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Literature summary extracted from

  • Zhang, X.; Rao, Z.; Zhang, L.; Xu, M.; Yang, T.
    Efficient 9alpha-hydroxy-4-androstene-3,17-dione production by engineered Bacillus subtilis co-expressing Mycobacterium neoaurum 3-ketosteroid 9alpha-hydroxylase and B. subtilis glucose 1-dehydrogenase with NADH regeneration (2016), SpringerPlus, 5, 1207 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.14.15.30 expressed in Bacillus subtilis strain 168 Mycolicibacterium neoaurum
1.14.15.30 genes kshA and kshB, DNA and amino acid sequence determination and analysis, cloning into plasmid pMA5, recombinant expression of C-terminally His-tagged KshA and KshB in Bacillus subtilis strain 168, coexpression with Bacillus subtilis glucose 1-dehydrogenase, method development and evaluation, overview. KSH activity can only be detected when KshA and KshB are coexpressed Mycolicibacterium neoaurum

Protein Variants

EC Number Protein Variants Comment Organism
1.14.15.30 additional information efficient 9alpha-hydroxy-4-androstene-3,17-dione production by engineered Bacillus subtilis co-expressing Mycobacterium neoaurum 3-ketosteroid 9alpha-hydroxylase and Bacillus subtilis glucose 1-dehydrogenase with NADH regeneration Mycolicibacterium neoaurum

Inhibitors

EC Number Inhibitors Comment Organism Structure
1.14.15.30 Co2+
-
 Mycolicibacterium neoaurum
1.14.15.30 Cu2+
-
 Mycolicibacterium neoaurum
1.14.15.30 Fe3+
-
 Mycolicibacterium neoaurum
1.14.15.30 Ni2+
-
 Mycolicibacterium neoaurum
1.14.15.30 Zn2+
-
 Mycolicibacterium neoaurum

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.14.15.30 1,4-androstadiene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2 Mycolicibacterium neoaurum
-
 9alpha-hydroxy-1,4-androstadiene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 1,4-androstadiene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2 Mycolicibacterium neoaurum JC-12
-
 9alpha-hydroxy-1,4-androstadiene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2 Mycolicibacterium neoaurum
-
 9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2 Mycolicibacterium neoaurum JC-12
-
 9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + H+ + O2 Mycolicibacterium neoaurum
-
 9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + H+ + O2 Mycolicibacterium neoaurum JC-12
-
 9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 additional information Mycolicibacterium neoaurum Mycobacterium neoaurum strain JC-12 is a 4-androstene-3,17-dione and 1,4-androstadiene-3,17-dione producing strain ?
-
 ?
1.14.15.30 additional information Mycolicibacterium neoaurum JC-12 Mycobacterium neoaurum strain JC-12 is a 4-androstene-3,17-dione and 1,4-androstadiene-3,17-dione producing strain ?
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.14.15.30 Mycolicibacterium neoaurum A0A0F7JI94 AND D2Y640 genes kshA and kshB encoding the enzyme subunits KshA and KshB
-
1.14.15.30 Mycolicibacterium neoaurum A0A0F7JI94 AND D2Y640 subunit KshA and subunit KshB
-
1.14.15.30 Mycolicibacterium neoaurum JC-12 A0A0F7JI94 AND D2Y640 genes kshA and kshB encoding the enzyme subunits KshA and KshB
-
1.14.15.30 Mycolicibacterium neoaurum JC-12 A0A0F7JI94 AND D2Y640 subunit KshA and subunit KshB
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.14.15.30 recombinant His-tagged KshA and KshB from Bacillus subtilis strain 168 by nickel affinity chromatography Mycolicibacterium neoaurum

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.14.15.30 0.57
-
 recombinant enzyme from Bacillus subtilis, pH 7.0, 22°C Mycolicibacterium neoaurum

Storage Stability

EC Number Storage Stability Organism
1.14.15.30 -20°C and 0°C, 50 mM Tris-HCl buffer (pH 7.0), 24 h, 28% and 67% loss of activity, respectively Mycolicibacterium neoaurum

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.14.15.30 1,4-androstadiene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
-
 Mycolicibacterium neoaurum 9alpha-hydroxy-1,4-androstadiene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 1,4-androstadiene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
-
 Mycolicibacterium neoaurum JC-12 9alpha-hydroxy-1,4-androstadiene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
-
 Mycolicibacterium neoaurum 9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
-
 Mycolicibacterium neoaurum JC-12 9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + H+ + O2
-
 Mycolicibacterium neoaurum 9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + H+ + O2
-
 Mycolicibacterium neoaurum JC-12 9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.30 additional information Mycobacterium neoaurum strain JC-12 is a 4-androstene-3,17-dione and 1,4-androstadiene-3,17-dione producing strain Mycolicibacterium neoaurum ?
-
 ?
1.14.15.30 additional information KshB, the reductase component of KSH, accepts electrons from NADH and transfers it to oxygenase component KshA, the cooperation of KshA and KshB is critically important for maintaining KSH activity Mycolicibacterium neoaurum ?
-
 ?
1.14.15.30 additional information Mycobacterium neoaurum strain JC-12 is a 4-androstene-3,17-dione and 1,4-androstadiene-3,17-dione producing strain Mycolicibacterium neoaurum JC-12 ?
-
 ?
1.14.15.30 additional information KshB, the reductase component of KSH, accepts electrons from NADH and transfers it to oxygenase component KshA, the cooperation of KshA and KshB is critically important for maintaining KSH activity Mycolicibacterium neoaurum JC-12 ?
-
 ?

Subunits

EC Number Subunits Comment Organism
1.14.15.30 ? x * 378000, subunit KshB, SDS-PAGE Mycolicibacterium neoaurum
1.14.15.30 ? x * 451000, subunit KshA, SDS-PAGE Mycolicibacterium neoaurum
1.14.15.30 More enzyme KSH consists of KshA and KshB components Mycolicibacterium neoaurum

Synonyms

EC Number Synonyms Comment Organism
1.14.15.30 3-ketosteroid 9alpha-hydroxylase
-
 Mycolicibacterium neoaurum
1.14.15.30 KSH
-
 Mycolicibacterium neoaurum
1.14.15.30 KshA reductase subunit Mycolicibacterium neoaurum
1.14.15.30 KshB oxygenase subunit Mycolicibacterium neoaurum

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.14.15.30 33
-
-
 Mycolicibacterium neoaurum
1.14.15.30 33
-
 recombinant enzyme Mycolicibacterium neoaurum

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.14.15.30 7
-
-
 Mycolicibacterium neoaurum
1.14.15.30 7
-
 assay at Mycolicibacterium neoaurum

General Information

EC Number General Information Comment Organism
1.14.15.30 metabolism 3-ketosteroid 9alpha-hydroxylase is a key enzyme in steroid metabolism and can catalyze the transformation of 4-androstene-3,17-dione to 9alpha-hydroxy-4-androstene-3,17-dione Mycolicibacterium neoaurum