Information on EC 3.7.1.B1 - 2,4-diacetylphloroglucinol hydrolase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
3.7.1.B1
preliminary BRENDA-supplied EC number
RECOMMENDED NAME
GeneOntology No.
2,4-diacetylphloroglucinol hydrolase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
2,4-diacetylphloroglucinol + H2O = acetylphloroglucinol + acetate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C-C-bond cleavage
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retro-Friedel-Crafts acylation
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
gene PSEBR_a2324
UniProt
Manually annotated by BRENDA team
gene PSEBR_a2324
UniProt
Manually annotated by BRENDA team
gene phlG
UniProt
Manually annotated by BRENDA team
gene phlG
UniProt
Manually annotated by BRENDA team
gene phlG
UniProt
Manually annotated by BRENDA team
gene phlG
UniProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
malfunction
metabolism
physiological function
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
2,4-diacetylphloroglucinol + H2O
4-acetylphloroglucinol + acetate
show the reaction diagram
additional information
?
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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
2,4-diacetylphloroglucinol + H2O
4-acetylphloroglucinol + acetate
show the reaction diagram
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Co2+
specifically and significantly activates the enzyme, shows a hyperbolic saturation curve in the micromolar range, binding to the enzyme might induce a conformational change that increases the catalytic effect of Zn2+ or might forma a distinct catalytic site besides that for Zn2+
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Cu2+
slight inhibition
diethyldicarbonate
60% inhibition at 0.5 mM
monoacetylphloroglucinol
competitive inhibition
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pyoluteorin
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strong inhibition of the enzyme, another antifungal compound produced by the bacterium
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additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.024 - 0.873
2,4-diacetylphloroglucinol
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.000026 - 33
2,4-diacetylphloroglucinol
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00062 - 17
2,4-diacetylphloroglucinol
206963
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.087
monoacetylphloroglucinol
recombinant enzyme, pH 7.0, 25°C
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
9.1
purified recombinant enzyme, pH 7.0, 25°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 8
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10% of maximal activity at pH 7.6, inactive at pH 8.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
16 - 42
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50% of maximal activity within this range, optimum at 30°C
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
35000
2 * 35000, recombinant enzyme, SDS-PAGE
40000
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x * 40000, recombinant His-tagged enzyme, SDS-PAGE
65000
recombinant enzyme, gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified recombinant enzyme, using a precipitant consisting of 100 mM citric acid, pH 5.0, 1.0 M lithium chloride, 8% w/v PEG 6000, X-ray diffraction structure determination and analysis at 1.87 A resolution
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purified wild-type and selenomethionine-labeled enzymes, hanging drop vapor diffusion technique, mixing of 0.001 ml of 20 mg/ml protein sample with an equal volume of the reservoir solution containing 20% PEG 4000, 17% isopropyl alcohol, and 0.1 M sodium citrate, pH 5.6, crystals appear in 1-2 days and reach maximum size in 1 week, at 16°C, X-ray diffraction structure determination and analysis at 2.0 A resolution, MAD phasing method, modeling
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
47 - 55
purified recombinant enzyme, 50% activity remaining at 47°C, inactivation at 55°C
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
2-mercaptoethanol or DTT is required for stability, the purified enzyme is rapidly inactivates without stabilizing compounds, the enzyme can be partially reactivated by 10 mM 2-mercaptoethanol to regain 20% of its initial activity, the enzyme is irreversibly inactivated by freezing and thawing
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant enzyme
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recombinant enzyme 22fold from Escherichia coli strain XL-1 Blue to homogeneity by ammonium sulfate fractionation, gel filtration, two steps of anion exchange chromatography, and hydrophobic interaction chromatograpy
recombinant His6-tagged enzyme from Escherichia coli strain BL21(DE3)/pME8032 by nickel affinity chromatography
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recombinant wild-type and mutant enzymes from Escherichia coli
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
gene phlG, from locus phlHGFACBDEI, DNA and amino acid sequence determination and analysis, sequence comparison, recombinant expression in Escherichia coli strain XL-1 Blue
gene phlG, located upstream of the phlACBD biosynthetic operon, between the phlF and phlH genes which encode pathway-specific regulators, genetic structure of the operon, functional recombinant expression of soluble His6-tagged enzyme in Escherichia coli strain BL21(DE3)/pME8032
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gene phlG, recombinant expression of wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) or B834 (DE3), the latter for the selenomethionine-labeled enzyme
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gene phlg, sequence comparison
gene PSEBR_a2324, DNA and amino acid sequence determination and analysis
recombinant expression
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
expression of gene phlG is not influenced by the substrate 2,4-diacetylphloroglucinol or the degradation product monoacetylphloroglucinol but is subject to positive control by the GacS/GacA two-component system and to negative control by the pathway-specific regulators PhlF and PhlH
gene phlG expression regulation study, overview
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
H214Q
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site-directed mutagenesis, mutation of a polar residue that is predicted to form hydrogen bond with the polar groups of the substrate, the mutant shows reduced activity compared to the wild-type enzyme
N132A
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site-directed mutagenesis, mutation of a polar residue that is predicted to form hydrogen bond with the polar groups of the substrate, the mutant shows reduced activity compared to the wild-type enzyme
Y229A
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site-directed mutagenesis, mutation of a polar residue that is predicted to form hydrogen bond with the polar groups of the substrate, the mutant shows reduced activity compared to the wild-type enzyme
Y229F
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site-directed mutagenesis, mutation of a polar residue that is predicted to form hydrogen bond with the polar groups of the substrate, the mutant shows reduced activity compared to the wild-type enzyme
E160A
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site-directed mutagenesis, the mutation of the zinc binding residue leads to reduced activity compared to the wild-type enzyme
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E274A
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site-directed mutagenesis, the mutation of the zinc binding residue leads to reduced activity compared to the wild-type enzyme
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H214A
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site-directed mutagenesis, mutation of a polar residue that is predicted to form hydrogen bond with the polar groups of the substrate, the mutant shows reduced activity compared to the wild-type enzyme
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Y121A
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site-directed mutagenesis, mutation of a polar residue that is predicted to form hydrogen bond with the polar groups of the substrate, the mutant shows reduced activity compared to the wild-type enzyme
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Y229F
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site-directed mutagenesis, mutation of a polar residue that is predicted to form hydrogen bond with the polar groups of the substrate, the mutant shows reduced activity compared to the wild-type enzyme
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additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
agriculture
additional information