Hepes, pH 8.0, 5 mM MgCl2
22
galactarate
Escherichia coli
-
-
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
1.5
3-galactarate
Agrobacterium tumefaciens
-
at pH 7.9 and 22C
197756
0.00029
galactarate
Oceanobacillus iheyensis
Q8EMJ9
mutant H45Q, in 50 mM Hepes, pH 8.0, 5 mM MgCl2
3374
0.00065
galactarate
Oceanobacillus iheyensis
Q8EMJ9
mutant R162N, in 50 mM Hepes, pH 8.0, 5 mM MgCl2
3374
0.0031
galactarate
Oceanobacillus iheyensis
Q8EMJ9
mutant Y90F, in 50 mM Hepes, pH 8.0, 5 mM MgCl2
3374
11
galactarate
Salmonella enterica subsp. enterica serovar Typhimurium str. LT2
Q8ZL58
pH 8.0, 25C
3374
11
galactarate
Oceanobacillus iheyensis
Q8EMJ9
wild-type, in 50 mM Hepes, pH 8.0, 5 mM MgCl2
3374
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7 - 9
-
low activity below pH 7.0 and above pH 9.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Escherichia coli (strain K12)
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
sitting drop vapor diffusion method, using 2 M ammonium sulfate and 100 mM Bis-Tris-HCl (pH 5.5)
-
three different crystal forms are grown by hanging drop vapor diffusion at room temperature: (1) selenomethionine (SeMet)-substituted wild type enzyme, (2) wild-type enzyme complexed with Mg2+ and L-lyxarohydroxamate, and (3) the K197A mutant enzyme complexed with Mg2+ and L-glucarate
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
galactarate stabilizes
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, column chromatography, and gel filtration
-
by centrifugation, sonication, on Ni-NTA column and by gel filtration
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
into a TOPO vector (pSGX3), and transformed into Escherichia coli TOP10 competent cells. Gene in the pSGX3 vector transformed into Escherichia coli strains XL1-Blue for transformation and BL21(DE3) for expression
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
H45Q
has no detectable activity
R162N
retains a small amount of activity
Y164F
has no detectable activity
Y90F
is catalytically impaired. Structure of the mutant in complex with Mg2+ and galactarate has a well-defined C-terminal segment through residue 387, well-ordered electron density for galactarate, and Mg2+ ions in both metal sites for both protomers comprising the asymmetric unit
H45Q
Oceanobacillus iheyensis DSM 14371
-
has no detectable activity
-
R162N
Oceanobacillus iheyensis DSM 14371
-
retains a small amount of activity
-
Y164F
Oceanobacillus iheyensis DSM 14371
-
has no detectable activity
-
Y90F
Oceanobacillus iheyensis DSM 14371
-
is catalytically impaired. Structure of the mutant in complex with Mg2+ and galactarate has a well-defined C-terminal segment through residue 387, well-ordered electron density for galactarate, and Mg2+ ions in both metal sites for both protomers comprising the asymmetric unit
-
K197A
inactive mutant enzyme, the structure of the K197A mutant enzyme complexed with Mg2+ and L-glucarate is determined by molecular replacement using the SeMet-substituted STM3697 structure as the search model