Please wait a moment until all data is loaded. This message will disappear when all data is loaded.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
2,2'-dihydroxybiphenyl + NADH + O2
2,2',3-trihydroxybiphenyl + NAD+ + H2O
-
-
-
-
?
2,2'-dihydroxybiphenyl + NADPH + O2
2,2',3-trihydroxybiphenyl + NADP+ + H2O
-
-
-
-
?
2,3-dihydroxybiphenyl + NADH + H+ + O2
?
-
-
-
?
2,5-dihydroxybiphenyl + NADH + O2
2,3,5-trihydroxybiphenyl + NAD+ + H2O
-
-
-
-
?
2-ethylphenol + NADH + O2
1,2-dihydroxy-3-ethylbenzene + NAD+ + H2O
-
-
-
-
?
2-hydroxybiphenyl + NADH + H+ + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
2-hydroxybiphenyl + NADPH + O2
2,3-dihydroxybiphenyl + NADP+ + H2O
2-hydroxybiphenyl + O2 + NADH + H+
3-phenylcatechol + NAD+ + H2O
2-methylphenol + NADH + O2
1,2-dihydroxy-3-methylbenzene + NAD+ + H2O
-
-
-
-
?
2-propylphenol + NADH + O2
1,2-dihydroxy-3-propylbenzene + NAD+ + H2O
2-sec-butylphenol + NADH + O2
2-sec-butylcatechol + NAD+ + H2O
2-sec-butylphenol + NADPH + O2
2-sec-butylcatechol + NADP+ + H2O
-
-
-
-
?
2-tert-butylphenol + NADH + O2
1,2-dihydroxy-3-tert-butylbenzene + NAD+ + H2O
3-hydroxybiphenyl + NADH + H+ + O2
3,4-dihydroxybiphenyl + NAD+ + H2O
activity of enzyme mutant M321A
-
-
?
guaiacol + NADH + O2
2,3-dihydroxy-methoxybenzene + NAD+ + H2O
additional information
?
-
2-hydroxybiphenyl + NADH + H+ + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
-
-
?
2-hydroxybiphenyl + NADH + H+ + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
-
-
-
?
2-hydroxybiphenyl + NADH + H+ + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
-
-
?
2-hydroxybiphenyl + NADH + H+ + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
it is suggested that Trp225, which is located in the active site, facilitates proper substrate entrance into the binding pocket
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
-
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
-
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
ternary complex mechanism in which the aromatic substrate has strict control in both the reductive and oxidative half-reaction in a way that reactions leading to substrate hydroxylation are favored over those leading to the futile formation of hydrogen peroxide. NAD+ release from the reduced enzyme-substrate complex is the slowest step in catalysis
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
the activity of the mutant enzyme HbpAind is six times lower than that of the wild-type enzyme
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
first enzyme of 2-hydroxybiphenyl degradation
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
first enzyme of the 2-hydroxybiphenyl degradation pathway
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
-
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
the activity of the mutant enzyme HbpAind is six times lower than that of the wild-type enzyme
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
ternary complex mechanism in which the aromatic substrate has strict control in both the reductive and oxidative half-reaction in a way that reactions leading to substrate hydroxylation are favored over those leading to the futile formation of hydrogen peroxide. NAD+ release from the reduced enzyme-substrate complex is the slowest step in catalysis
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
-
-
?
2-hydroxybiphenyl + NADH + O2
2,3-dihydroxybiphenyl + NAD+ + H2O
-
first enzyme of 2-hydroxybiphenyl degradation
-
-
?
2-hydroxybiphenyl + NADPH + O2
2,3-dihydroxybiphenyl + NADP+ + H2O
-
-
-
-
?
2-hydroxybiphenyl + NADPH + O2
2,3-dihydroxybiphenyl + NADP+ + H2O
-
-
-
-
?
2-hydroxybiphenyl + O2 + NADH + H+
3-phenylcatechol + NAD+ + H2O
-
-
-
-
?
2-hydroxybiphenyl + O2 + NADH + H+
3-phenylcatechol + NAD+ + H2O
-
-
-
-
?
2-propylphenol + NADH + O2
1,2-dihydroxy-3-propylbenzene + NAD+ + H2O
-
-
-
-
?
2-propylphenol + NADH + O2
1,2-dihydroxy-3-propylbenzene + NAD+ + H2O
-
-
-
-
?
2-sec-butylphenol + NADH + O2
2-sec-butylcatechol + NAD+ + H2O
-
-
-
-
?
2-sec-butylphenol + NADH + O2
2-sec-butylcatechol + NAD+ + H2O
-
-
-
-
?
2-tert-butylphenol + NADH + O2
1,2-dihydroxy-3-tert-butylbenzene + NAD+ + H2O
-
-
-
-
?
2-tert-butylphenol + NADH + O2
1,2-dihydroxy-3-tert-butylbenzene + NAD+ + H2O
-
-
-
-
?
guaiacol + NADH + O2
2,3-dihydroxy-methoxybenzene + NAD+ + H2O
-
i.e. 2-methoxyphenol
-
-
?
guaiacol + NADH + O2
2,3-dihydroxy-methoxybenzene + NAD+ + H2O
-
i.e. 2-methoxyphenol
-
-
?
indole + NADH + O2
?
-
-
-
-
?
indole + NADH + O2
?
-
-
-
-
?
additional information
?
-
-
Ile244 is located in the substrate binding pocket and is involved in accomodating the phenyl substituent of the phenol
-
-
?
additional information
?
-
-
Asp222 is involved in substrate activation in HbpA
-
-
?
additional information
?
-
-
the substrates partially uncouple oxygen activation from hydroxylation with resultant reduction of both atoms of oxygen to form hydrogen peroxide
-
-
?
additional information
?
-
comparisons of substrate binding structures, overview
-
-
-
additional information
?
-
-
comparisons of substrate binding structures, overview
-
-
-
additional information
?
-
wild-type enzyme HbpA has a broad substrate range and catalyzes the regioselective ortho-hydroxylation of a wide range of 2-substituted phenols to the corresponding catechols. It possess pro-S enantioselectivity towards the production of several chiral sulfoxides, whereas its mutant variant M321F exhibits improved enantioselectivity, while mutant M321A shows altered regioselectivity by oxidizing 3-hydroxybiphenyl, and thus enabling the production of a distinct antioxidant, 3,4-dihydroxybiphenyl. Identification of substrates and products by GC/MS
-
-
-
additional information
?
-
-
Ile244 is located in the substrate binding pocket and is involved in accomodating the phenyl substituent of the phenol
-
-
?
additional information
?
-
-
Asp222 is involved in substrate activation in HbpA
-
-
?
additional information
?
-
-
the substrates partially uncouple oxygen activation from hydroxylation with resultant reduction of both atoms of oxygen to form hydrogen peroxide
-
-
?
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
0.0034 - 0.004
2,2'-Dihydroxybiphenyl
0.0018 - 0.0787
2-Hydroxybiphenyl
0.0057
2-sec-Butylphenol
-
reaction with NADH or NADPH and O2
0.0292
O2
-
reaction with 2-hydroxybiphenyl and NADH
additional information
additional information
-
Km-values for wild-type and mutant enzymes
-
0.0034
2,2'-Dihydroxybiphenyl
-
reaction with NADPH and O2
0.004
2,2'-Dihydroxybiphenyl
-
reaction with NADH and O2
0.0018
2-Hydroxybiphenyl
mutant M321F, pH 7.5, 30°C
0.0019
2-Hydroxybiphenyl
-
-
0.0028
2-Hydroxybiphenyl
-
reaction with NADH and O2
0.0028
2-Hydroxybiphenyl
mutant M223E, pH 7.5, 30°C
0.0031
2-Hydroxybiphenyl
-
reaction with NADPH and O2
0.0031
2-Hydroxybiphenyl
pH 7.5, 30°C, wild-type enzyme
0.0031
2-Hydroxybiphenyl
recombinant wild-type enzyme, pH 7.5, 30°C
0.0031
2-Hydroxybiphenyl
pH 7.5, temperature not specified in the publication, mutant enzyme H48A
0.0033
2-Hydroxybiphenyl
pH 7.5, temperature not specified in the publication, wild-type enzyme
0.0048
2-Hydroxybiphenyl
pH 7.5, temperature not specified in the publication R242A
0.005
2-Hydroxybiphenyl
pH 7.5, temperature not specified in the publication, mutant enzyme D117A
0.006
2-Hydroxybiphenyl
mutant W97A, pH 7.5, 30°C
0.0065
2-Hydroxybiphenyl
mutant M223K, pH 7.5, 30°C
0.0123
2-Hydroxybiphenyl
mutant M223A, pH 7.5, 30°C
0.0153
2-Hydroxybiphenyl
mutant M321V, pH 7.5, 30°C
0.0156
2-Hydroxybiphenyl
mutant M321L, pH 7.5, 30°C
0.0169
2-Hydroxybiphenyl
mutant W97Y, pH 7.5, 30°C
0.0228
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme W225Y
0.0236
2-Hydroxybiphenyl
mutant M223I, pH 7.5, 30°C
0.0266
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme G255F
0.0278
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme R242E
0.0295
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme W225A
0.0462
2-Hydroxybiphenyl
mutant M321A, pH 7.5, 30°C
0.0787
2-Hydroxybiphenyl
mutant M223Q, pH 7.5, 30°C
0.0097
NADH
-
reaction with 2-hydroxybiphenyl
0.0216
NADH
-
reaction with 2,2'-dihydroxybiphenyl and O2
0.0268
NADH
-
reaction with 2-hydroxybiphenyl and O2
0.102
NADH
pH 7.5, 30°C, mutant enzyme W225Y
0.149
NADH
pH 7.5, 30°C, wild-type enzyme
0.222
NADH
pH 7.5, 30°C, mutant enzyme R242Q
0.253
NADH
pH 7.5, 30°C, mutant enzyme W225A
0.336
NADH
pH 7.5, 30°C, mutant enzyme R242A
0.427
NADH
pH 7.5, 30°C, mutant enzyme R242E
0.531
NADH
pH 7.5, 30°C, mutant enzyme G255F
0.0943
NADPH
-
reaction with 2,2'-dihydroxybiphenyl and O2
0.137
NADPH
-
reaction with 2-hydroxybiphenyl and O2
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
9 - 9.4
2,2'-Dihydroxybiphenyl
0.15 - 15.6
2-Hydroxybiphenyl
10.2 - 15.8
2-sec-Butylphenol
16.2
O2
-
reaction with 2-hydroxybiphenyl and NADH
additional information
additional information
-
turnover-numbers for mutant enzymes
-
9
2,2'-Dihydroxybiphenyl
-
reaction with NADH and O2
9.4
2,2'-Dihydroxybiphenyl
-
reaction with NADPH and O2
0.15
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme R242E
0.46
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme W225A
0.46
2-Hydroxybiphenyl
mutant W97Y, pH 7.5, 30°C
0.48
2-Hydroxybiphenyl
mutant W97A, pH 7.5, 30°C
0.71
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme G255F
0.73
2-Hydroxybiphenyl
mutant M223E, pH 7.5, 30°C
1.24
2-Hydroxybiphenyl
mutant M223A, pH 7.5, 30°C
1.4
2-Hydroxybiphenyl
-
turnover rate refers to the enzyme monomer and not to the tetramer
1.83
2-Hydroxybiphenyl
mutant M223I, pH 7.5, 30°C
1.97
2-Hydroxybiphenyl
mutant M223K, pH 7.5, 30°C
2.26
2-Hydroxybiphenyl
pH 7.5, 30°C, wild-type enzyme
2.26
2-Hydroxybiphenyl
recombinant wild-type enzyme, pH 7.5, 30°C
2.3
2-Hydroxybiphenyl
-
mutant enzyme HbpAind
2.75
2-Hydroxybiphenyl
mutant M321V, pH 7.5, 30°C
2.79
2-Hydroxybiphenyl
mutant M321L, pH 7.5, 30°C
3.01
2-Hydroxybiphenyl
mutant M321A, pH 7.5, 30°C
3.94
2-Hydroxybiphenyl
mutant M321F, pH 7.5, 30°C
4.51
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme W225Y
4.54
2-Hydroxybiphenyl
mutant M223Q, pH 7.5, 30°C
11.9
2-Hydroxybiphenyl
-
wild-type enzyme
12.8
2-Hydroxybiphenyl
-
reaction with NADPH and O2
14.9
2-Hydroxybiphenyl
-
reaction with NADH and O2
15.6
2-Hydroxybiphenyl
-
wild-type enzyme
10.2
2-sec-Butylphenol
-
reaction with NADPH and O2
14.5
2-sec-Butylphenol
-
wild-type enzyme
15.8
2-sec-Butylphenol
-
reaction with NADH and O2
0.005
indole
-
wild-type enzyme
0.09
indole
-
mutant enzyme HbpAind
0.14
NADH
pH 7.5, 30°C, mutant enzyme R242A
0.15
NADH
pH 7.5, 30°C, mutant enzyme R242E
0.24
NADH
pH 7.5, 30°C, mutant enzyme R242Q
1.34
NADH
pH 7.5, 30°C, mutant enzyme G255F
3.16
NADH
pH 7.5, 30°C, wild-type enzyme
3.45
NADH
pH 7.5, 30°C, mutant enzyme W225A
5.8
NADH
pH 7.5, 30°C, mutant enzyme W225Y
9.8
NADH
-
reaction with 2,2'-dihydroxybiphenyl and O2
16.2
NADH
-
reaction with 2-hydroxybiphenyl and O2
11.2
NADPH
-
reaction with 2,2'-dihydroxybiphenyl and O2
18.8
NADPH
-
reaction with 2-hydroxybiphenyl and O2
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
5.4 - 2188.9
2-Hydroxybiphenyl
5.4
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme R242A
20
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme W225A
27.2
2-Hydroxybiphenyl
mutant W97Y, pH 7.5, 30°C
30
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme G255F
57.7
2-Hydroxybiphenyl
mutant M223Q, pH 7.5, 30°C
65.2
2-Hydroxybiphenyl
mutant M321A, pH 7.5, 30°C
77.5
2-Hydroxybiphenyl
mutant M223I, pH 7.5, 30°C
80
2-Hydroxybiphenyl
mutant W97A, pH 7.5, 30°C
100.8
2-Hydroxybiphenyl
mutant M223A, pH 7.5, 30°C
178.9
2-Hydroxybiphenyl
mutant M321L, pH 7.5, 30°C
179.7
2-Hydroxybiphenyl
mutant M321V, pH 7.5, 30°C
200
2-Hydroxybiphenyl
pH 7.5, 30°C, mutant enzyme W225Y
260.7
2-Hydroxybiphenyl
mutant M223E, pH 7.5, 30°C
303.1
2-Hydroxybiphenyl
mutant M223K, pH 7.5, 30°C
729
2-Hydroxybiphenyl
recombinant wild-type enzyme, pH 7.5, 30°C
730
2-Hydroxybiphenyl
pH 7.5, 30°C, wild-type enzyme
2188.9
2-Hydroxybiphenyl
mutant M321F, pH 7.5, 30°C
0.314
NADH
pH 7.5, 30°C, mutant enzyme R242A
0.351
NADH
pH 7.5, 30°C, mutant enzyme R242E
1.08
NADH
pH 7.5, 30°C, mutant enzyme R242Q
2.53
NADH
pH 7.5, 30°C, mutant enzyme G255F
14
NADH
pH 7.5, 30°C, mutant enzyme W225A
20
NADH
pH 7.5, 30°C, wild-type enzyme
57
NADH
pH 7.5, 30°C, mutant enzyme W225Y
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
hanging drop method, structure of the enzyme with bound 2-hydroxybiphenyl, as well as several variants, at a resolution of 2.3-2.5 A to investigate structure function correlations of the enzyme. An observed hydrogen bond between 2-hydroxybiphenyl and His48 in the active site confirms the role of this residue in substrate deprotonation. The entrance to the active site is confirmed by generating variant G255F which exhibits only 7% of the wild-type's specific activity of product formation, suggesting inhibition of substrate entrance into the active site by the large aromatic residue. Residue Arg242 is suggested to facilitate FAD movement and reduction as was previously reported in studies on the homologous protein para-hydroxybenzoate hydroxylase. In addition, it is suggested that Trp225,which is located in the active site, facilitates proper substrate entrance into the binding pocket in contrast to aklavinone-11-hydroxylase and para-hydroxybenzoate hydroxylase in which a residue at a similar position is responsible for substrate deprotonation. Structure function correlations described in this work will aid in the design of variants with improved activity and altered selectivity for potential industrial applications
hanging-drop vapour-diffusion, optimized precipitant solution contains 1.6 M ammonium sulfate, 100 mM sodium chloride and 100 mM MES-NaOH, pH 7.5, crystals of native and SeMet-HbpA diffract to 2.01 and 2.25 A, respectively
-
purified recombinant enzyme HbpA mutant M321A, X-ray diffraction structure determination and anylysis at 2.78 A resolution, molecular replacement and structure modeling
single crystals of the enzyme are used to determine its structure in two forms: an FAD-bound form that allows characterisation of the active site, and an apo form that, although lacking flavin, gives extra information on the location of residues and structure of mobile loops that are absent from the FAD complex
structure in complex with FAD. Structural comparison of p-hydroxybenzoate hydroxylase (PobA) from Pseudomonas putida, EC 1.14.13.2, 2-hydroxybiphenyl 3-monooxygenase (HbpA) from Pseudomonas nitroreducens, EC 1.14.13.44, and 2-methyl-3-hydroxypyridine-5-carboxylic acid oxygenase (MHPCO) from Mesorhizobium japonicum. The portions of the residues required for substrate and FAD binding are identical, including the betaalphabeta fold and beta-sheet wall
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
G255F
the variant exhibits 7% compared to the specific activity of the wild-type enzyme on NADH and 2,3-dihydroxybiphenyl, suggesting inhibition of substrate entrance into the active site by the large aromatic residue
I244V
-
mutant enzyme has a 30% higher specific activity with 2-sec-butylphenol, guaiacol, and 2-hydroxybiphenyl. The Km-value for guaiacol decreases with this mutant, but the Km-value for 2-hydroxybiphenyl increase
M223A
site-directed saturation mutagenesis, the mutant shows reduced activity compared to wild-type enzyme
M223E
site-directed saturation mutagenesis, the mutant shows reduced activity compared to wild-type enzyme
M223I
site-directed saturation mutagenesis, the mutant shows reduced activity compared to wild-type enzyme
M223K
site-directed saturation mutagenesis, the mutant shows reduced activity compared to wild-type enzyme
M223Q
site-directed saturation mutagenesis, the mutant shows reduced activity compared to wild-type enzyme
M321A
site-directed saturation mutagenesis, the mutant variant demonstrates altered regioselectivity by oxidizing 3-hydroxybiphenyl, and thus enabling the production of a distinct antioxidant, 3,4-dihydroxybiphenyl, with similar ferric reducing capacity to the well-studied piceatannol. Mutant enzyme crystal structure analysis and comparison to the wild-type enzyme structure
M321F
site-directed saturation mutagenesis, wild-type HbpA possess pro-S enantioselectivity towards the production of several chiral sulfoxides, whereas the mutant M321F exhibits improved enantioselectivity and increased activity compared to wild-type
M321L
site-directed saturation mutagenesis, the mutant shows reduced activity compared to wild-type enzyme
M321V
site-directed saturation mutagenesis, the mutant shows reduced activity compared to wild-type enzyme
P320X
site-directed mutagenesis, all mutations of Pro320 in the library result in activity loss. Therefore, it appears that a proline at this position is crucial for the catalytic reaction. The short distance of Pro320 from the FAD cofactor implies its involvement in enabling the movement of FAD, which is imperative for HbpA activity. Pro320 is conserved among flavin monooxygenases. In aklavinone-11-hydroxylase (RdmE), para-hydroxybenzoate hydroxylase (pHBH), and phenol hydroxylase (PH), this position next to the FAD is occupied by Pro315, Pro293, and Pro364, respectively
R242A
the variant is not active on NADH and 2,3-dihydroxybiphenyl
R242E
the variant is not active on NADH and 2,3-dihydroxybiphenyl
R242Q
the variant is not active on NADH and 2,3-dihydroxybiphenyl
V368A/L417F
-
double replacement improves the efficiency of substrate hydroxylation by reducing the uncoupled oxidation of NADH. With guaiacol as substrate, the Vmax is increased and the Km-value is decreased. With 2-tert-butylphenol as substrate the turnover number is increased more than 5fold
W225A
the variant exhibits 7% compared to the specific activity of the wild-type enzyme on NADH and 2,3-dihydroxybiphenyl
W225Y
the variant exhibits 122% compared to the specific activity of the wild-type enzyme on NADH and 2,3-dihydroxybiphenyl
W97A
site-directed saturation mutagenesis, the mutant shows reduced activity compared to wild-type enzyme
W97Y
site-directed saturation mutagenesis, the mutant shows reduced activity compared to wild-type enzyme
I244V
-
mutant enzyme has a 30% higher specific activity with 2-sec-butylphenol, guaiacol, and 2-hydroxybiphenyl. The Km-value for guaiacol decreases with this mutant, but the Km-value for 2-hydroxybiphenyl increase
-
V368A/L417F
-
double replacement improves the efficiency of substrate hydroxylation by reducing the uncoupled oxidation of NADH. With guaiacol as substrate, the Vmax is increased and the Km-value is decreased. With 2-tert-butylphenol as substrate the turnover number is increased more than 5fold
-
additional information
-
direct enzyme evolution of EC 1.14.13.44 results in an enzyme variant HbpAind, that hydroxylates indole and indole derivatives such as hydroxyindoles and 5-bromoindole. The wild-type protein does not catalyze theses reactions. HbpAind contains amino acid substitutions D222V and V368A. The activity for indole hydroxylation is increased 18fold in this variant
additional information
site-directed saturation mutagenesis of HbpA and library screening, altering 2-hydroxybiphenyl 3-monooxygenase regioselectivity by protein engineering for the production of a new antioxidant. Modulation of the enzyme activity and selectivity via mutation of several residues in the active site pocket
additional information
-
direct enzyme evolution of EC 1.14.13.44 results in an enzyme variant HbpAind, that hydroxylates indole and indole derivatives such as hydroxyindoles and 5-bromoindole. The wild-type protein does not catalyze theses reactions. HbpAind contains amino acid substitutions D222V and V368A. The activity for indole hydroxylation is increased 18fold in this variant
-
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
Meyer, A.; Schmid, A.; Held, M.; Westphal, A.H.; Rothlisberger, M.; Kohler, H.P.E.; Van Berkel, W.J.H.; Witholt, B.
Changing the substrate reactivity of 2-hydroxybiphenyl 3-monooxygenase from Pseudomonas azelaica HBP1 by directed evolution
J. Biol. Chem.
277
5575-5582
2002
Pseudomonas nitroreducens, Pseudomonas nitroreducens HBP1
brenda
Meyer, A.; Wursten, M.; Schmid, A.; Kohler, H.P.; Witholt, B.
Hydroxylation of indole by laboratory-evolved 2-hydroxybiphenyl 3-monooxygenase
J. Biol. Chem.
277
34161-34167
2002
Pseudomonas nitroreducens, Pseudomonas nitroreducens HBP1
brenda
Suske, W.A.; Van Berkel, W.J.H.; Kohler, H.P.E.
Catalytic mechanism of 2-hydroxybiphenyl 3-monooxygenase, a flavoprotein from Pseudomonas azelaica HBP1
J. Biol. Chem.
274
33355-33365
1999
Pseudomonas nitroreducens, Pseudomonas nitroreducens HBP1
brenda
Suske, W.A.; Held, M.; Schmid, A.; Fleischmann, T.; Wubbolts, M.G.; Kohler, H.P.E.
Purification and characterization of 2-hydroxybiphenyl 3-monooxygenase, a novel NADH-dependent, FAD-containing aromatic hydroxylase from Pseudomonas azelaica HBP1
J. Biol. Chem.
272
24257-24265
1997
Pseudomonas nitroreducens, Pseudomonas nitroreducens HBP1
brenda
Meyer, A.; Tanner, D.; Schmid, A.; Sargent, D.F.; Kohler, H.P.; Witholt, B.
Crystallization and preliminary X-ray analysis of native and selenomethionine 2-hydroxybiphenyl 3-monooxygenase
Acta Crystallogr. Sect. D
59
741-743
2003
Pseudomonas nitroreducens
brenda
Held, M.; Schmid, A.; Kohler, H.P.; Suske, W.; Witholt, B.; Wubbolts, M.
An integrated process for the production of toxic catechols from toxic phenols based on a designer biocatalyst
Biotechnol. Bioeng.
62
641-648
1999
Escherichia coli, Escherichia coli JM101
brenda
Kanteev, M.; Bregman-Cohen, A.; Deri, B.; Shahar, A.; Adir, N.; Fishman, A.
A crystal structure of 2-hydroxybiphenyl 3-monooxygenase with bound substrate provides insights into the enzymatic mechanism
Biochim. Biophys. Acta
1854
1906-1913
2015
Pseudomonas nitroreducens (O06647)
brenda
Jensen, C.N.; Mielke, T.; Farrugia, J.E.; Frank, A.; Man, H.; Hart, S.; Turkenburg, J.P.; Grogan, G.
Structures of the Apo and FAD-bound forms of 2-hydroxybiphenyl 3-monooxygenase (HbpA) locate activity hotspots identified by using directed evolution
ChemBioChem
16
968-976
2015
Pseudomonas nitroreducens (O06647)
brenda
Lazar, J.T.; Shuvalova, L.; Rosas-Lemus, M.; Kiryukhina, O.; Satchell, K.J.F.; Minasov, G.
Structural comparison of p-hydroxybenzoate hydroxylase (PobA) from Pseudomonas putida with PobA from other Pseudomonas spp. and other monooxygenases
Acta Crystallogr. F Struct. Biol. Commun.
75
507-514
2019
Pseudomonas nitroreducens (O06647), Pseudomonas nitroreducens
brenda
Lazar, J.T.; Shuvalova, L.; Rosas-Lemus, M.; Kiryukhina, O.; Satchell, K.J.F.; Minasov, G.
Structural comparison of p-hydroxybenzoate hydroxylase (PobA) from Pseudomonas putida with PobA from other Pseudomonas spp. and other monooxygenases
Acta Crystallogr. Sect. F
75
507-514
2019
Pseudomonas nitroreducens (O06647), Pseudomonas nitroreducens
brenda
Bregman-Cohen, A.; Deri, B.; Maimon, S.; Pazy, Y.; Fishman, A.
Altering 2-hydroxybiphenyl 3-monooxygenase regioselectivity by protein engineering for the production of a new antioxidant
ChemBioChem
19
583-590
2018
Pseudomonas nitroreducens (O06647)
brenda