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Literature summary for 3.1.8.2 extracted from

  • Li, D.; Zhang, Y.; Song, H.; Lu, L.; Liu, D.; Yuan, Y.
    Aminoalcohol-induced activation of organophosphorus hydrolase (OPH) towards diisopropylfluorophosphate (DFP) (2017), PLoS ONE, 12, e0169937 .
    View publication on PubMedView publication on EuropePMC

Activating Compound

Activating Compound Comment Organism Structure
1,1',1''-nitrilotri(propan-2-ol)
-
Sphingobium fuliginis
1-amino-2-propanol
-
Sphingobium fuliginis
2-amino-2-methyl-1-propanol
-
Sphingobium fuliginis
diethanolamine
-
Sphingobium fuliginis
Monoethanolamine
-
Sphingobium fuliginis
additional information organophosphorus hydrolase (OPH)-catalyzed hydrolysis of diisopropylfluorophosphate (DFP), an analogue molecule of G-type warfare agents, is activated by aminoalcohols. OPH reaction are remarkably increased in the buffers containing aminoalcohols with C2 between nitrogen and oxygen in their structures, including triethanolamine (TEA), diethanolamine, monoethanolamine, 1-amino-2-propanol, 2-amino-2-methyl-1-propanol, and triisopropanolamine. The activation of OPH by aminoalcohol buffers can be attributed to the reduction of fluoride inhibition, which would be beneficial to the hydrolase-based detoxification of organophosphofluoridate Sphingobium fuliginis
triethanolamine best activator, EDTA abolishes the triethanolamine-induced activation of enzyme OPH Sphingobium fuliginis

Cloned(Commentary)

Cloned (Comment) Organism
gene opd, recombinant expression of His6-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3), subcloning in Escherichia coli strain DH5alpha Sphingobium fuliginis

Protein Variants

Protein Variants Comment Organism
F132Y site-directed mutagenesis, the mutant exhibits a 3fold increased activity with diisopropyl fluorophosphate compared to wild-type Sphingobium fuliginis
F132Y/ L140Y site-directed mutagenesis, the mutant exhibits a several folds increased activity with diisopropyl fluorophosphate compared to wild-type Sphingobium fuliginis
L140Y site-directed mutagenesis, the mutant exhibits a 5fold increased activity with diisopropyl fluorophosphate compared to wild-type Sphingobium fuliginis

Inhibitors

Inhibitors Comment Organism Structure
EDTA inhibits wild-type and mutant enzymes at 50 mM Sphingobium fuliginis
fluoride product fluoride effectively inhibits organophosphorus hydrolase-catalyzed hydrolysis of diisopropylfluorophosphate (DFP) by a linear mixed inhibition, which is partially released by triethanolamine adding at initial or later reaction stage Sphingobium fuliginis
additional information the activation of organophosphorus hydrolase by aminoalcohol buffers can be attributed to the reduction of fluoride inhibition, which would be beneficial to the hydrolase-based detoxification of organophosphofluoridate Sphingobium fuliginis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten kinetic analysis, substrate diisopropyl fluorophosphate in presence of aminoalcohol activators Sphingobium fuliginis
2.92
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM 1-amino-2-propanol Sphingobium fuliginis
2.95
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM 2-amino-2-methyl-1-propanol Sphingobium fuliginis
2.99
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 1 mM NaF Sphingobium fuliginis
3.01
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme Sphingobium fuliginis
3.08
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM ethanol Sphingobium fuliginis
3.1
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y/L140Y Sphingobium fuliginis
3.15
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 400 mM monoethanolamine Sphingobium fuliginis
3.18
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM methanol Sphingobium fuliginis
3.2
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y Sphingobium fuliginis
3.2
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant L140Y Sphingobium fuliginis
3.35
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 400 mM diethanolamine Sphingobium fuliginis
3.38
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM glycerol Sphingobium fuliginis
3.57
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 2 mM NaF Sphingobium fuliginis
3.87
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 200 mM thiodipropanol and 1 mM NaF Sphingobium fuliginis
3.97
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM thiodiglycol Sphingobium fuliginis
4
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y/L140Y, with 300 mM triethanolamine Sphingobium fuliginis
4.06
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM thiodipropanol and 1 mM NaF Sphingobium fuliginis
4.1
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triethanolamine Sphingobium fuliginis
4.17
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM methylamine and 300 mM ethanol Sphingobium fuliginis
4.2
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y, with 300 mM triethanolamine Sphingobium fuliginis
4.21
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triisopropanolamine Sphingobium fuliginis
4.23
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 100 mM thiodipropanol and 1 mM NaF Sphingobium fuliginis
4.32
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM ethylamine and 300 mM ethanol Sphingobium fuliginis
4.32
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 400 mM triethanolamine Sphingobium fuliginis
4.41
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM diethylamine and 300 mM ethanol Sphingobium fuliginis
4.44
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM methylamine Sphingobium fuliginis
4.45
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 3 mM NaF Sphingobium fuliginis
4.45
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triethylamine and 300 mM ethanol Sphingobium fuliginis
4.5
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant L140Y, with 300 mM triethanolamine Sphingobium fuliginis
4.52
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triethylamine Sphingobium fuliginis
4.54
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 200 mM thiodipropanol and 2 mM NaF Sphingobium fuliginis
4.56
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM ethylamine Sphingobium fuliginis
4.58
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM thiodipropanol and 2 mM NaF Sphingobium fuliginis
4.61
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM 3-amino-1-propanol Sphingobium fuliginis
4.67
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM diethylamine Sphingobium fuliginis
4.77
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM thiodipropanol Sphingobium fuliginis
4.87
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 100 mM thiodipropanol and 2 mM NaF Sphingobium fuliginis
4.89
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM thiodipropanol and 3 mM NaF Sphingobium fuliginis
5.04
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 200 mM thiodipropanol and 3 mM NaF Sphingobium fuliginis
5.58
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 100 mM thiodipropanol and 3 mM NaF Sphingobium fuliginis

Metals/Ions

Metals/Ions Comment Organism Structure
additional information the binuclear metal center, comprised of either two equivalents of Zn2+ in the native enzyme or mixed metal ions Cd2+, Co2+, Mn2+, and Ni2+ in the metal-substituted catalysts, which generates the activated water molecules that initiate nucleophilic attack on the phosphorus atom of the substrate, resulting in the cleavage of phosphoester bond and the release of leaving group Sphingobium fuliginis
Zn2+ required two equivalents of Zn2+ per native enzyme molecule Sphingobium fuliginis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
diisopropyl fluorophosphate + H2O Sphingobium fuliginis an analogue molecule of G-type warfare agents diisopropyl phosphate + fluoride
-
?

Organism

Organism UniProt Comment Textmining
Sphingobium fuliginis P0A433 formerly Flavobacterium sp. strain ATCC 27551
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His6-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography Sphingobium fuliginis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(RS)-propan-2-yl methylphosphonofluoridate + H2O i.e. sarin Sphingobium fuliginis isopropyl phosphate methylphosphonate + fluoride
-
?
3-[fluoro(methyl)phosphoryl]oxy-2,2-dimethylbutane + H2O i.e. soman Sphingobium fuliginis Pinacolyl methylphosphonate + fluoride
-
?
diethyl fluorophosphate + H2O
-
Sphingobium fuliginis diethyl phosphate + fluoride
-
?
diisopropyl fluorophosphate + H2O
-
Sphingobium fuliginis diisopropyl phosphate + fluoride
-
?
diisopropyl fluorophosphate + H2O an analogue molecule of G-type warfare agents Sphingobium fuliginis diisopropyl phosphate + fluoride
-
?
additional information the substrates of enzyme OPH include organophosphate insecticides paraoxon, parathion, methylparathion, coumaphos, and diazinon, as well as potent nerve agents sarin, soman, and their analogue diisopropylfluorophosphate (DFP), cf. EC 3.1.8.1 and 3.1.8.2 Sphingobium fuliginis ?
-
?

Synonyms

Synonyms Comment Organism
More cf. EC 3.1.8.1 Sphingobium fuliginis
opd
-
Sphingobium fuliginis
organophosphorus hydrolase
-
Sphingobium fuliginis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Sphingobium fuliginis

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.0115
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM glycerol Sphingobium fuliginis
0.0117
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme Sphingobium fuliginis
0.0123
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM ethanol Sphingobium fuliginis
0.0123
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM methanol Sphingobium fuliginis
0.0142
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM 3-amino-1-propanol Sphingobium fuliginis
0.0148
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triethylamine Sphingobium fuliginis
0.0151
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM thiodipropanol Sphingobium fuliginis
0.0155
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triethylamine and 300 mM ethanol Sphingobium fuliginis
0.0163
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM diethylamine Sphingobium fuliginis
0.0178
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM diethylamine and 300 mM ethanol Sphingobium fuliginis
0.018
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM ethylamine Sphingobium fuliginis
0.0193
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM methylamine Sphingobium fuliginis
0.0195
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM 1-amino-2-propanol Sphingobium fuliginis
0.0195
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM ethylamine and 300 mM ethanol Sphingobium fuliginis
0.0209
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 400 mM monoethanolamine Sphingobium fuliginis
0.0211
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM 2-amino-2-methyl-1-propanol Sphingobium fuliginis
0.0216
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM methylamine and 300 mM ethanol Sphingobium fuliginis
0.0295
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM thiodiglycol Sphingobium fuliginis
0.031
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant L140Y Sphingobium fuliginis
0.0312
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 400 mM diethanolamine Sphingobium fuliginis
0.0361
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triisopropanolamine Sphingobium fuliginis
0.042
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triethanolamine Sphingobium fuliginis
0.0477
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 400 mM triethanolamine Sphingobium fuliginis
0.048
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y/L140Y Sphingobium fuliginis
0.049
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y Sphingobium fuliginis
0.142
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant L140Y, with 300 mM triethanolamine Sphingobium fuliginis
0.182
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y, with 300 mM triethanolamine Sphingobium fuliginis
0.217
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y/L140Y, with 300 mM triethanolamine Sphingobium fuliginis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8
-
assay at Sphingobium fuliginis

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
3.21
-
fluoride pH 8.0, 25°C, recombinant wild-type enzyme Sphingobium fuliginis
5.17
-
fluoride pH 8.0, 25°C, recombinant wild-type enzyme, with 100 mM triethanolamine Sphingobium fuliginis
5.45
-
fluoride pH 8.0, 25°C, recombinant wild-type enzyme, with 200 mM triethanolamine Sphingobium fuliginis
7.51
-
fluoride pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triethanolamine Sphingobium fuliginis

General Information

General Information Comment Organism
additional information the metal-substituted catalysts generates the activated water molecules that initiate nucleophilic attack on the phosphorus atom of the substrate, resulting in the cleavage of phosphoester bond and the release of leaving group Sphingobium fuliginis

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.0031
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM 3-amino-1-propanol Sphingobium fuliginis
0.0032
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM thiodipropanol Sphingobium fuliginis
0.0033
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triethylamine Sphingobium fuliginis
0.0034
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM glycerol Sphingobium fuliginis
0.0035
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM diethylamine Sphingobium fuliginis
0.0035
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triethylamine and 300 mM ethanol Sphingobium fuliginis
0.0039
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme Sphingobium fuliginis
0.0039
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM methanol Sphingobium fuliginis
0.004
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM diethylamine and 300 mM ethanol Sphingobium fuliginis
0.004
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM ethanol Sphingobium fuliginis
0.004
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM ethylamine Sphingobium fuliginis
0.0044
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM methylamine Sphingobium fuliginis
0.0045
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM ethylamine and 300 mM ethanol Sphingobium fuliginis
0.0052
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM methylamine and 300 mM ethanol Sphingobium fuliginis
0.0066
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 400 mM monoethanolamine Sphingobium fuliginis
0.0067
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM 1-amino-2-propanol Sphingobium fuliginis
0.0072
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM 2-amino-2-methyl-1-propanol Sphingobium fuliginis
0.0074
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM thiodiglycol Sphingobium fuliginis
0.0086
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triisopropanolamine Sphingobium fuliginis
0.0093
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 400 mM diethanolamine Sphingobium fuliginis
0.0098
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant L140Y Sphingobium fuliginis
0.01
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 300 mM triethanolamine Sphingobium fuliginis
0.011
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant wild-type enzyme, with 400 mM triethanolamine Sphingobium fuliginis
0.0153
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y Sphingobium fuliginis
0.016
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y/L140Y Sphingobium fuliginis
0.032
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant L140Y, with 300 mM triethanolamine Sphingobium fuliginis
0.043
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y, with 300 mM triethanolamine Sphingobium fuliginis
0.054
-
diisopropyl fluorophosphate pH 8.0, 25°C, recombinant mutant F132Y/L140Y, with 300 mM triethanolamine Sphingobium fuliginis