Information on EC 2.1.2.11 - 3-methyl-2-oxobutanoate hydroxymethyltransferase

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

EC NUMBER
COMMENTARY hide
2.1.2.11
-
RECOMMENDED NAME
GeneOntology No.
3-methyl-2-oxobutanoate hydroxymethyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
5,10-methylenetetrahydrofolate + 3-methyl-2-oxobutanoate + H2O = tetrahydrofolate + 2-dehydropantoate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydroxymethyl group transfer
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-
-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Biosynthesis of secondary metabolites
-
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Metabolic pathways
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Pantothenate and CoA biosynthesis
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pantothenate biosynthesis
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phosphopantothenate biosynthesis I
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phosphopantothenate biosynthesis III
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SYSTEMATIC NAME
IUBMB Comments
5,10-methylenetetrahydrofolate:3-methyl-2-oxobutanoate hydroxymethyltransferase
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CAS REGISTRY NUMBER
COMMENTARY hide
56093-17-5
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
wild-type strains biA1 and yA2, strain G1765
Uniprot
Manually annotated by BRENDA team
strains 168, BD170, UR1, UR4
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-
Manually annotated by BRENDA team
strain K12
-
-
Manually annotated by BRENDA team
BCG, strain 1173-P2, live vaccine against tuberculosis, used for superficial bladder cancer immunotherapy
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-
Manually annotated by BRENDA team
strain KIT 10468
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-
Manually annotated by BRENDA team
genomic gene panB, and a second putative MOHMT enzyme, RHE_PE00443, similar to the product of panB, is encoded on plasmid p42e
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Manually annotated by BRENDA team
genomic gene panB, and a second putative MOHMT enzyme, RHE_PE00443, similar to the product of panB, is encoded on plasmid p42e
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Manually annotated by BRENDA team
serovar typhimurium
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
metabolism
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
5,10-methylenetetrahydrofolate + 2-oxo-4-methylthiobutyrate
tetrahydrofolate + ?
show the reaction diagram
-
about 50% of the activity with 3-methyl-2-oxobutanoate
-
-
?
5,10-methylenetetrahydrofolate + 2-oxobutyrate
?
show the reaction diagram
5,10-methylenetetrahydrofolate + 2-oxobutyrate
tetrahydrofolate + ?
show the reaction diagram
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about 50% of the activity with 3-methyl-2-oxobutanoate
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-
?
5,10-methylenetetrahydrofolate + 2-oxopentanoate
?
show the reaction diagram
5,10-methylenetetrahydrofolate + 2-oxopentanoate
tetrahydrofolate + ?
show the reaction diagram
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about 30% of the activity with 3-methyl-2-oxobutanoate
-
-
?
5,10-methylenetetrahydrofolate + 3-methyl-2-oxobutanoate
tetrahydrofolate + 2-dehydropantoate
show the reaction diagram
5,10-methylenetetrahydrofolate + 3-methyl-2-oxopentanoate
?
show the reaction diagram
5,10-methylenetetrahydrofolate + 3-methyl-2-oxopentanoate
tetrahydrofolate + ?
show the reaction diagram
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about 65% of the activity with 3-methyl-2-oxobutanoate
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?
5,10-methylenetetrahydrofolate + pyruvate
tetrahydrofolate + ?
show the reaction diagram
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about 20% of the activity with 3-methyl-2-oxobutanoate
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-
?
formaldehyde + 3-methyl-2-oxobutanoate
2-dehydropantoate
show the reaction diagram
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enzyme catalyzes methylenetetrahydrofolate-independent hydroxymethyltransferase reaction between free formaldehyde and alpha-ketoisovalerate, formaldehyde is unlikely to be the natural substrate
-
?
formaldehyde + tetrahydrofolate
methylentetrahydrofolate
show the reaction diagram
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
5,10-methylenetetrahydrofolate + 3-methyl-2-oxobutanoate
tetrahydrofolate + 2-dehydropantoate
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
5,10-methylenetetrahydrofolate
tetrahydrofolate
additional information
-
no requirement of pyridoxal 5-phosphate as cofactor
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
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Km: 0.27 mM, the reaction is divalent metal-dependent with descending order of preference: Mg2+, Zn2+, Co2+, Ni2+, Ca2+; substrate enolization is divalent metal-dependent with a preference for metal ions in decreasing order: Mg2+, Zn2+, Co2+, Ni2+, Ca2+
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3-Methyl-2-butanone
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5 mM, 27% inhibition
coenzyme A
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above 1 mM
D-valine
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5 mM, 16% inhibition
formaldehyde
Isovalerate
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5 mM, 39% inhibition
L-valine
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5 mM, 23% inhibition
Pantoate
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0.05 mM or above
pantothenate
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0.5 mM or above
pyruvate
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5 mM, 38% inhibition
tetrahydrofolate
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2.9
2-oxobutyrate
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25
2-Oxopentanoate
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0.24 - 1.1
3-methyl-2-oxobutanoate
5.9
3-methyl-2-oxopentanoate
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0.82
5,10-methylenetetrahydrofolate
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5.9
formaldehyde
0.15 - 0.16
ketopantoate
0.18
tetrahydrofolate
0.25
tetrahydropteroyldiglutamate
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-
0.29
tetrahydropteroylheptaglutamate
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0.17
tetrahydropteroylhexaglutamate
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0.33
tetrahydropteroylmonoglutamate
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0.1
Tetrahydropteroylpentaglutamate
0.18
tetrahydropteroyltriglutamate
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-
additional information
additional information
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.78 - 0.783
3-methyl-2-oxobutanoate
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0025
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strain BD170
0.00254
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strain UR1
0.00258
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strain UR4
0.00262
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strain 168
0.007
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strain K12
0.01 - 0.027
recombinant enzyme expressed in Escherichia coli
0.019
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recombinant enzyme
0.162
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strain Hfr3000 YA139/pCEJ02
0.348
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strain Hfr3000 YA139/pCEJ01
0.666
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strain Hfr3000 YA139/pSAL38
3890
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purified recombinant enzyme from strain Hfr3000 YA139/pCEJ01
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 7.5
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; rate of substrate enolization is pH-dependent with optimal activity in the range of
8
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at or below
additional information
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 9
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about 50% of activity maximum at pH 6 and 9, inactive below pH 5
10
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no ketopantoate formation above pH 10
additional information
-
stable and active over a broad pH-range
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
70 - 80
-
reverse reaction, activity decreases rapidly above 80C
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Burkholderia pseudomallei (strain 1710b)
Burkholderia thailandensis (strain E264 / ATCC 700388 / DSM 13276 / CIP 106301)
Escherichia coli (strain K12)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Neisseria meningitidis serogroup B (strain MC58)
Neisseria meningitidis serogroup B (strain MC58)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
120000
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gel filtration
174000
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recombinant enzyme, gel filtration
255000
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sedimentation equilibrium
285000
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gel filtration
340000
recombinant enzyme expressed in Escherichia coli, gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 29337, calculated from the amino acid sequence, x * 35000, recombinant enzyme expressed in Escherichia coli BL21, SDS-PAGE
decamer
hexamer
octamer
8 * 37700, calculated from the amino acid sequence, 8 * 40000, recombinant enzyme expressed in Escherichia coli, SDS-PAGE
tetramer
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recombinant enzyme expressed in Escherichia coli BL21 (DE3): 4 * 29000, SDS-PAGE, 4 * 29202, electrospray ionization/mass spectrometry, 4 * 29366, calculated from the amino acid sequence
additional information
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enzyme is degraded by the proteasome. Maintenance of the physiological levels of 3-methyl-2-oxobutanoate hydroxymethyltransferase and of malonyl-CoA acyl carrier protein transacylase requires Mycobacterium proteasomal ATPase and proteasome accessory factor in addition to proteasome protease activity
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
no modification
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
comparative analysis of the Escherichia coli ketopantoate hydroxymethyltransferase crystal structure confirms that it is a member of the (betaalpha)8 phosphoenolpyruvate/pyruvate superfamily
hanging drop vapor diffusion, crystal structure at 1.9 A resolution in complex with its product ketopantoate, the enzyme adopts the (betaalpha)8 barrel fold and the active site contains a ketopantoate bidentately coordiated to Mg2+
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.5
-
rapid inactivation below
485700
5 - 10
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stable
485700
5.5
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loss of proper protein folding at pH values lower than 5.5
485710
additional information
-
stable and active over a broad pH-range
485701
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
55
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treatment at 55C largely destroys enzyme in crude extract, partially purified enzyme is more heat-stabile
80
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rapid denaturation above
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate fractionation inactivates, resistant to urea denaturation
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not inactivated by borohydride reduction in the presence of excess substrates
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4C, 100 mM potassium phosphate, pH 6.8, 1 mM EDTA, 0.5 mM dithiothreitol, 6 months, almost no loss of activity
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
19.3fold purification of recombinant enzyme 50fold overexpressed in E. coli panB mutant Hfr3000 YA139/pCEJ01
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affinity purification of recombinant PanB
purification of recombinant enzyme expressed in Escherichia coli BL21 (DE3)
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purification of recombinant enzyme overexpressed in Escherichia coli BL21
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strain K12, 2450fold purification
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
gene panB, phylogenetic analysis, genes panBC in Rhizobiales with multipartite genomes, overview
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panB gene coding for enzyme is located together with panE in the intervall purE-tre of the chromosome
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panB gene encoding enzyme is cloned and overexpressed in Escherichia coli BL21 (DE3)
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panB gene encoding enzyme is cloned, sequenced and 50fold overexpressed in Escherichia coli panB mutant Hfr3000 YA139 containing plasmid pCEJ01, gene is 792 bp long and encodes a protein of 264 amino acids, gene is likely to be cotranscribed with at least one other gene, panB, panC and panD genes are closely clustered at 3.1 min of the Escherichia coli K12 genetic map
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panB gene encoding enzyme is cloned, sequenced and expressed in Escherichia coli as an active octameric enzyme, ORF encodes a protein of 349 amino acids, panB gene is closely linked to nimO gene on Aspergillus nidulans linkage group VII
panB gene encoding enzyme is cloned, sequenced and overexpressed in Escherichia coli BL21, ORF of 846 bp encodes a protein of 281 amino acids, N-terminal 37 amino acids are essential for enzyme function
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panB gene encoding enzyme is clustered together with other pantothenate biosynthetic genes panC and panD at 3 min of the chromosome map, panBCD operon
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PCR amplification of panB gene, which is identical to that of Mycobacterium bovis BCG
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pharmacology
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