Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary for 1.14.13.24 extracted from

  • Chen, X.; Tang, H.; Liu, Y.; Xu, P.; Xue, Y.; Lin, K.; Cui, C.
    Purification and initial characterization of 3-hydroxybenzoate 6-hydroxylase from a halophilic Martelella strain AD-3 (2018), Front. Microbiol., 9, 1335 .
    View publication on PubMedView publication on EuropePMC
Search for more literature for 1.14.13.24

Cloned(Commentary)

Cloned (Comment) Organism
gene 3HB6H is encoded in a cluster that potentially encodes for gentisic acid degradation, sequence comparisons and phylogenetic analysis and tree recombinant expression of N-terminally His-tagged enzyme in Escherichia coli strain BL21(DE3) Martelella sp. AD-3

Protein Variants

Protein Variants Comment Organism
A308G site-directed mutagenesis, the mutant shows reduced activity compared to wild-type enzyme, comparison of FAD contents Martelella sp. AD-3
Q305P site-directed mutagenesis, the mutant shows reduced activity compared to wild-type enzyme, comparison of FAD contents Martelella sp. AD-3
Y221F site-directed mutagenesis, the mutant shows reduced activity compared to wild-type enzyme, comparison of FAD contents Martelella sp. AD-3
Y306H site-directed mutagenesis, the mutant shows reduced activity compared to wild-type enzyme, comparison of FAD contents Martelella sp. AD-3

Inhibitors

Inhibitors Comment Organism Structure
Co2+ inhibits the enzyme activity at 0.5 mM by about 30% Martelella sp. AD-3
Cu2+ inhibits the enzyme activity at 0.5 mM by 83.2% Martelella sp. AD-3
Fe3+ inhibits the enzyme activity at 0.5 mM by about 50% Martelella sp. AD-3
Mg2+ inhibits the enzyme activity at 0.5 mM by about 50% Martelella sp. AD-3
Zn2+ inhibits the enzyme activity at 0.5 mM by 77.4% Martelella sp. AD-3

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information purified 3HB6H displays typical Michaelis-Menten kinetics, Lineweaver-Burk plots Martelella sp. AD-3
0.0337
-
 NADH pH 8.0, 30°C, mutant A308G Martelella sp. AD-3
0.038
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant A308G Martelella sp. AD-3
0.0463
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant Y306H Martelella sp. AD-3
0.0726
-
 3-hydroxybenzoate pH 8.0, 30°C, recombinant wild-type enzyme Martelella sp. AD-3
0.0867
-
 NADH pH 8.0, 30°C, mutant Y221F Martelella sp. AD-3
0.0913
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant Q305P Martelella sp. AD-3
0.0959
-
 NADH pH 8.0, 30°C, mutant Q305P Martelella sp. AD-3
0.1037
-
 NADH pH 8.0, 30°C, mutant Y306H Martelella sp. AD-3
0.1041
-
 NADH pH 8.0, 30°C, recombinant enzyme Martelella sp. AD-3
0.1642
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant Y221F Martelella sp. AD-3

Metals/Ions

Metals/Ions Comment Organism Structure
additional information enzyme activity is highest without salinity and metal salts. Ca2+ and Mn2+ have a poor effect on enzyme activity at 0.5 mM. With salinity ranging from 0 to 85.56 mM, the increase in salinity clearly causes decreased enzyme activity, 3% activity remaining at 85.56 mM salinity Martelella sp. AD-3

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
3-hydroxybenzoate + NADH + H+ + O2 Martelella sp. AD-3
-
 2,5-dihydroxybenzoate + NAD+ + H2O
-
 ?

Organism

Organism UniProt Comment Textmining
Martelella sp. AD-3
-
-
-

Posttranslational Modification

Posttranslational Modification Comment Organism
lipoprotein the recombinant 3HB6H expressed in Escherichia coil contains phosphatidylglycerol, while phosphatidylethanolamine is not detected Martelella sp. AD-3

Purification (Commentary)

Purification (Comment) Organism
recombinant N-terminally His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography Martelella sp. AD-3

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
25.2
-
 purified recombinant enzyme, pH 8.0, 37°C Martelella sp. AD-3

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3-hydroxybenzoate + NADH + H+ + O2
-
 Martelella sp. AD-3 2,5-dihydroxybenzoate + NAD+ + H2O
-
 ?

Subunits

Subunits Comment Organism
homodimer 2 * 46000, recombinant His-tagged enzyme SDS-PAGE Martelella sp. AD-3

Synonyms

Synonyms Comment Organism
3-hydroxybenzoate 6-hydroxylase
-
 Martelella sp. AD-3
3HB6H
-
 Martelella sp. AD-3

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 recombinant enzyme Martelella sp. AD-3

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
16 50 enzyme activity is maintained at 16-37°C, and quickly lost at higher temperatures. The enzyme precipitates initially at 42°C and precipitates completely at 50°C Martelella sp. AD-3

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1
-
 NADH pH 8.0, 30°C, mutant Y221F Martelella sp. AD-3
1.5
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant Y306H Martelella sp. AD-3
1.6
-
 NADH pH 8.0, 30°C, mutant Q305P Martelella sp. AD-3
1.6
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant Q305P Martelella sp. AD-3
1.6
-
 NADH pH 8.0, 30°C, mutant Y306H Martelella sp. AD-3
1.7
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant Y221F Martelella sp. AD-3
2
-
 NADH pH 8.0, 30°C, mutant A308G Martelella sp. AD-3
2.3
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant A308G Martelella sp. AD-3
5.7
-
 3-hydroxybenzoate pH 8.0, 30°C, recombinant wild-type enzyme Martelella sp. AD-3
7.8
-
 NADH pH 8.0, 30°C, recombinant enzyme Martelella sp. AD-3

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8
-
 recombinant enzyme, phosphate buffer Martelella sp. AD-3

Cofactor

Cofactor Comment Organism Structure
FAD residues 305, 306 and 308 are important for FAD binding. FAD concentration in the purified 3HB6H (0.29 mM) is 0.30 mM, and FAD/enzyme ratio is 1.02/0.05 determined by HPLC analysis Martelella sp. AD-3
NADH
-
 Martelella sp. AD-3

General Information

General Information Comment Organism
evolution sequence comparisons and phylogenetic analysis, phylogenetic tree is constructed with 3-hydroxybenzoate 6-hydroxylases, salicylate 1-hydroxylases and 3-hydroxybenzoate 4-hydroxylases from other 17 strains and confirmed that it is most closely related to 3HB6H from Pseudomonas alcaligenes strain NCIMB 9867 Martelella sp. AD-3
metabolism 3-hydroxybenzoate 6-hydroxylase can convert 3-hydroxybenzoate which is an important intermediate in the biodegradation of many aromatic hydrocarbons. 3-Hydroxybenzoate is metabolized by entering the TCA cycle through the gentisate pathway Martelella sp. AD-3
additional information residues Tyr221 and Gln305 of 3HB6H from Martelella sp. strain AD-3 are involved in substrate binding Martelella sp. AD-3
physiological function 3-hydroxybenzoate 6-hydroxylase, an NADH-dependent flavoprotein, can convert 3-hydroxybenzoate which is an important intermediate in the biodegradation of many aromatic hydrocarbons Martelella sp. AD-3

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
10.35
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant Y221F Martelella sp. AD-3
11.53
-
 NADH pH 8.0, 30°C, mutant Y221F Martelella sp. AD-3
15.43
-
 NADH pH 8.0, 30°C, mutant Y306H Martelella sp. AD-3
16.68
-
 NADH pH 8.0, 30°C, mutant Q305P Martelella sp. AD-3
32.4
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant Y306H Martelella sp. AD-3
59.35
-
 NADH pH 8.0, 30°C, mutant A308G Martelella sp. AD-3
60.53
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant A308G Martelella sp. AD-3
62.61
-
 3-hydroxybenzoate pH 8.0, 30°C, mutant Q305P Martelella sp. AD-3
74.93
-
 NADH pH 8.0, 30°C, recombinant enzyme Martelella sp. AD-3
78.51
-
 3-hydroxybenzoate pH 8.0, 30°C, recombinant wild-type enzyme Martelella sp. AD-3