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Information on EC 1.14.15.30 - 3-ketosteroid 9alpha-monooxygenase and Organism(s) Mycobacterium tuberculosis and UniProt Accession P71875
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1.14.15.30 3-ketosteroid 9alpha-monooxygenaseIUBMB Comments
The enzyme is involved in the cholesterol degradation pathway of several bacterial pathogens, such as Mycobacterium tuberculosis. It forms a two-component system with a ferredoxin reductase (KshB). The enzyme contains a Rieske-type iron-sulfur center and non-heme iron. The product of the enzyme is unstable, and spontaneously converts to 3-hydroxy-9,10-seconandrost-1,3,5(10)-triene-9,17-dione.
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Word Map
- 1.14.15.30
- 3-ketosteroids
- cholesterol
-
rieske
- rhodochrous
- tuberculosis
- 4-androstene-3,17-dione
-
two-component
- oxygenases
-
cholic
- sterols
- erythropolis
- bile
-
mouth
- ruber
- coa
- smegmatis
- medicine
- drug development
The taxonomic range for the selected organisms is: Mycobacterium tuberculosis
The expected taxonomic range for this enzyme is: Bacteria, Archaea
The expected taxonomic range for this enzyme is: Bacteria, Archaea
Reaction Schemes
+
2
+
2
+
=
+
2
+
+
2
reduced ferredoxin [iron-sulfur] cluster
+
2
+
=
+
2
oxidized ferredoxin [iron-sulfur] cluster
+
Synonyms
ksha1, ksha2, kshab, ksha5, ksha3, 3-ketosteroid-9alpha-hydroxylase, 3-ketosteroid 9-alpha-hydroxylase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
3-ketosteroid 9alpha-hydroxylase
KSH
-
3-ketosteroid 9alpha-hydroxylase is comprised of KshA (oxygenase) and KshB (oxygenase-reductase)
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
androsta-1,4-diene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2 = 9alpha-hydroxyandrosta-1,4-diene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
androsta-1,4-diene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2 = 9alpha-hydroxyandrosta-1,4-diene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
KSH enzymes employ an electron transport chain that starts with the oxidation of NADH. The electrons are transferred to the flavin cofactor (FAD) of the ferredoxin reductase component KshB and then transported to the plant type iron-sulfur cluster of KshB. The Rieske iron-sulfur cluster of the KshA oxygenase component subsequently accepts the electrons from KshB. The electrons end up at the nonheme iron situated in the active site of KshA. The mononuclear iron is the site where O2 is bound and activated and the substrate is hydroxylated
androsta-1,4-diene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2 = 9alpha-hydroxyandrosta-1,4-diene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
KshAB-catalyzed 9alpha-hydroxylation of 3-ketosteroids. 1,4-Diene ketosteroids undergo subsequent nonenzymatic cleavage of ring B and aromatization of ring A
PATHWAY SOURCE
PATHWAYS
-
-
SYSTEMATIC NAME
IUBMB Comments
androsta-1,4-diene-3,17-dione,[reduced ferredoxin]:oxygen oxidoreductase (9alpha-hydroxylating)
The enzyme is involved in the cholesterol degradation pathway of several bacterial pathogens, such as Mycobacterium tuberculosis. It forms a two-component system with a ferredoxin reductase (KshB). The enzyme contains a Rieske-type iron-sulfur center and non-heme iron. The product of the enzyme is unstable, and spontaneously converts to 3-hydroxy-9,10-seconandrost-1,3,5(10)-triene-9,17-dione.
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
1,4-androstadiene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-1,4-androstadiene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
1,4-androstadiene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + O2
9alpha-hydroxy-1,4-androstadiene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
3-oxo-23,24-bisnorcholest-4-en-22-oic acid + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-3-oxo-23,24-bisnorcholest-4-en-22-oic acid + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
3-oxo-23,24-bisnorcholest-4-en-22-oyl-CoA + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-3-oxo-23,24-bisnorcholest-4-en-22-oyl-CoA + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate-CoA + reduced ferredoxin [iron-sulfur] cluster + H+ + O2
?
-
-
-
r
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oyl-CoA + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-3-oxo-23,24-bisnorcholesta-1,4-dien-22-oyl-CoA + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
4-androstene-3,17-dione + O2 + NADH + H+
9alpha-hydroxy-4-androstene-3,17-dione + H2O + NAD+
-
-
-
?
androsta-1,4-diene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxyandrosta-1,4-diene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
3-oxo-23,24-bisnorcholest-4-en-22-oic acid + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-3-oxo-23,24-bisnorcholest-4-en-22-oic acid + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
3-oxo-23,24-bisnorcholest-4-en-22-oyl-CoA + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-3-oxo-23,24-bisnorcholest-4-en-22-oyl-CoA + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate-CoA + reduced ferredoxin [iron-sulfur] cluster + H+ + O2
?
-
-
-
r
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oyl-CoA + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-3-oxo-23,24-bisnorcholesta-1,4-dien-22-oyl-CoA + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
4-androstene-3,17-dione + O2 + NADH + H+
? + H2O + NAD+
-
-
-
?
5alpha-androstane-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-5alpha-androstane-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
-
?
androsta-1,4-diene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxyandrosta-1,4-diene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
androsta-1,4-diene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxyandrosta-1,4-diene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
androsta-1,4-diene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxyandrosta-1,4-diene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
androsta-1,4-diene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxyandrosta-1,4-diene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
-
?
androsta-1,4-diene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxyandrosta-1,4-diene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
additional information
?
-
-
substrate specificity, overview. KshAB also transforms CoA thioester steroids
-
-
?
additional information
?
-
substrate specificity, overview. KshAB also transforms CoA thioester steroids
-
-
?
additional information
?
-
substrate specificity, overview. KshAB also transforms CoA thioester steroids
-
-
?
additional information
?
-
cholesterol-degrading KshAMtb from Mycobacterium tuberculosis has the highest catalytic efficiency for CoA-thioesterified substrates, substrate specificity comparisons of KSH enzymes, overview
-
-
?
additional information
?
-
-
cholesterol-degrading KshAMtb from Mycobacterium tuberculosis has the highest catalytic efficiency for CoA-thioesterified substrates, substrate specificity comparisons of KSH enzymes, overview
-
-
?
additional information
?
-
KSH of Mycobacterium tuberculosis can use 3-ketosteroids as substrates and shows high preference for the CoA thioester intermediate of cholesterol side chain degradation compared to the tested C17-ketosteroids
-
-
?
additional information
?
-
-
substrate specificity, overview. KshAB also transforms CoA thioester steroids
-
-
?
additional information
?
-
substrate specificity, overview. KshAB also transforms CoA thioester steroids
-
-
?
additional information
?
-
substrate specificity, overview. KshAB also transforms CoA thioester steroids
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
1,4-androstadiene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-1,4-androstadiene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
4-androstene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-4-androstene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
androsta-1,4-diene-3,17-dione + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxyandrosta-1,4-diene-3,17-dione + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
3-oxo-23,24-bisnorcholest-4-en-22-oyl-CoA + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-3-oxo-23,24-bisnorcholest-4-en-22-oyl-CoA + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oyl-CoA + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2
9alpha-hydroxy-3-oxo-23,24-bisnorcholesta-1,4-dien-22-oyl-CoA + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
FAD
flavin co-factor of the ferredoxin reductase component KshB
Fe-S center
Rieske-type Fe-S cluster, Rieske monooxygenase
[2Fe-2S]-center
8.2 mol of iron and 2.4 mol of sulfur per mol of KshA protomer
[2Fe-2S]-center
a Rieske iron-sulfur cluster of the KshA oxygenase component and a plant type iron-sulfur cluster of KshB. The oxygenase component, which performs the substrate hydroxylation, is an iron-sulfur protein and contains a non-heme iron situated at the active site. The Rieske iron-sulfur cluster and the non-heme Fe2+ catalytic centre are located relatively far away from each other, but the typical head-to-tail trimer arrangement positions the Rieske Fe2-S2 in close proximity to the non-heme Fe2+ of the neighbouring KshA subunit, enabling transport of electrons between KshA subunits
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Fe2+
Fe2+
the oxygenase component, which performs the substrate hydroxylation, is an iron-sulfur protein and contains a non-heme iron situated at the active site. The iron is bidentate bound to the carboxyl group of the aspartate leaving two sites available for exogenous ligands. This metal centre is more labile compared to the covalently bound heme-iron. Non-heme iron is a highly catalytic platform able to bind O2 and steroid substrate simultaneously in different orientations
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.11
1,4-androstadiene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 25°C
0.07
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate
pH 7.0, 22°C, KshAB
0.017
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate-CoA
pH 7.0, 22°C, KshAB
0.003
3-oxo-23,24-bisnorcholesta-4-ene-22-oate
pH 7.0, 22°C, KshAB
0.0068
3-oxo-23,24-bisnorcholesta-4-ene-22-oyl-CoA
pH 7.0, 22°C, KshAB
0.1 - 1
androsta-1,4-diene-3,17-dione
pH 7.0, 22°C, KshAB
1.2
O2
in the presence of 1,4-androstadiene-3,17-dione, Km above 1.2 mM, in 0.1 M potassium phosphate, pH 7.0, at 25°C
0.003
3-oxo-23,24-bisnorchol-4-en-22-oic acid
in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.0068
3-oxo-23,24-bisnorchol-4-en-22-oic acid-CoA
in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.07
3-oxo-23,24-bisnorchola-1,4-dien-22-oic acid
in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.017
3-oxo-23,24-bisnorchola-1,4-dien-22-oic acid-CoA
in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.07
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate
pH 7.0, 22°C, KshAB
0.017
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate-CoA
pH 7.0, 22°C, KshAB
0.003
3-oxo-23,24-bisnorcholesta-4-ene-22-oate
pH 7.0, 22°C, KshAB
0.0068
3-oxo-23,24-bisnorcholesta-4-ene-22-oyl-CoA
pH 7.0, 22°C, KshAB
0.09
O2
apparent Km value, in the presence of 3-oxo-23,24-bisnorchola-1,4-dien-22-oic acid-CoA, in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.024
4-androstene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 25°C
0.024
4-androstene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.1 - 1
androsta-1,4-diene-3,17-dione
pH 7.0, 22°C, KshAB
0.11
androsta-1,4-diene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 22°C
additional information
additional information
steady-state kinetic analysis
-
additional information
additional information
-
steady-state kinetic analysis
-
additional information
additional information
-
steady-state kinetic parameters for KshAB, overview
-
additional information
additional information
steady-state kinetic parameters for KshAB, overview
-
additional information
additional information
steady-state kinetic parameters for KshAB, overview
-
additional information
additional information
-
steady-state kinetic parameters for KshAB, overview
-
additional information
additional information
steady-state kinetic parameters for KshAB, overview
-
additional information
additional information
steady-state kinetic parameters for KshAB, overview
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.8
1,4-androstadiene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 25°C
0.25
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate
pH 7.0, 22°C, KshAB
2.7
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate-CoA
pH 7.0, 22°C, KshAB
0.08
3-oxo-23,24-bisnorcholesta-4-ene-22-oate
pH 7.0, 22°C, KshAB
0.61
3-oxo-23,24-bisnorcholesta-4-ene-22-oyl-CoA
pH 7.0, 22°C, KshAB
0.8
androsta-1,4-diene-3,17-dione
pH 7.0, 22°C, KshAB
7
O2
in the presence of 1,4-androstadiene-3,17-dione, kcat above 7 1/sec, in 0.1 M potassium phosphate, pH 7.0, at 25°C
0.08
3-oxo-23,24-bisnorchol-4-en-22-oic acid
in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.61
3-oxo-23,24-bisnorchol-4-en-22-oic acid-CoA
in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.25
3-oxo-23,24-bisnorchola-1,4-dien-22-oic acid
in 0.1 M potassium phosphate, pH 7.0, at 22°C
2.7
3-oxo-23,24-bisnorchola-1,4-dien-22-oic acid-CoA
in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.25
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate
pH 7.0, 22°C, KshAB
2.7
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate-CoA
pH 7.0, 22°C, KshAB
0.08
3-oxo-23,24-bisnorcholesta-4-ene-22-oate
pH 7.0, 22°C, KshAB
0.61
3-oxo-23,24-bisnorcholesta-4-ene-22-oyl-CoA
pH 7.0, 22°C, KshAB
2.5
O2
apparent Km value, in the presence of 3-oxo-23,24-bisnorchola-1,4-dien-22-oic acid-CoA, in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.07
4-androstene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 25°C
0.07
4-androstene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.8
androsta-1,4-diene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 22°C
0.8
androsta-1,4-diene-3,17-dione
pH 7.0, 22°C, KshAB
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
7.6
1,4-androstadiene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 25°C
3.57
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate
pH 7.0, 22°C, KshAB
158.8
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate-CoA
pH 7.0, 22°C, KshAB
26.67
3-oxo-23,24-bisnorcholesta-4-ene-22-oate
pH 7.0, 22°C, KshAB
89.71
3-oxo-23,24-bisnorcholesta-4-ene-22-oyl-CoA
pH 7.0, 22°C, KshAB
7.27
androsta-1,4-diene-3,17-dione
pH 7.0, 22°C, KshAB
2.45
O2
in the presence of 1,4-androstadiene-3,17-dione, in 0.1 M potassium phosphate, pH 7.0, at 25°C
30
3-oxo-23,24-bisnorchol-4-en-22-oic acid
in 0.1 M potassium phosphate, pH 7.0, at 22°C
90
3-oxo-23,24-bisnorchol-4-en-22-oic acid-CoA
in 0.1 M potassium phosphate, pH 7.0, at 22°C
3.5
3-oxo-23,24-bisnorchola-1,4-dien-22-oic acid
in 0.1 M potassium phosphate, pH 7.0, at 22°C
160
3-oxo-23,24-bisnorchola-1,4-dien-22-oic acid-CoA
in 0.1 M potassium phosphate, pH 7.0, at 22°C
3.57
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate
pH 7.0, 22°C, KshAB
158.8
3-oxo-23,24-bisnorcholesta-1,4-dien-22-oate-CoA
pH 7.0, 22°C, KshAB
26.67
3-oxo-23,24-bisnorcholesta-4-ene-22-oate
pH 7.0, 22°C, KshAB
89.71
3-oxo-23,24-bisnorcholesta-4-ene-22-oyl-CoA
pH 7.0, 22°C, KshAB
29
O2
apparent Km value, in the presence of 3-oxo-23,24-bisnorchola-1,4-dien-22-oic acid-CoA, in 0.1 M potassium phosphate, pH 7.0, at 22°C
3
4-androstene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 25°C
3
4-androstene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 22°C
7.27
androsta-1,4-diene-3,17-dione
pH 7.0, 22°C, KshAB
7.6
androsta-1,4-diene-3,17-dione
in 0.1 M potassium phosphate, pH 7.0, at 22°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
22
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
gene kshA; the enzyme harbors a Rieske oxygenase, encoded by kshA
UniProt
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
evolution
KSH is classified as a group I RO in this classification system. Group I contains a broad range of mono- and dioxygenases with low amino acid sequence similarity and various protein sizes. The oxygenases are alpha-monomers. KshA contains a Rieske domain, coordinating the Rieske Fe2-S2 cluster, and a catalytic domain with the typical helix-Grip fold, which is part of the StAR (steroidogenic acute regulatory protein) related lipid transfer (START) domain superfamily. The catalytic domain is composed of a beta-sheet flanked by alpha-helices
malfunction
deletion of KSH activity in sterol degrading bacteria results in blockage of steroid ring opening and is used to produce valuable C19-steroids such as 4-androstene-3,17-dione and 1,4-androstadiene-3,17-dione
physiological function
malfunction
-
DELTAkshA and DELTAkshB deletion mutants are unable to use cholesterol and 4-androstene-3,17-dione as primary carbon and energy sources and are unable to metabolize 5alpha-androstane-3,17-dione. The deletion of either of these genes leads to rapid death of the microorganism in murine infection models and in macrophages
physiological function
additional information
physiological function
the enzyme plays a role of cholesterol catabolism, overview
physiological function
a plant type iron-sulfur cluster of KshB. The 3-ketosteroid 9alpha-hydroxylase activity is a two component Rieske non-heme monooxygenase comprised of the oxygenase KshA and the reductase KshB, and is a key enzyme in bacterial steroid degradation, essential for the pathogenicity of Mycobacterium tuberculosis. KSH initiates opening of the steroid polycyclic ring structure
physiological function
KshA is the oxygenase component of 3-ketosteroid 9alpha-hydroxylase, a Rieske oxygenase involved in the bacterial degradation of steroids
physiological function
the enzyme is essential for the pathogenicity of Mycobacterium tuberculosis
physiological function
-
3-ketosteroid 9alpha-hydroxylase is an essential factor in the pathogenesis of Mycobacterium tuberculosis
physiological function
the enzyme plays a role of cholesterol catabolism, overview
additional information
-
enzyme substrate binding pocket analysis using molecular modeling, overview. Modeling of substrates into KshA crystal structure using KshA structure PDB 2ZYL
additional information
enzyme substrate binding pocket analysis using molecular modeling, overview. Modeling of substrates into KshA crystal structure using KshA structure PDB 2ZYL
additional information
enzyme substrate binding pocket analysis using molecular modeling, overview. Modeling of substrates into KshA crystal structure using KshA structure PDB 2ZYL
additional information
structure-function relationship of KSH enzymes and components, overview
additional information
the enzyme has a steroid-binding pocket with the substrate's C17 oriented toward the active site opening. Comparisons suggest that Tyr245 and Phe297 are determinants of KshA1 specificity. The enzyme has a flexible 16-residue mouth loop, which in some structures completely occlude the substrate-binding pocket from the bulk solvent
additional information
-
the enzyme has a steroid-binding pocket with the substrate's C17 oriented toward the active site opening. Comparisons suggest that Tyr245 and Phe297 are determinants of KshA1 specificity. The enzyme has a flexible 16-residue mouth loop, which in some structures completely occlude the substrate-binding pocket from the bulk solvent
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
-
the enzyme comprises a Rieske oxygenase, encoded by kshA, and a reductase, encoded by kshB
additional information
the enzyme comprises a Rieske oxygenase, encoded by kshA, and a reductase, encoded by kshB
additional information
the enzyme comprises a Rieske oxygenase, encoded by kshA, and a reductase, encoded by kshB
additional information
KshAB is a two-component Rieske oxygenase comprising an oxygenase (KshA) and a reductase (KshB), structures comparisons of different KSH enzymes. The substrate binding residues within the active site pocket are positioned nearly identically in enzyme KshA1 from Rhodococcus rhodochrous and KshA from Mycobacterium tuberculosis
additional information
-
KshAB is a two-component Rieske oxygenase comprising an oxygenase (KshA) and a reductase (KshB), structures comparisons of different KSH enzymes. The substrate binding residues within the active site pocket are positioned nearly identically in enzyme KshA1 from Rhodococcus rhodochrous and KshA from Mycobacterium tuberculosis
additional information
typical head-to-tail trimer arrangement of KshA enzymes, structure overview
additional information
-
the enzyme comprises a Rieske oxygenase, encoded by kshA, and a reductase, encoded by kshB
additional information
the enzyme comprises a Rieske oxygenase, encoded by kshA, and a reductase, encoded by kshB
additional information
the enzyme comprises a Rieske oxygenase, encoded by kshA, and a reductase, encoded by kshB
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
component KshA, sitting drop vapor diffusion method, using 1.1 M sodium malonate, 0.1 M HEPES, pH 7.0, and 0.5% (v/v) Jeffamine M600
purified recombinant enzyme in complex with substrate androsta-1,4-diene-3,17-dione, using 500 mM NaH2PO4, 125 mM K2HPO4, 4% PEG 1000, 20 mM Tris, 100 mM phosphate-citrate, pH 4.2, X-ray diffraction structure determination and analysis at 2.46-2.52 A resolution
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
Ni2+-NTA agarose column chromatography and Source15Q column chromatography
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
drug development
the enzyme can be a target for inhibition in treatment of tuberculosis
medicine
KSH inhibitory compounds may find application in combatting tuberculosis
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Hu, Y.; Van Der Geize, R.; Besra, G.; Gurcha, S.; Liu, A.; Rohde, M.; Singh, M.; Coates, A.
3-Ketosteroid 9alpha-hydroxylase is an essential factor in the pathogenesis of Mycobacterium tuberculosis
Mol. Microbiol.
75
107-121
2010
Mycobacterium tuberculosis, Mycobacterium tuberculosis H37Rv
Capyk, J.; DAngelo, I.; Strynadka, N.; Eltis, L.
Characterization of 3-ketosteroid 9alpha-hydroxylase, a Rieske oxygenase in the cholesterol degradation pathway of Mycobacterium tuberculosis
J. Biol. Chem.
284
9937-9946
2009
Mycobacterium tuberculosis (P71875), Mycobacterium tuberculosis, Mycobacterium tuberculosis H37Rv (P71875)
Capyk, J.; Casabon, I.; Gruninger, R.; Strynadka, N.; Eltis, L.
Activity of 3-ketosteroid 9alpha-hydroxylase (KshAB) indicates cholesterol side chain and ring degradation occur simultaneously in Mycobacterium tuberculosis
J. Biol. Chem.
286
40717-40724
2011
Mycobacterium tuberculosis, Mycobacterium tuberculosis (P71875), Mycobacterium tuberculosis (P9WJ93), Mycobacterium tuberculosis H37Rv (P71875), Mycobacterium tuberculosis H37Rv (P9WJ93)
Petrusma, M.; van der Geize, R.; Dijkhuizen, L.
3-Ketosteroid 9alpha-hydroxylase enzymes Rieske non-heme monooxygenases essential for bacterial steroid degradation
Antonie van Leeuwenhoek
106
157-172
2014
Mycobacterium tuberculosis (P71875), Mycobacterium tuberculosis H37Rv (P71875), Mycolicibacterium smegmatis, Mycolicibacterium smegmatis mc2 155, Rhodococcus erythropolis, Rhodococcus erythropolis SQ1, Rhodococcus jostii, Rhodococcus jostii (Q0RXD9), Rhodococcus jostii (Q0S812), Rhodococcus rhodochrous, Rhodococcus rhodochrous DSM 43269
Penfield, J.S.; Worrall, L.J.; Strynadka, N.C.; Eltis, L.D.
Substrate specificities and conformational flexibility of 3-ketosteroid 9alpha-hydroxylases
J. Biol. Chem.
289
25523-25536
2014
Mycobacterium tuberculosis (P71875), Mycobacterium tuberculosis, Mycobacterium tuberculosis H37Rv (P71875), Rhodococcus rhodochrous (F1CMX0), Rhodococcus rhodochrous (F1CMY8), Rhodococcus rhodochrous DSM 43269 (F1CMX0), Rhodococcus rhodochrous DSM 43269 (F1CMY8), Rhodococcus rhodochrous DSM 43269
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